MagnetoWiki - User contributions [en]

Electronic Esophagus

2024-09-25T02:22:23Z

MBass: removed details in case my own writing flags me as plagiarizing myself

=Parts=

==Electronics==

Arduino- compatible microcontroller (Teensy-LC)
[https://www.pjrc.com/teensy/teensyLC.html Teensy-LC]

A capacitive touch sensor (Adafruit MPR121)
[https://www.adafruit.com/product/1982 MPR121 Capacitive Touch Sensor]

stepper motor driver (Pololu DRV8825)
[https://www.pololu.com/product/2133/ DRV8825 Stepper Motor Driver]

stepper motor (NEMA-17, 200 steps/rev, 12V, 350mA)
[https://www.adafruit.com/product/324 Stepper motor, NEMA-17, 200 steps/rev, 12V 350mA]

==Hardware==
1 [https://www.adafruit.com/product/1176 Aluminum Flex Shaft Coupler - 5mm to 8mm]

1 [https://www.adafruit.com/product/1181 Linear Ball Bearing - 8mm diameter]

1 M8 x 250mm Fully Threaded Rod, 304 Stainless Steel, Right Hand Threads [https://www.amazon.com/dp/B078H2ZM3C?ref=ppx_yo2_dt_b_product_details&th=1 like this one]

1 Linear Motion Rod 8mmx 250mm [https://www.amazon.com/dp/B08JFZQG2H?ref=ppx_yo2_dt_b_product_details&th=1 like these]

1 M8-1.25 Hex Nut [https://www.amazon.com/Persberg-M8-1-25-Height-Stainless-120153/dp/B089PR8DVP/ref=sr_1_1_sspa?crid=29DW29A9AWQEP&keywords=m8+hex+nut&qid=1642699078&s=industrial&sprefix=m8+he%2Cindustrial%2C87&sr=1-1-spons&psc=1&spLa=ZW5jcnlwdGVkUXVhbGlmaWVyPUEyUEtNOVlXSlkyR00wJmVuY3J5cHRlZElkPUEwMzc5MzU5MjJSWVcyRDg5MDY4TSZlbmNyeXB0ZWRBZElkPUEwNzUzNTQ3M0Q4TEZKWTJZRDdCViZ3aWRnZXROYW1lPXNwX2F0ZiZhY3Rpb249Y2xpY2tSZWRpcmVjdCZkb05vdExvZ0NsaWNrPXRydWU= like these]

==3D Printed Parts==
The 4 parts are currently available as STL and 3MF files.
[https://drive.google.com/drive/folders/1_GCll42MNhbK981q99WKfd3auBnq7ENU?usp=sharing Download here]

==Software==
[https://learn.adafruit.com/adafruit-mpr121-12-key-capacitive-touch-sensor-breakout-tutorial/wiring Adafruit Tutorial]

[https://www.arduino.cc/en/software Arduino IDE]

[https://www.pjrc.com/teensy/teensyduino.html Teensyduino add-on]

[https://circuitjournal.com/arduino-serial-to-spreadsheet ArduSpreadsheet add-on]

=Assembly=

==Electronics==
[https://drive.google.com/drive/folders/1_GCll42MNhbK981q99WKfd3auBnq7ENU Fritzing schematic.]

==3D Printed Parts and Hardware==

===3D Printed Parts===
[https://drive.google.com/drive/folders/1WG1S4pQOFqJO8LHcBtilgX4-BnkKr9EW 3D printed parts] 4 parts ("NemaMount," "MidBody," 20mLbody" and "Plunger").

=Code=
Arduino code is written in C/C++

//include relevant libraries
#include <Wire.h>
#include "Adafruit_MPR121.h"
#include <AccelStepper.h>

//define MPR121 info
#ifndef _BV
#define _BV(bit) (1 << (bit))
#endif
Adafruit_MPR121 cap = Adafruit_MPR121();
uint16_t lasttouched = 0;
uint16_t currtouched = 0;

//define DRV8825 stuff:
#define dirPin 2
#define stepPin 3
#define motorInterfaceType 1

// Create a new instance of the AccelStepper class:
AccelStepper stepper = AccelStepper(motorInterfaceType, stepPin, dirPin);

//setup code here, to run once:
void setup() {
stepper.setMaxSpeed(1000); //set max stepper speed in steps per second
Serial.begin(9600); //start serial monitor
while (!Serial) { // needed to keep from starting too fast!
delay(10);
}
Serial.println("Electronic Esophagus test start");
// Default address is 0x5A, if tied to 3.3V its 0x5B
// If tied to SDA its 0x5C and if SCL then 0x5D
if (!cap.begin(0x5A)) {
Serial.println("MPR121 not found, check wiring?");
while (1);
}
Serial.println("MPR121 found!");
// to change threshold sensitivites (touched, released). Defaults are (12,6). Valuess from 0-255 = less-more sensitive.
cap.setThresholds(24, 12);
}

// main code here, to run repeatedly:
void loop() {
// Get the currently touched pads
currtouched = cap.touched();

for (uint8_t i=0; i<12; i++) {
// it if *is* touched and *wasnt* touched before, alert!
if ((currtouched & _BV(i)) && !(lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" touched");
// set the position to 0:
stepper.setCurrentPosition(0);
// Run the motor forward at 400 steps/second until the motor reaches 12 steps (~0.125 revolutions):
while(stepper.currentPosition() != 12)
{
stepper.setSpeed(400);
stepper.runSpeed();
}
delayMicroseconds(500);
}

// if it *was* touched and now *isnt*, alert!
if (!(currtouched & _BV(i)) && (lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" released");
stepper.setCurrentPosition(0);
stepper.stop(); // Stop as fast as possible: sets new target
stepper.runToPosition(); // Now stopped after quickstop
delayMicroseconds(500);
}
}
// reset our state
lasttouched = currtouched;
// comment out "return" line for detailed data from the sensor!
return;
// debugging info, what
Serial.print("\t\t\t\t\t\t\t\t\t\t\t\t\t 0x"); Serial.println(cap.touched(), HEX);
Serial.print("Filt: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.filteredData(i)); Serial.print("\t");
}
Serial.println();
Serial.print("Base: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.baselineData(i)); Serial.print("\t");
}
Serial.println();
// put a delay so it isn't overwhelming
delay(100);
}

=References=
Dr. D-Flo. (2017, Feb 20). DIY Syringe Pump (Food 3D Printer - Part 1) [Video]. YouTube. [https://www.youtube.com/watch?v=UHa-OKb_CiM&t=212s]

Elizalde G & Sclafani A, Flavor preferences conditioned by intragastric polycose infusions: A detailed analysis using an electronic esophagus preparation, Physiology & Behavior 47: 63-67, 1990. [https://pubmed.ncbi.nlm.nih.gov/2109327/]

Longley M et al. An open source device for operant licking in rats. PeerJ 5:e2981, 2017. [https://peerj.com/articles/2981/]

Wijen B et al. Open-source syringe pump library. PLOS ONE 9(9): e107216, 2014. [https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0107216]

=People=
Marena Bass

[[Category:Flavor Preference]]

=Research Support=
T32 DC000044, National Institutes of Health (NIH/NIDCD) Bass, Marena N
Florida State University (FSU) Chemical Senses Training Program (CTP) Grant Award
Role: Trainee (08/07/2019 - 08/06/2021)

Electronic Esophagus

2024-05-03T13:53:45Z

MBass: added research funding

Selection, construction and programming of the apparatus hardware was modified from open-source syringe pump manuscripts by Longley et al (2017) and Wijen et al (2014). A capacitive touch sensor (Adafruit MPR121) is connected to the rats’ sipper tubes and programmed to detect each lick from the spout with millisecond accuracy. The capacitive touch sensor relays its information to an Arduino- compatible microcontroller (Teensy-LC) which subsequently signals a stepper motor driver (Pololu DRV8825) to turn a stepper motor (NEMA-17, 200 steps/rev, 12V, 350mA), resulting in the release of 4–8 μl liquid per lick from a 20 ml syringe connected to the stepper motor (Figure 4). The syringe pumps liquid into the rat’s chronic gastric catheter at a rate of 1 ml/min over 10 min. This infusion rate is slow enough that the stomach easily accommodates the fill and allows for gastric emptying into the intestine at a normal rate.

Using a protocol adapted from Elizalde & Sclafani (1990), unconditioned stimuli (glucose or saline) will be infused intragastrically as the animals orally sample distinct flavored solutions. With this "Electronic Esophagus" preparation utilized by Sclafani and others, animals form robust and persistent preferences for the flavored solution that is paired with glucose infusions.

=Parts=

==Electronics==
[https://www.pjrc.com/teensy/teensyLC.html Teensy-LC]

[https://www.adafruit.com/product/1982 MPR121 Capacitive Touch Sensor]

[https://www.pololu.com/product/2133/ DRV8825 Stepper Motor Driver]

[https://www.adafruit.com/product/324 Stepper motor, NEMA-17, 200 steps/rev, 12V 350mA]

==Hardware==
1 [https://www.adafruit.com/product/1176 Aluminum Flex Shaft Coupler - 5mm to 8mm]

1 [https://www.adafruit.com/product/1181 Linear Ball Bearing - 8mm diameter]

1 M8 x 250mm Fully Threaded Rod, 304 Stainless Steel, Right Hand Threads [https://www.amazon.com/dp/B078H2ZM3C?ref=ppx_yo2_dt_b_product_details&th=1 like this one]

1 Linear Motion Rod 8mmx 250mm [https://www.amazon.com/dp/B08JFZQG2H?ref=ppx_yo2_dt_b_product_details&th=1 like these]

1 M8-1.25 Hex Nut [https://www.amazon.com/Persberg-M8-1-25-Height-Stainless-120153/dp/B089PR8DVP/ref=sr_1_1_sspa?crid=29DW29A9AWQEP&keywords=m8+hex+nut&qid=1642699078&s=industrial&sprefix=m8+he%2Cindustrial%2C87&sr=1-1-spons&psc=1&spLa=ZW5jcnlwdGVkUXVhbGlmaWVyPUEyUEtNOVlXSlkyR00wJmVuY3J5cHRlZElkPUEwMzc5MzU5MjJSWVcyRDg5MDY4TSZlbmNyeXB0ZWRBZElkPUEwNzUzNTQ3M0Q4TEZKWTJZRDdCViZ3aWRnZXROYW1lPXNwX2F0ZiZhY3Rpb249Y2xpY2tSZWRpcmVjdCZkb05vdExvZ0NsaWNrPXRydWU= like these]

==3D Printed Parts==
The 4 parts are currently available as STL and 3MF files.
[https://drive.google.com/drive/folders/1_GCll42MNhbK981q99WKfd3auBnq7ENU?usp=sharing Download here]

==Software==
[https://learn.adafruit.com/adafruit-mpr121-12-key-capacitive-touch-sensor-breakout-tutorial/wiring Adafruit Tutorial]

[https://www.arduino.cc/en/software Arduino IDE]

[https://www.pjrc.com/teensy/teensyduino.html Teensyduino add-on]

[https://circuitjournal.com/arduino-serial-to-spreadsheet ArduSpreadsheet add-on]

=Assembly=

==Electronics==
The MPR121 capacitive touch sensor can be connected to up to 12 capacitive electrodes (e.g., 12 sipper tubes). Its GND, 3.3V, SCL and SDA pins are connected to the corresponding Teensy-LC pins. The 5V and GND Teensy-LC pins provide the logic power supply to the DRV8825 motor driver, which is connected to a 12V DC power supply with a 100 uF decoupling capacitor to power to the stepper motor. The direction (DIR), step (STP) and enable (ENA) pins of the motor driver receive signals from the Teensy-LC digital pins D0, D1 and D2 respectively. Because the reset (RES) and sleep (SLP) motor driver pins are active low pins, they are connected to the logic power supply to ensure they remain inactive. [https://drive.google.com/drive/folders/1_GCll42MNhbK981q99WKfd3auBnq7ENU Fritzing schematic of electronic esophagus circuit.]

==3D Printed Parts and Hardware==

===3D Printed Parts===
Three of the four [https://drive.google.com/drive/folders/1WG1S4pQOFqJO8LHcBtilgX4-BnkKr9EW 3D printed parts] ("NemaMount," "MidBody" and 20mLbody") are attached together with [https://supergluecorp.com/product/super-glue-tube/ super glue] to form the body of the apparatus.

The "Plunger" will house some of the hardware, including the M8-1.25 Hex Nut and the linear ball bearing.

===Hardware===
The M8-1.25 Hex Nut can be inserted into the hexagonal cutaway in the 3D-printed "Plunger" and subsequently positioned so that the hole in the Hex Nut is in line with the continuous hole of the "Plunger."

The linear ball bearing fits snugly into the larger diameter continuous hole below the hex nut.

=Code=
Arduino code is written in C/C++

//include relevant libraries
#include <Wire.h>
#include "Adafruit_MPR121.h"
#include <AccelStepper.h>

//define MPR121 info
#ifndef _BV
#define _BV(bit) (1 << (bit))
#endif
Adafruit_MPR121 cap = Adafruit_MPR121();
uint16_t lasttouched = 0;
uint16_t currtouched = 0;

//define DRV8825 stuff:
#define dirPin 2
#define stepPin 3
#define motorInterfaceType 1

// Create a new instance of the AccelStepper class:
AccelStepper stepper = AccelStepper(motorInterfaceType, stepPin, dirPin);

//setup code here, to run once:
void setup() {
stepper.setMaxSpeed(1000); //set max stepper speed in steps per second
Serial.begin(9600); //start serial monitor
while (!Serial) { // needed to keep from starting too fast!
delay(10);
}
Serial.println("Electronic Esophagus test start");
// Default address is 0x5A, if tied to 3.3V its 0x5B
// If tied to SDA its 0x5C and if SCL then 0x5D
if (!cap.begin(0x5A)) {
Serial.println("MPR121 not found, check wiring?");
while (1);
}
Serial.println("MPR121 found!");
// to change threshold sensitivites (touched, released). Defaults are (12,6). Valuess from 0-255 = less-more sensitive.
cap.setThresholds(24, 12);
}

// main code here, to run repeatedly:
void loop() {
// Get the currently touched pads
currtouched = cap.touched();

for (uint8_t i=0; i<12; i++) {
// it if *is* touched and *wasnt* touched before, alert!
if ((currtouched & _BV(i)) && !(lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" touched");
// set the position to 0:
stepper.setCurrentPosition(0);
// Run the motor forward at 400 steps/second until the motor reaches 12 steps (~0.125 revolutions):
while(stepper.currentPosition() != 12)
{
stepper.setSpeed(400);
stepper.runSpeed();
}
delayMicroseconds(500);
}

// if it *was* touched and now *isnt*, alert!
if (!(currtouched & _BV(i)) && (lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" released");
stepper.setCurrentPosition(0);
stepper.stop(); // Stop as fast as possible: sets new target
stepper.runToPosition(); // Now stopped after quickstop
delayMicroseconds(500);
}
}
// reset our state
lasttouched = currtouched;
// comment out "return" line for detailed data from the sensor!
return;
// debugging info, what
Serial.print("\t\t\t\t\t\t\t\t\t\t\t\t\t 0x"); Serial.println(cap.touched(), HEX);
Serial.print("Filt: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.filteredData(i)); Serial.print("\t");
}
Serial.println();
Serial.print("Base: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.baselineData(i)); Serial.print("\t");
}
Serial.println();
// put a delay so it isn't overwhelming
delay(100);
}

=References=
Dr. D-Flo. (2017, Feb 20). DIY Syringe Pump (Food 3D Printer - Part 1) [Video]. YouTube. [https://www.youtube.com/watch?v=UHa-OKb_CiM&t=212s]

Elizalde G & Sclafani A, Flavor preferences conditioned by intragastric polycose infusions: A detailed analysis using an electronic esophagus preparation, Physiology & Behavior 47: 63-67, 1990. [https://pubmed.ncbi.nlm.nih.gov/2109327/]

Longley M et al. An open source device for operant licking in rats. PeerJ 5:e2981, 2017. [https://peerj.com/articles/2981/]

Wijen B et al. Open-source syringe pump library. PLOS ONE 9(9): e107216, 2014. [https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0107216]

=People=
Marena Bass

[[Category:Flavor Preference]]

=Research Support=
T32 DC000044, National Institutes of Health (NIH/NIDCD) Bass, Marena N
Florida State University (FSU) Chemical Senses Training Program (CTP) Grant Award
Role: Trainee (08/07/2019 - 08/06/2021)

MBJ Selenate Mass Spec

2023-09-25T14:53:22Z

MBass: /* Agenda */

=Background=

[[Selenium|Sodium Selenate]] (Na2Se04) activates PP2A (1).

[[CTA and Selenate]] experiments show that male Sprague Dawley rats given systemic injections of 0.5mg/kg sodium selenate two hours before taste aversion conditioning form attenuated taste aversions that extinguish rapidly. Selenate also decreases LiCl-induced phospho-MAP kinase in the NTS and lateral PBN.

This experiment will use LCMS to identify differences in protein phosphorylation in the brain two hours after rats receive 0.5mg/kg selenate vs vehicle.

=Agenda=

'''07/15/2023 - Tissue Collection:
'''
10 rats (5 males, 5 females) were overdosed with euthasol and rapidly decapitated two hours after receiving 1ml/kg i.p. injections of 0.5mg/kg sodium selenate or vehicle (0.15M NaCl). Brains were dissected and washed in ice-cold PP2A storage buffer (50mM Tris-HCl, pH 7.4, 0.1mM EDTA, 0.1mM EGTA, 1mM DTT, 250mM Sucrose, 0.1%(v/v) N-PER detergent, 1 Pierce protease inhibitor mini tablet per 10mL solution), and frozen at -80*C.

{| class="wikitable"
|+ Experiment schedule
|-
! Subject !! Sex !! Treatment !! Weight (g) !! Injection vol (ml) !! Injection time (PM) !! Time of brain dissection (PM)
|-
| MBJ01 || M || NaCl || 371 || 0.37 || 2:17 || 4:28
|-
| MBJ02 ||F || Sel || 257 || 0.26 || 2:26 || 4:34
|-
| MBJ03 || M || Sel || 348 || 0.35 || 2:33 ||
|-
| MBJ04 || F || NaCl || 250 || 0.25 || 2:40 ||
|-
| MBJ05 || M || NaCl || 346 || 0.35 || 2:46 ||5:00
|-
| MBJ06 || F || Sel || 247 || 0.25 || 2:53 ||
|-
| MBJ07 || M || Sel || 324 || 0.33 || 2:59 ||
|-
| MBJ08 || F || NaCl || 245 || 0.25 || 3:04 ||
|-
| MBJ09 || M || NaCl || 335 || 0.34 || 3:10 ||
|-
| MBJ10 || F || Sel || 263 || 0.26 || 3:16 || 5:33
|}

'''7/28/2023 - Protein Extraction:'''
For each frozen tissue sample (MBJ01-10), Amygdalar lobe (A) and caudal brainstem (B) regions were dissected and homogenized in PP2A storage buffer (50mM Tris-HCl, pH 7.4, 0.1mM EDTA, 0.1mM EGTA, 1mM DTT, 250mM Sucrose, 0.1%(v/v) N-PER detergent, 1 Pierce protease inhibitor mini tablet per 10mL solution)
* 1:3 ratio of brain tissue:buffer
* Dounce homogenization: 50-60 strokes on ice
* The homogenate was centrifuged at 100,000 x g, 4°C for 1 hour
* The supernatants were collected and stored at 4°C overnight

'''7/29/2023 - Sample clean-up: detergent and salt removal
'''
The supernatants were buffer exchanged into TEE buffer (50mM Tris-HCl pH 7.4, 0.1mM EDTA, 0.1mM EGTA) using PD-10 desalting columns (Cytiva #17085101)
The samples were then aliquoted and frozen at -20°C

'''7/31/2023 - Bradford and BCA assays to determine protein concentration
'''
Both Bradford and BCA Assays were conducted to determine the protein concentration. Results generated from both assays were nearly identical +/- 0.02ug/ml
The protein concentrations estimated from the BCA assay were used for all subsequent calculations

'''9/18/2023 - Reduction/Alkylation/Digestion
'''

'''''Reduction'''''
* Add 60 uL of each sample to a premade tube of 0.04g urea → [final] = 8M urea
* Vortex until dissolved
* Place in 37 deg C water bath for 2 mins
* Add 5 uL TCEP bond breaker to each sample
* pipette/mix 5x
* Pulse 5 sec
* Incubate in 37 deg C water bath for 60 mins

'''''Alkylation'''
''
* Add 10 uL of 20mM iodoacetamide stock solution to each sample
* Vortex 2 seconds
* Pulse 5 seconds
* Incubate in 37 deg C water bath for 20 mins
* Add 400 uL water to each sample → ~500 uL final volume

'''''Digestion
'''''
* Reconstitute 20ug Trypsin (Promega #V5111) by adding 200 ul of supplied buffer → [final] = 1ug/10ul trypsin
* Add enough trypsin for a 1:20 ratio of trypsin:protein
* Incubate all samples in 37°C water bath overnight
Next day:
* Remove samples from water bath
* Quench with 2.6 uL of 10% Formic Acid
* Store at -20°C

'''Final sample
'''
* Total number of samples = 17
* 09A, 09B, and 10B were excluded from this batch due to possible cross-contamination
* Buffer (before reduction/alkylation/digestion) = 50mM Tris-HCl pH 7.4, 0.1mM EDTA, 0.1mM EGTA
* Final volume = ~500uL per sample

'''09/21/2023 - Samples delivered to Translational Lab for Mass Spec analysis
'''
* Samples passed through S-trap column for final clean-up (removal of urea and salts)

=References=
Corcoran NM, Martin D, Hutter-Paier B, Windisch M, Nguyen T, Nheu L, Sundstrom LE, Costello AJ, Hovens CM. Sodium selenate specifically activates PP2A phosphatase, dephosphorylates tau and reverses memory deficits in an Alzheimer’s disease model. Journal of Clinical Neuroscience 17: 1025- 1033, 2010.[https://pubmed.ncbi.nlm.nih.gov/20537899/]

=People=
Marena Bass

[[Category:CTA Experiments]]
[[Category:Taste_Aversion]]

MBJ Selenate Mass Spec

2023-09-25T14:45:17Z

MBass: /* Agenda */

=Background=

[[Selenium|Sodium Selenate]] (Na2Se04) activates PP2A (1).

[[CTA and Selenate]] experiments show that male Sprague Dawley rats given systemic injections of 0.5mg/kg sodium selenate two hours before taste aversion conditioning form attenuated taste aversions that extinguish rapidly. Selenate also decreases LiCl-induced phospho-MAP kinase in the NTS and lateral PBN.

This experiment will use LCMS to identify differences in protein phosphorylation in the brain two hours after rats receive 0.5mg/kg selenate vs vehicle.

=Agenda=

'''07/15/2023 - Tissue Collection:
'''
10 rats (5 males, 5 females) were overdosed with euthasol and rapidly decapitated two hours after receiving 1ml/kg i.p. injections of 0.5mg/kg sodium selenate or vehicle (0.15M NaCl). Brains were dissected and washed in ice-cold PP2A storage buffer (50mM Tris-HCl, pH 7.4, 0.1mM EDTA, 0.1mM EGTA, 1mM DTT, 250mM Sucrose, 0.1%(v/v) N-PER detergent, 1 Pierce protease inhibitor mini tablet per 10mL solution), and frozen at -80*C.

{| class="wikitable"
|+ Experiment schedule
|-
! Subject !! Sex !! Treatment !! Weight !! Injection vol (ml) !! Injection time (PM) !! Time of brain dissection (PM)
|-
| MBJ01 || M || NaCl || 371 || 0.37 || 2:17 || 4:28
|-
| MBJ02 ||F || Sel || 257 || 0.26 || 2:26 || 4:34
|-
| MBJ03 || M || Sel || 348 || 0.35 || 2:33 ||
|-
| MBJ04 || F || NaCl || 250 || 0.25 || 2:40 ||
|-
| MBJ05 || M || NaCl || 346 || 0.35 || 2:46 ||5:00
|-
| MBJ06 || F || Sel || 247 || 0.25 || 2:53 ||
|-
| MBJ07 || M || Sel || 324 || 0.33 || 2:59 ||
|-
| MBJ08 || F || NaCl || 245 || 0.25 || 3:04 ||
|-
| MBJ09 || M || NaCl || 335 || 0.34 || 3:10 ||
|-
| MBJ10 || F || Sel || 263 || 0.26 || 3:16 || 5:33
|}

'''7/28/2023 - Protein Extraction:'''
For each frozen tissue sample (MBJ01-10), Amygdalar lobe (A) and caudal brainstem (B) regions were dissected and homogenized in PP2A storage buffer (50mM Tris-HCl, pH 7.4, 0.1mM EDTA, 0.1mM EGTA, 1mM DTT, 250mM Sucrose, 0.1%(v/v) N-PER detergent, 1 Pierce protease inhibitor mini tablet per 10mL solution)
* 1:3 ratio of brain tissue:buffer
* Dounce homogenization: 50-60 strokes on ice
* The homogenate was centrifuged at 100,000 x g, 4°C for 1 hour
* The supernatants were collected and stored at 4°C overnight

'''7/29/2023 - Sample clean-up: detergent and salt removal
'''
The supernatants were buffer exchanged into TEE buffer (50mM Tris-HCl pH 7.4, 0.1mM EDTA, 0.1mM EGTA) using PD-10 desalting columns (Cytiva #17085101)
The samples were then aliquoted and frozen at -20°C

'''7/31/2023 - Bradford and BCA assays to determine protein concentration
'''
Both Bradford and BCA Assays were conducted to determine the protein concentration. Results generated from both assays were nearly identical +/- 0.02ug/ml
The protein concentrations estimated from the BCA assay were used for all subsequent calculations

'''9/18/2023 - Reduction/Alkylation/Digestion
'''

'''''Reduction'''''
* Add 60 uL of each sample to a premade tube of 0.04g urea → [final] = 8M urea
* Vortex until dissolved
* Place in 37 deg C water bath for 2 mins
* Add 5 uL TCEP bond breaker to each sample
* pipette/mix 5x
* Pulse 5 sec
* Incubate in 37 deg C water bath for 60 mins

'''''Alkylation'''
''
* Add 10 uL of 20mM iodoacetamide stock solution to each sample
* Vortex 2 seconds
* Pulse 5 seconds
* Incubate in 37 deg C water bath for 20 mins
* Add 400 uL water to each sample → ~500 uL final volume

'''''Digestion
'''''
* Reconstitute 20ug Trypsin (Promega #V5111) by adding 200 ul of supplied buffer → [final] = 1ug/10ul trypsin
* Add enough trypsin for a 1:20 ratio of trypsin:protein
* Incubate all samples in 37°C water bath overnight
Next day:
* Remove samples from water bath
* Quench with 2.6 uL of 10% Formic Acid
* Store at -20°C

'''Final sample
'''
* Total number of samples = 17
* 09A, 09B, and 10B were excluded from this batch due to possible cross-contamination
* Buffer (before reduction/alkylation/digestion) = 50mM Tris-HCl pH 7.4, 0.1mM EDTA, 0.1mM EGTA
* Final volume = ~500uL per sample

'''09/21/2023 - Samples delivered to Translational Lab for Mass Spec analysis
'''
* Samples passed through S-trap column for final clean-up (removal of urea and salts)

=References=
Corcoran NM, Martin D, Hutter-Paier B, Windisch M, Nguyen T, Nheu L, Sundstrom LE, Costello AJ, Hovens CM. Sodium selenate specifically activates PP2A phosphatase, dephosphorylates tau and reverses memory deficits in an Alzheimer’s disease model. Journal of Clinical Neuroscience 17: 1025- 1033, 2010.[https://pubmed.ncbi.nlm.nih.gov/20537899/]

=People=
Marena Bass

[[Category:CTA Experiments]]
[[Category:Taste_Aversion]]

MBJ Selenate Mass Spec

2023-09-25T14:40:57Z

MBass: /* Agenda */

=Background=

[[Selenium|Sodium Selenate]] (Na2Se04) activates PP2A (1).

[[CTA and Selenate]] experiments show that male Sprague Dawley rats given systemic injections of 0.5mg/kg sodium selenate two hours before taste aversion conditioning form attenuated taste aversions that extinguish rapidly. Selenate also decreases LiCl-induced phospho-MAP kinase in the NTS and lateral PBN.

This experiment will use LCMS to identify differences in protein phosphorylation in the brain two hours after rats receive 0.5mg/kg selenate vs vehicle.

=Agenda=

'''07/15/2023 - Tissue Collection:
'''
10 rats (5 males, 5 females) were overdosed with euthasol and rapidly decapitated two hours after receiving 1ml/kg i.p. injections of 0.5mg/kg sodium selenate or vehicle (0.15M NaCl). Brains were dissected and washed in ice-cold PP2A storage buffer (50mM Tris-HCl, pH 7.4, 0.1mM EDTA, 0.1mM EGTA, 1mM DTT, 250mM Sucrose, 0.1%(v/v) N-PER detergent, 1 Pierce protease inhibitor mini tablet per 10mL solution), and frozen at -80*C.

{| class="wikitable"
|+ Experiment schedule
|-
! Subject !! Sex !! Treatment !! Weight !! Injection vol (ml) !! Injection time (PM) !! Time of brain dissection (PM)
|-
| MBJ01 || M || NaCl || 371 || 0.37 || 2:17 || 4:28
|-
| MBJ02 ||F || Sel || 257 || 0.26 || 2:26 || 4:34
|-
| MBJ03 || M || Sel || 348 || 0.35 || 2:33 ||
|-
| MBJ04 || F || NaCl || 250 || 0.25 || 2:40 ||
|-
| MBJ05 || M || NaCl || 346 || 0.35 || 2:46 ||5:00
|-
| MBJ06 || F || Sel || 247 || 0.25 || 2:53 ||
|-
| MBJ07 || M || Sel || 324 || 0.33 || 2:59 ||
|-
| MBJ08 || F || NaCl || 245 || 0.25 || 3:04 ||
|-
| MBJ09 || M || NaCl || 335 || 0.34 || 3:10 ||
|-
| MBJ10 || F || Sel || 263 || 0.26 || 3:16 || 5:33
|}

'''7/28/2023 - Protein Extraction:'''
For each frozen tissue sample (MBJ01-10), Amygdalar lobe (A) and caudal brainstem (B) regions were dissected and homogenized in PP2A storage buffer (50mM Tris-HCl, pH 7.4, 0.1mM EDTA, 0.1mM EGTA, 1mM DTT, 250mM Sucrose, 0.1%(v/v) N-PER detergent, 1 Pierce protease inhibitor mini tablet per 10mL solution)
* 1:3 ratio of brain tissue:buffer
* Dounce homogenization: 50-60 strokes on ice
* The homogenate was centrifuged at 100,000 x g, 4°C for 1 hour
* The supernatants were collected and stored at 4°C overnight

'''7/29/2023 - Sample clean-up: detergent and salt removal
'''
The supernatants were buffer exchanged into TEE buffer (50mM Tris-HCl pH 7.4, 0.1mM EDTA, 0.1mM EGTA) using PD-10 desalting columns (Cytiva #17085101)
The samples were then aliquoted and frozen at -20°C

'''7/31/2023 - Bradford and BCA assays to determine protein concentration
'''
Both Bradford and BCA Assays were conducted to determine the protein concentration. Results generated from both assays were nearly identical +/- 0.02ug/ml
The protein concentrations estimated from the BCA assay were used for all subsequent calculations

'''9/18/2023 - Reduction/Alkylation/Digestion
'''

'''''Reduction'''''
* Add 60 uL of each sample to a premade tube of 0.04g urea → [final] = 8M urea
* Vortex until dissolved
* Place in 37 deg C water bath for 2 mins
* Add 5 uL TCEP bond breaker to each sample
* pipette/mix 5x
* Pulse 5 sec
* Incubate in 37 deg C water bath for 60 mins

'''''Alkylation'''
''
* Add 10 uL of 20mM iodoacetamide stock solution to each sample
* Vortex 2 seconds
* Pulse 5 seconds
* Incubate in 37 deg C water bath for 20 mins
* Add 400 uL water to each sample → ~500 uL final volume

'''''Digestion
'''''
* Reconstitute 20ug Trypsin (Promega #V5111) by adding 200 ul of supplied buffer → [final] = 1ug/10ul trypsin
* Add enough trypsin for a 1:20 ratio of trypsin:protein
* Incubate all samples in 37°C water bath overnight
Next day:
* Remove samples from water bath
* Quench with 2.6 uL of 10% Formic Acid
* Store at -20°C

'''Final sample
'''
* Total number of samples = 17
* 09A, 09B, and 10B were excluded from this batch due to possible cross-contamination
* Buffer (before reduction/alkylation/digestion) = 50mM Tris-HCl pH 7.4, 0.1mM EDTA, 0.1mM EGTA
* Final volume = ~500uL per sample

=References=
Corcoran NM, Martin D, Hutter-Paier B, Windisch M, Nguyen T, Nheu L, Sundstrom LE, Costello AJ, Hovens CM. Sodium selenate specifically activates PP2A phosphatase, dephosphorylates tau and reverses memory deficits in an Alzheimer’s disease model. Journal of Clinical Neuroscience 17: 1025- 1033, 2010.[https://pubmed.ncbi.nlm.nih.gov/20537899/]

=People=
Marena Bass

[[Category:CTA Experiments]]
[[Category:Taste_Aversion]]

MBJ Selenate Mass Spec

2023-09-25T14:39:43Z

MBass: /* Agenda */ formatting again

=Background=

[[Selenium|Sodium Selenate]] (Na2Se04) activates PP2A (1).

[[CTA and Selenate]] experiments show that male Sprague Dawley rats given systemic injections of 0.5mg/kg sodium selenate two hours before taste aversion conditioning form attenuated taste aversions that extinguish rapidly. Selenate also decreases LiCl-induced phospho-MAP kinase in the NTS and lateral PBN.

This experiment will use LCMS to identify differences in protein phosphorylation in the brain two hours after rats receive 0.5mg/kg selenate vs vehicle.

=Agenda=

'''07/15/2023 - Tissue Collection:
'''
10 rats (5 males, 5 females) were overdosed with euthasol and rapidly decapitated two hours after receiving 1ml/kg i.p. injections of 0.5mg/kg sodium selenate or vehicle (0.15M NaCl). Brains were dissected and washed in ice-cold PP2A storage buffer (50mM Tris-HCl, pH 7.4, 0.1mM EDTA, 0.1mM EGTA, 1mM DTT, 250mM Sucrose, 0.1%(v/v) N-PER detergent, 1 Pierce protease inhibitor mini tablet per 10mL solution), and frozen at -80*C.

{| class="wikitable"
|+ Experiment schedule
|-
! Subject !! Sex !! Treatment !! Weight !! Injection vol (ml) !! Injection time (PM) !! Time of brain dissection (PM)
|-
| MBJ01 || M || NaCl || 371 || 0.37 || 2:17 || 4:28
|-
| MBJ02 ||F || Sel || 257 || 0.26 || 2:26 || 4:34
|-
| MBJ03 || M || Sel || 348 || 0.35 || 2:33 ||
|-
| MBJ04 || F || NaCl || 250 || 0.25 || 2:40 ||
|-
| MBJ05 || M || NaCl || 346 || 0.35 || 2:46 ||5:00
|-
| MBJ06 || F || Sel || 247 || 0.25 || 2:53 ||
|-
| MBJ07 || M || Sel || 324 || 0.33 || 2:59 ||
|-
| MBJ08 || F || NaCl || 245 || 0.25 || 3:04 ||
|-
| MBJ09 || M || NaCl || 335 || 0.34 || 3:10 ||
|-
| MBJ10 || F || Sel || 263 || 0.26 || 3:16 || 5:33
|}

'''7/28/2023 - Protein Extraction:'''
For each frozen tissue sample (MBJ01-10), Amygdalar lobe (A) and caudal brainstem (B) regions were dissected and homogenized in PP2A storage buffer (50mM Tris-HCl, pH 7.4, 0.1mM EDTA, 0.1mM EGTA, 1mM DTT, 250mM Sucrose, 0.1%(v/v) N-PER detergent, 1 Pierce protease inhibitor mini tablet per 10mL solution)
1:3 ratio of brain tissue:buffer
* Dounce homogenization: 50-60 strokes on ice
* The homogenate was centrifuged at 100,000 x g, 4°C for 1 hour
* The supernatants were collected and stored at 4°C overnight

'''7/29/2023 - Sample clean-up: detergent and salt removal
'''
The supernatants were buffer exchanged into TEE buffer (50mM Tris-HCl pH 7.4, 0.1mM EDTA, 0.1mM EGTA) using PD-10 desalting columns (Cytiva #17085101)
The samples were then aliquoted and frozen at -20°C

'''7/31/2023 - Bradford and BCA assays to determine protein concentration
'''
Both Bradford and BCA Assays were conducted to determine the protein concentration. Results generated from both assays were nearly identical +/- 0.02ug/ml
The protein concentrations estimated from the BCA assay were used for all subsequent calculations

'''9/18/2023 - Reduction/Alkylation/Digestion
'''

'''''Reduction'''''
* Add all 60 uL of each sample to a premade tube of 0.04g urea → [final] = 8M urea
* Vortex until dissolved
* Place in 37 deg C water bath for 2 mins
* Add 5 uL TCEP bond breaker to each sample
* pipette/mix 5x
* Pulse 5 sec
* Incubate in 37 deg C water bath for 60 mins

'''''Alkylation'''
''
* Add 10 uL of 20mM iodoacetamide stock solution to each sample
* Vortex 2 seconds
* Pulse 5 seconds
* Incubate in 37 deg C water bath for 20 mins
* Add 400 uL water to each sample → ~500 uL final volume

'''''Digestion
'''''
* Reconstitute 20ug Trypsin (Promega #V5111) by adding 200 ul of supplied buffer → [final] = 1ug/10ul trypsin
* Add enough trypsin for a 1:20 ratio of trypsin:protein
* Incubate all samples in 37°C water bath overnight
Next day:
* Remove samples from water bath
* Quench with 2.6 uL of 10% Formic Acid
* Store at -20°C

'''Final sample
'''
* Total number of samples = 17
* 09A, 09B, and 10B were excluded from this batch due to possible cross-contamination
* Buffer (before reduction/alkylation/digestion) = 50mM Tris-HCl pH 7.4, 0.1mM EDTA, 0.1mM EGTA
* Final volume = ~500uL per sample

=References=
Corcoran NM, Martin D, Hutter-Paier B, Windisch M, Nguyen T, Nheu L, Sundstrom LE, Costello AJ, Hovens CM. Sodium selenate specifically activates PP2A phosphatase, dephosphorylates tau and reverses memory deficits in an Alzheimer’s disease model. Journal of Clinical Neuroscience 17: 1025- 1033, 2010.[https://pubmed.ncbi.nlm.nih.gov/20537899/]

=People=
Marena Bass

[[Category:CTA Experiments]]
[[Category:Taste_Aversion]]

MBJ Selenate Mass Spec

2023-09-25T14:37:47Z

MBass: /* Agenda */ formatting

=Background=

[[Selenium|Sodium Selenate]] (Na2Se04) activates PP2A (1).

[[CTA and Selenate]] experiments show that male Sprague Dawley rats given systemic injections of 0.5mg/kg sodium selenate two hours before taste aversion conditioning form attenuated taste aversions that extinguish rapidly. Selenate also decreases LiCl-induced phospho-MAP kinase in the NTS and lateral PBN.

This experiment will use LCMS to identify differences in protein phosphorylation in the brain two hours after rats receive 0.5mg/kg selenate vs vehicle.

=Agenda=

'''07/15/2023 - Tissue Collection:
'''
10 rats (5 males, 5 females) were overdosed with euthasol and rapidly decapitated two hours after receiving 1ml/kg i.p. injections of 0.5mg/kg sodium selenate or vehicle (0.15M NaCl). Brains were dissected and washed in ice-cold PP2A storage buffer (50mM Tris-HCl, pH 7.4, 0.1mM EDTA, 0.1mM EGTA, 1mM DTT, 250mM Sucrose, 0.1%(v/v) N-PER detergent, 1 Pierce protease inhibitor mini tablet per 10mL solution), and frozen at -80*C.

{| class="wikitable"
|+ Experiment schedule
|-
! Subject !! Sex !! Treatment !! Weight !! Injection vol (ml) !! Injection time (PM) !! Time of brain dissection (PM)
|-
| MBJ01 || M || NaCl || 371 || 0.37 || 2:17 || 4:28
|-
| MBJ02 ||F || Sel || 257 || 0.26 || 2:26 || 4:34
|-
| MBJ03 || M || Sel || 348 || 0.35 || 2:33 ||
|-
| MBJ04 || F || NaCl || 250 || 0.25 || 2:40 ||
|-
| MBJ05 || M || NaCl || 346 || 0.35 || 2:46 ||5:00
|-
| MBJ06 || F || Sel || 247 || 0.25 || 2:53 ||
|-
| MBJ07 || M || Sel || 324 || 0.33 || 2:59 ||
|-
| MBJ08 || F || NaCl || 245 || 0.25 || 3:04 ||
|-
| MBJ09 || M || NaCl || 335 || 0.34 || 3:10 ||
|-
| MBJ10 || F || Sel || 263 || 0.26 || 3:16 || 5:33
|}

'''7/28/2023 - Protein Extraction:'''
For each frozen tissue sample (MBJ01-10), Amygdalar lobe (A) and caudal brainstem (B) regions were dissected and homogenized in PP2A storage buffer (50mM Tris-HCl, pH 7.4, 0.1mM EDTA, 0.1mM EGTA, 1mM DTT, 250mM Sucrose, 0.1%(v/v) N-PER detergent, 1 Pierce protease inhibitor mini tablet per 10mL solution)
1:3 ratio of brain tissue:buffer
Dounce homogenization: 50-60 strokes on ice
The homogenate was centrifuged at 100,000 x g, 4°C for 1 hour
The supernatants were collected and stored at 4°C overnight

'''7/29/2023 - Sample clean-up: detergent and salt removal
'''
The supernatants were buffer exchanged into TEE buffer (50mM Tris-HCl pH 7.4, 0.1mM EDTA, 0.1mM EGTA) using PD-10 desalting columns (Cytiva #17085101)
The samples were then aliquoted and frozen at -20°C

'''7/31/2023 - Bradford and BCA assays to determine protein concentration
'''
Both Bradford and BCA Assays were conducted to determine the protein concentration. Results generated from both assays were nearly identical +/- 0.02ug/ml
The protein concentrations estimated from the BCA assay were used for all subsequent calculations

'''9/18/2023 - Reduction/Alkylation/Digestion
'''

'''''Reduction'''''
* Add all 60 uL of each sample (see table 1 below) to a premade tube of 0.04g urea → [final] = 8M urea
* Vortex until dissolved
* Place in 37 deg C water bath for 2 mins
* Add 5 uL TCEP bond breaker to each sample
* pipette/mix 5x
* Pulse 5 sec
* Incubate in 37 deg C water bath for 60 mins

'''''Alkylation'''
''
* Add 10 uL of 20mM iodoacetamide stock solution to each sample
* Vortex 2 seconds
* Pulse 5 seconds
* Incubate in 37 deg C water bath for 20 mins
* Add 400 uL water to each sample → ~500 uL final volume

'''''Digestion
'''''
* Reconstitute 20ug Trypsin (Promega #V5111) by adding 200 ul of supplied buffer → [final] = 1ug/10ul trypsin
* Add enough trypsin for a 1:20 ratio of trypsin:protein
* Incubate all samples in 37°C water bath overnight
Next day:
* Remove samples from water bath
* Quench with 2.6 uL of 10% Formic Acid
* Store at -20°C

'''Final sample
'''
Total number of samples = 17
09A, 09B, and 10B were excluded from this batch due to possible cross-contamination
Buffer (before reduction/alkylation/digestion) = 50mM Tris-HCl pH 7.4, 0.1mM EDTA, 0.1mM EGTA
Final volume = ~500uL per sample

=References=
Corcoran NM, Martin D, Hutter-Paier B, Windisch M, Nguyen T, Nheu L, Sundstrom LE, Costello AJ, Hovens CM. Sodium selenate specifically activates PP2A phosphatase, dephosphorylates tau and reverses memory deficits in an Alzheimer’s disease model. Journal of Clinical Neuroscience 17: 1025- 1033, 2010.[https://pubmed.ncbi.nlm.nih.gov/20537899/]

=People=
Marena Bass

[[Category:CTA Experiments]]
[[Category:Taste_Aversion]]

MBJ Selenate Mass Spec

2023-09-25T14:34:21Z

MBass: /* Agenda */

=Background=

[[Selenium|Sodium Selenate]] (Na2Se04) activates PP2A (1).

[[CTA and Selenate]] experiments show that male Sprague Dawley rats given systemic injections of 0.5mg/kg sodium selenate two hours before taste aversion conditioning form attenuated taste aversions that extinguish rapidly. Selenate also decreases LiCl-induced phospho-MAP kinase in the NTS and lateral PBN.

This experiment will use LCMS to identify differences in protein phosphorylation in the brain two hours after rats receive 0.5mg/kg selenate vs vehicle.

=Agenda=

'''07/15/2023 - Tissue Collection:
'''
10 rats (5 males, 5 females) were overdosed with euthasol and rapidly decapitated two hours after receiving 1ml/kg i.p. injections of 0.5mg/kg sodium selenate or vehicle (0.15M NaCl). Brains were dissected and washed in ice-cold PP2A storage buffer (50mM Tris-HCl, pH 7.4, 0.1mM EDTA, 0.1mM EGTA, 1mM DTT, 250mM Sucrose, 0.1%(v/v) N-PER detergent, 1 Pierce protease inhibitor mini tablet per 10mL solution), and frozen at -80*C.

{| class="wikitable"
|+ Experiment schedule
|-
! Subject !! Sex !! Treatment !! Weight !! Injection vol (ml) !! Injection time (PM) !! Time of brain dissection (PM)
|-
| MBJ01 || M || NaCl || 371 || 0.37 || 2:17 || 4:28
|-
| MBJ02 ||F || Sel || 257 || 0.26 || 2:26 || 4:34
|-
| MBJ03 || M || Sel || 348 || 0.35 || 2:33 ||
|-
| MBJ04 || F || NaCl || 250 || 0.25 || 2:40 ||
|-
| MBJ05 || M || NaCl || 346 || 0.35 || 2:46 ||5:00
|-
| MBJ06 || F || Sel || 247 || 0.25 || 2:53 ||
|-
| MBJ07 || M || Sel || 324 || 0.33 || 2:59 ||
|-
| MBJ08 || F || NaCl || 245 || 0.25 || 3:04 ||
|-
| MBJ09 || M || NaCl || 335 || 0.34 || 3:10 ||
|-
| MBJ10 || F || Sel || 263 || 0.26 || 3:16 || 5:33
|}

'''7/28/2023 - Protein Extraction:'''
For each frozen tissue sample (MBJ01-10), Amygdalar lobe (A) and caudal brainstem (B) regions were dissected and homogenized in PP2A storage buffer (50mM Tris-HCl, pH 7.4, 0.1mM EDTA, 0.1mM EGTA, 1mM DTT, 250mM Sucrose, 0.1%(v/v) N-PER detergent, 1 Pierce protease inhibitor mini tablet per 10mL solution)
1:3 ratio of brain tissue:buffer
Dounce homogenization: 50-60 strokes on ice
The homogenate was centrifuged at 100,000 x g, 4°C for 1 hour
The supernatants were collected and stored at 4°C overnight

'''Sample clean-up: detergent and salt removal (7/29/2023)
'''
The supernatants were buffer exchanged into TEE buffer (50mM Tris-HCl pH 7.4, 0.1mM EDTA, 0.1mM EGTA) using PD-10 desalting columns (Cytiva #17085101)
The samples were then aliquoted and frozen at -20°C

'''Bradford and BCA assays to determine protein concentration (7/31/2023)
'''
Both Bradford and BCA Assays were conducted to determine the protein concentration. Results generated from both assays were nearly identical +/- 0.02ug/ml
The protein concentrations estimated from the BCA assay were used for all subsequent calculations

'''Reduction/Alkylation/Digestion (9/18/2023)
'''

Reduction
Add all 60 uL of each sample (see table 1 below) to a premade tube of 0.04g urea → [final] = 8M urea
Vortex until dissolved
Place in 37 deg C water bath for 2 mins

Add 5 uL TCEP bond breaker to each sample
pipette/mix 5x
Pulse 5 sec
Incubate in 37 deg C water bath for 60 mins

Alkylation
Add 10 uL of 20mM iodoacetamide stock solution to each sample
Vortex 2 seconds
Pulse 5 seconds
Incubate in 37 deg C water bath for 20 mins
Add 400 uL water to each sample → ~500 uL final volume

Digestion
Reconstitute 20ug Trypsin (Promega #V5111) by adding 200 ul of supplied buffer → [final] = 1ug/10ul trypsin
Add enough trypsin for a 1:20 ratio of trypsin:protein (see table 1 below)
Incubate all samples in 37°C water bath overnight
Next day:
Remove samples from water bath
Quench with 2.6 uL of 10% Formic Acid
Store at -20°C

'''Final sample
'''
Total number of samples = 17
09A, 09B, and 10B were excluded from this batch due to possible cross-contamination
Buffer (before reduction/alkylation/digestion) = 50mM Tris-HCl pH 7.4, 0.1mM EDTA, 0.1mM EGTA
Final volume = ~500uL per sample

=References=
Corcoran NM, Martin D, Hutter-Paier B, Windisch M, Nguyen T, Nheu L, Sundstrom LE, Costello AJ, Hovens CM. Sodium selenate specifically activates PP2A phosphatase, dephosphorylates tau and reverses memory deficits in an Alzheimer’s disease model. Journal of Clinical Neuroscience 17: 1025- 1033, 2010.[https://pubmed.ncbi.nlm.nih.gov/20537899/]

=People=
Marena Bass

[[Category:CTA Experiments]]
[[Category:Taste_Aversion]]

MBJ Selenate Mass Spec

2023-07-16T16:59:01Z

MBass: added reference, people, category sections

MBJ Selenate Mass Spec

2023-07-16T16:42:55Z

MBass: /* Agenda */ typo

MBJ Selenate Mass Spec

2023-07-16T16:42:07Z

MBass: Created page with "=Background= Sodium Selenate (Na2Se04) activates PP2A (1). CTA and Selenate experiments show that male Sprague Dawley rats given systemic injections of 0.5m..."

MBG Selenate Lying on Belly

2023-07-06T13:21:26Z

MBass: edited organization of background and agenda

Selenate modulation of LiCl-induced Lying-on-Belly

=Background=

[[Selenium|Sodium Selenate]] (Na2Se04) activates PP2A (1).

[[CTA and Selenate]] experiments show that male Sprague Dawley rats given systemic injections of 0.5mg/kg sodium selenate two hours before taste aversion conditioning form attenuated taste aversions that extinguish rapidly. Selenate also decreases LiCl-induced phospho-MAP kinase in the NTS and lateral PBN.

Results suggest that selenate interferes with the regulation of learning-induced intracellular signaling by PP2A, and prevents consolidation of memories acquired during CTA learning. However, an alternative explanation is that sodium selenate affects LiCl-induced malaise and prevents the rats from associating the conditioned stimulus (saccharin taste) with LiCl by dampening the effects of LiCl toxicity.

Rats typically display lying-on-belly (LOB) behavior in response to LiCl-induced malaise. The time to onset of LOB behavior has been shown to decrease as the dose of LiCl increases (2), so the time to onset of LOB can be used to infer the magnitude of LiCl-induced malaise.

The [[MBF Selenate Lying on Belly]] experiments determined that a 20 ml/kg i.p. injection of 0.15 LiCl reliably induces LOB behavior in male Sprague Dawley rats within 30 minutes.
*The low dose of LiCl (0.15M, 6 ml/kg, i.p.) did not induce LOB behavior
*The moderate dose of LiCl (0.15M, 12 ml/kg, i.p.) reliably induced pica but did not induce LOB in all of the rats.

This experiment aims to determine if sodium selenate pretreatment weakens the toxic effects of LiCl by measuring the time to onset of LOB after LiCl (0.15M, 20 ml/kg, i.p.) administration in selenate-pretreated vs control rats.

=Agenda=

MBG01-08
*A 0.5mg/ml solution of sodium selenate in 0.15M NaCl was prepared the day before the experiment.
*Male Sprague Dawley rats (n=8/group) were given selenate (1 ml/kg, i.p.) or NaCl (0.15M, 1 ml/kg, i.p.) two hours before receiving LiCl (0.15M, 20 ml/kg, i.p.) or NaCl (0.15M, 20 ml/kg, i.p.).
*Immediately after the LiCl injections, rats were placed in a test cage and video recorded for 30 minutes.
*Recordings were stopped after 30 minutes and the rats were returned to their home cages.
*Videos were scored for the time to onset of LOB for all rats.

{| class="wikitable"
|+ Drug Injection Schedule
|-
! Subject !! Day 1 !!Day 2 !! Day 3 !! Day 4
|-
| MBG01 || NaCl/NaCl || Sel/NaCl || Sel/LiCl || NaCl/LiCl
|-
| MBG02 || Sel/NaCl || NaCl/NaCl || NaCl/LiCl || Sel/LiCl
|-
| MBG03 || NaCl/LiCl || Sel/LiCl || Sel/NaCl || NaCl/NaCl
|-
| MBG04 || Sel/LiCl || NaCl/LiCl || NaCl/NaCl || Sel/NaCl
|-
| MBG05 || NaCl/LiCl || Sel/NaCl || NaCl/NaCl || Sel/LiCl
|-
| MBG06 || Sel/LiCl || NaCl/LiCl || Sel/NaCl || NaCl/NaCl
|-
| MBG07 || NaCl/NaCl || Sel/LiCl || NaCl/LiCl || Sel/NaCl
|-
| MBG08 || Sel/LiCl || NaCl/NaCl || Sel/LiCl || NaCl/LiCl
|}

=Results=
*Selenate pretreatment did not affect the time to onset of LOB.
*Two-way ANOVAs with pretreatment (vehicle or selenate) and LiCl condition (LiCl or NaCl) as factors revealed a significant effect of LiCl condition, but not pretreatment, on time to LOB onset (F[1,28] = 35.15, p<1e-05).

=References=
Corcoran NM, Martin D, Hutter-Paier B, Windisch M, Nguyen T, Nheu L, Sundstrom LE, Costello AJ, Hovens CM. Sodium selenate specifically activates PP2A phosphatase, dephosphorylates tau and reverses memory deficits in an Alzheimer’s disease model. Journal of Clinical Neuroscience 17: 1025- 1033, 2010.[https://pubmed.ncbi.nlm.nih.gov/20537899/]

Nunnink M, Davenport RA, Ortega B, Houpt TA. d-Cycloserine enhances conditioned taste aversion learning in rats. Pharmacology Biochemistry and Behavior 87(3): 321-330, 2007. [https://www.sciencedirect.com/science/article/pii/S0091305707001517?via%3Dihub]

=People=
Marena Bass

[[Category:CTA Experiments]]
[[Category:Taste_Aversion]]

MBG Selenate Lying on Belly

2023-07-05T21:30:09Z

MBass: /* Agenda */ added table caption

Selenate modulation of LiCl-induced Lying-on-Belly

=Background=

[[Selenium|Sodium Selenate]] (Na2Se04) activates PP2A (1).

[[CTA and Selenate]] experiments show that male Sprague Dawley rats given systemic injections of 0.5mg/kg sodium selenate two hours before taste aversion conditioning form attenuated taste aversions that extinguish rapidly. Selenate also decreases LiCl-induced phospho-MAP kinase in the NTS and lateral PBN.

Results suggest that selenate interferes with the regulation of learning-induced intracellular signaling by PP2A, and prevents consolidation of memories acquired during CTA learning. However, an alternative explanation is that sodium selenate affects LiCl-induced malaise and prevents the rats from associating the conditioned stimulus (saccharin taste) with LiCl by dampening the effects of LiCl toxicity.

Rats typically display lying-on-belly (LOB) behavior in response to LiCl-induced malaise. The time to onset of LOB behavior has been shown to decrease as the dose of LiCl increases (2), so the time to onset of LOB can be used to infer the magnitude of LiCl-induced malaise. This experiment aims to determine if sodium selenate pretreatment weakens the toxic effects of LiCl by measuring the time to onset of LOB after LiCl (0.15M, 20 ml/kg, i.p.) administration in selenate-pretreated vs control rats.

=Agenda=
The first set of LOB experiments (MBF01-06) determined that a 20 ml/kg i.p. injection of 0.15 LiCl reliably induces LOB behavior in male Sprague Dawley rats within 30 minutes.
*The low dose of LiCl (0.15M, 6 ml/kg, i.p.) did not induce LOB behavior
*The moderate dose of LiCl (0.15M, 12 ml/kg, i.p.) reliably induced pica but did not induce LOB in all of the rats.

MBG01-08
*A 0.5mg/ml solution of sodium selenate in 0.15M NaCl was prepared the day before the experiment.
*Male Sprague Dawley rats (n=8/group) were given selenate (1 ml/kg, i.p.) or NaCl (0.15M, 1 ml/kg, i.p.) two hours before receiving LiCl (0.15M, 20 ml/kg, i.p.) or NaCl (0.15M, 20 ml/kg, i.p.).
*Immediately after the LiCl injections, rats were placed in a test cage and video recorded for 30 minutes.
*Recordings were stopped after 30 minutes and the rats were returned to their home cages.
*Videos were scored for the time to onset of LOB for all rats.

{| class="wikitable"
|+ Drug Injection Schedule
|-
! Subject !! Day 1 !!Day 2 !! Day 3 !! Day 4
|-
| MBG01 || NaCl/NaCl || Sel/NaCl || Sel/LiCl || NaCl/LiCl
|-
| MBG02 || Sel/NaCl || NaCl/NaCl || NaCl/LiCl || Sel/LiCl
|-
| MBG03 || NaCl/LiCl || Sel/LiCl || Sel/NaCl || NaCl/NaCl
|-
| MBG04 || Sel/LiCl || NaCl/LiCl || NaCl/NaCl || Sel/NaCl
|-
| MBG05 || NaCl/LiCl || Sel/NaCl || NaCl/NaCl || Sel/LiCl
|-
| MBG06 || Sel/LiCl || NaCl/LiCl || Sel/NaCl || NaCl/NaCl
|-
| MBG07 || NaCl/NaCl || Sel/LiCl || NaCl/LiCl || Sel/NaCl
|-
| MBG08 || Sel/LiCl || NaCl/NaCl || Sel/LiCl || NaCl/LiCl
|}

=Results=
*Selenate pretreatment did not affect the time to onset of LOB.
*Two-way ANOVAs with pretreatment (vehicle or selenate) and LiCl condition (LiCl or NaCl) as factors revealed a significant effect of LiCl condition, but not pretreatment, on time to LOB onset (F[1,28] = 35.15, p<1e-05).

=References=
Corcoran NM, Martin D, Hutter-Paier B, Windisch M, Nguyen T, Nheu L, Sundstrom LE, Costello AJ, Hovens CM. Sodium selenate specifically activates PP2A phosphatase, dephosphorylates tau and reverses memory deficits in an Alzheimer’s disease model. Journal of Clinical Neuroscience 17: 1025- 1033, 2010.[https://pubmed.ncbi.nlm.nih.gov/20537899/]

Nunnink M, Davenport RA, Ortega B, Houpt TA. d-Cycloserine enhances conditioned taste aversion learning in rats. Pharmacology Biochemistry and Behavior 87(3): 321-330, 2007. [https://www.sciencedirect.com/science/article/pii/S0091305707001517?via%3Dihub]

=People=
Marena Bass

[[Category:CTA Experiments]]
[[Category:Taste_Aversion]]

MBG Selenate Lying on Belly

2023-07-05T21:29:32Z

MBass: /* Agenda */ added injection schedule

Selenate modulation of LiCl-induced Lying-on-Belly

=Background=

[[Selenium|Sodium Selenate]] (Na2Se04) activates PP2A (1).

[[CTA and Selenate]] experiments show that male Sprague Dawley rats given systemic injections of 0.5mg/kg sodium selenate two hours before taste aversion conditioning form attenuated taste aversions that extinguish rapidly. Selenate also decreases LiCl-induced phospho-MAP kinase in the NTS and lateral PBN.

Results suggest that selenate interferes with the regulation of learning-induced intracellular signaling by PP2A, and prevents consolidation of memories acquired during CTA learning. However, an alternative explanation is that sodium selenate affects LiCl-induced malaise and prevents the rats from associating the conditioned stimulus (saccharin taste) with LiCl by dampening the effects of LiCl toxicity.

Rats typically display lying-on-belly (LOB) behavior in response to LiCl-induced malaise. The time to onset of LOB behavior has been shown to decrease as the dose of LiCl increases (2), so the time to onset of LOB can be used to infer the magnitude of LiCl-induced malaise. This experiment aims to determine if sodium selenate pretreatment weakens the toxic effects of LiCl by measuring the time to onset of LOB after LiCl (0.15M, 20 ml/kg, i.p.) administration in selenate-pretreated vs control rats.

=Agenda=
The first set of LOB experiments (MBF01-06) determined that a 20 ml/kg i.p. injection of 0.15 LiCl reliably induces LOB behavior in male Sprague Dawley rats within 30 minutes.
*The low dose of LiCl (0.15M, 6 ml/kg, i.p.) did not induce LOB behavior
*The moderate dose of LiCl (0.15M, 12 ml/kg, i.p.) reliably induced pica but did not induce LOB in all of the rats.

MBG01-08
*A 0.5mg/ml solution of sodium selenate in 0.15M NaCl was prepared the day before the experiment.
*Male Sprague Dawley rats (n=8/group) were given selenate (1 ml/kg, i.p.) or NaCl (0.15M, 1 ml/kg, i.p.) two hours before receiving LiCl (0.15M, 20 ml/kg, i.p.) or NaCl (0.15M, 20 ml/kg, i.p.).
*Immediately after the LiCl injections, rats were placed in a test cage and video recorded for 30 minutes.
*Recordings were stopped after 30 minutes and the rats were returned to their home cages.
*Videos were scored for the time to onset of LOB for all rats.

{| class="wikitable"
|+ Caption text
|-
! Subject !! Day 1 !!Day 2 !! Day 3 !! Day 4
|-
| MBG01 || NaCl/NaCl || Sel/NaCl || Sel/LiCl || NaCl/LiCl
|-
| MBG02 || Sel/NaCl || NaCl/NaCl || NaCl/LiCl || Sel/LiCl
|-
| MBG03 || NaCl/LiCl || Sel/LiCl || Sel/NaCl || NaCl/NaCl
|-
| MBG04 || Sel/LiCl || NaCl/LiCl || NaCl/NaCl || Sel/NaCl
|-
| MBG05 || NaCl/LiCl || Sel/NaCl || NaCl/NaCl || Sel/LiCl
|-
| MBG06 || Sel/LiCl || NaCl/LiCl || Sel/NaCl || NaCl/NaCl
|-
| MBG07 || NaCl/NaCl || Sel/LiCl || NaCl/LiCl || Sel/NaCl
|-
| MBG08 || Sel/LiCl || NaCl/NaCl || Sel/LiCl || NaCl/LiCl
|}

=Results=
*Selenate pretreatment did not affect the time to onset of LOB.
*Two-way ANOVAs with pretreatment (vehicle or selenate) and LiCl condition (LiCl or NaCl) as factors revealed a significant effect of LiCl condition, but not pretreatment, on time to LOB onset (F[1,28] = 35.15, p<1e-05).

=References=
Corcoran NM, Martin D, Hutter-Paier B, Windisch M, Nguyen T, Nheu L, Sundstrom LE, Costello AJ, Hovens CM. Sodium selenate specifically activates PP2A phosphatase, dephosphorylates tau and reverses memory deficits in an Alzheimer’s disease model. Journal of Clinical Neuroscience 17: 1025- 1033, 2010.[https://pubmed.ncbi.nlm.nih.gov/20537899/]

Nunnink M, Davenport RA, Ortega B, Houpt TA. d-Cycloserine enhances conditioned taste aversion learning in rats. Pharmacology Biochemistry and Behavior 87(3): 321-330, 2007. [https://www.sciencedirect.com/science/article/pii/S0091305707001517?via%3Dihub]

=People=
Marena Bass

[[Category:CTA Experiments]]
[[Category:Taste_Aversion]]

MBF Selenate Lying on Belly

2023-07-05T21:09:08Z

MBass: /* Background */ edited last sentence for relevance to this experiment

Selenate modulation of LiCl-induced Lying-on-Belly

=Background=

[[Selenium|Sodium Selenate]] (Na2Se04) activates PP2A (1).

[[CTA and Selenate]] experiments show that male Sprague Dawley rats given systemic injections of 0.5mg/kg sodium selenate two hours before taste aversion conditioning form attenuated taste aversions that extinguish rapidly. Selenate also decreases LiCl-induced phospho-MAP kinase in the NTS and lateral PBN.

Results suggest that selenate interferes with the regulation of learning-induced intracellular signaling by PP2A, and prevents consolidation of memories acquired during CTA learning. However, an alternative explanation is that sodium selenate affects LiCl-induced malaise and prevents the rats from associating the conditioned stimulus (saccharin taste) with LiCl by dampening the effects of LiCl toxicity.

Rats typically display lying-on-belly (LOB) behavior in response to LiCl-induced malaise. The time to onset of LOB behavior has been shown to decrease as the dose of LiCl increases (2), so the time to onset of LOB can be used to infer the magnitude of LiCl-induced malaise. This experiment aims to determine if sodium selenate pretreatment weakens the toxic effects of LiCl by measuring the time to onset of LOB after injections of 20ml/kg, 12ml/kg and 6ml/kg LiCl (0.15M, i.p.) in selenate-pretreated vs control rats.

=Agenda=

* A 0.5mg/ml solution of sodium selenate in 0.15M NaCl was prepared the day before the experiment.

* Male Sprague Dawley rats (n=6/group) were given selenate (1 ml/kg, i.p.) or NaCl (0.15M, 1 ml/kg, i.p.) two hours before receiving 6ml/kg, 12ml/kg or 20ml/kg injections of LiCl (0.15M, i.p.) or NaCl (0.15M, i.p.).

* Immediately after the LiCl injections, rats were returned to their home cages and video recorded for 30-60 minutes.

*Videos were scored for the time to onset of LOB for all rats.

{| class="wikitable"
|+ Drug Injection Summary
|-
! Subject !! Day1 (6ml/kg) !! Day2 (12ml/kg) !! Day3 (20ml/kg) !! Day4 (12ml/kg) !! Day5 (12ml/kg) !! Day6 (12ml/kg)
|-
| MBF01 || NaCl/NaCl || Sel/NaCl || Sel/LiCl || NaCl/LiCl || Sel/LiCl || NaCl/NaCl
|-
| MBF02 || Sel/NaCl || Sel/LiCl || NaCl/LiCl || NaCl/NaCl || NaCl/LiCl || Sel/NaCl
|-
| MBF03 || Sel/LiCl || NaCl/LiCl || Sel/NaCl || NaCl/NaCl || Sel/LiCl || Sel/NaCl
|-
| MBF04 || NaCl/LiCl || NaCl/NaCl || NaCl/NaCl || Sel/LiCl || NaCl/LiCl || Sel/NaCl
|-
| MBF05 || NaCl/LiCl || Sel/LiCl || NaCl/NaCl || Sel/NaCl || NaCl/NaCl || NaCl/LiCl
|-
| MBF06 || Sel/LiCl || Sel/NaCl || Sel/NaCl || NaCl/LiCl || NaCl/NaCl || Sel/LiCl
|}

=Results=

=References=
Corcoran NM, Martin D, Hutter-Paier B, Windisch M, Nguyen T, Nheu L, Sundstrom LE, Costello AJ, Hovens CM. Sodium selenate specifically activates PP2A phosphatase, dephosphorylates tau and reverses memory deficits in an Alzheimer’s disease model. Journal of Clinical Neuroscience 17: 1025- 1033, 2010.[https://pubmed.ncbi.nlm.nih.gov/20537899/]

Nunnink M, Davenport RA, Ortega B, Houpt TA. d-Cycloserine enhances conditioned taste aversion learning in rats. Pharmacology Biochemistry and Behavior 87(3): 321-330, 2007. [https://www.sciencedirect.com/science/article/pii/S0091305707001517?via%3Dihub]

=People=
Marena Bass

[[Category:CTA Experiments]]
[[Category:Taste_Aversion]]

MBF Selenate Lying on Belly

2023-07-05T21:03:00Z

MBass: added agenda

Selenate modulation of LiCl-induced Lying-on-Belly

=Background=

[[Selenium|Sodium Selenate]] (Na2Se04) activates PP2A (1).

[[CTA and Selenate]] experiments show that male Sprague Dawley rats given systemic injections of 0.5mg/kg sodium selenate two hours before taste aversion conditioning form attenuated taste aversions that extinguish rapidly. Selenate also decreases LiCl-induced phospho-MAP kinase in the NTS and lateral PBN.

Results suggest that selenate interferes with the regulation of learning-induced intracellular signaling by PP2A, and prevents consolidation of memories acquired during CTA learning. However, an alternative explanation is that sodium selenate affects LiCl-induced malaise and prevents the rats from associating the conditioned stimulus (saccharin taste) with LiCl by dampening the effects of LiCl toxicity.

Rats typically display lying-on-belly (LOB) behavior in response to LiCl-induced malaise. The time to onset of LOB behavior has been shown to decrease as the dose of LiCl increases (2), so the time to onset of LOB can be used to infer the magnitude of LiCl-induced malaise. This experiment aims to determine if sodium selenate pretreatment weakens the toxic effects of LiCl by measuring the time to onset of LOB after LiCl (0.15M, 20 ml/kg, i.p.) administration in selenate-pretreated vs control rats.

=Agenda=

* A 0.5mg/ml solution of sodium selenate in 0.15M NaCl was prepared the day before the experiment.

* Male Sprague Dawley rats (n=6/group) were given selenate (1 ml/kg, i.p.) or NaCl (0.15M, 1 ml/kg, i.p.) two hours before receiving 6ml/kg, 12ml/kg or 20ml/kg injections of LiCl (0.15M, i.p.) or NaCl (0.15M, i.p.).

* Immediately after the LiCl injections, rats were returned to their home cages and video recorded for 30-60 minutes.

*Videos were scored for the time to onset of LOB for all rats.

{| class="wikitable"
|+ Drug Injection Summary
|-
! Subject !! Day1 (6ml/kg) !! Day2 (12ml/kg) !! Day3 (20ml/kg) !! Day4 (12ml/kg) !! Day5 (12ml/kg) !! Day6 (12ml/kg)
|-
| MBF01 || NaCl/NaCl || Sel/NaCl || Sel/LiCl || NaCl/LiCl || Sel/LiCl || NaCl/NaCl
|-
| MBF02 || Sel/NaCl || Sel/LiCl || NaCl/LiCl || NaCl/NaCl || NaCl/LiCl || Sel/NaCl
|-
| MBF03 || Sel/LiCl || NaCl/LiCl || Sel/NaCl || NaCl/NaCl || Sel/LiCl || Sel/NaCl
|-
| MBF04 || NaCl/LiCl || NaCl/NaCl || NaCl/NaCl || Sel/LiCl || NaCl/LiCl || Sel/NaCl
|-
| MBF05 || NaCl/LiCl || Sel/LiCl || NaCl/NaCl || Sel/NaCl || NaCl/NaCl || NaCl/LiCl
|-
| MBF06 || Sel/LiCl || Sel/NaCl || Sel/NaCl || NaCl/LiCl || NaCl/NaCl || Sel/LiCl
|}

=Results=

=References=
Corcoran NM, Martin D, Hutter-Paier B, Windisch M, Nguyen T, Nheu L, Sundstrom LE, Costello AJ, Hovens CM. Sodium selenate specifically activates PP2A phosphatase, dephosphorylates tau and reverses memory deficits in an Alzheimer’s disease model. Journal of Clinical Neuroscience 17: 1025- 1033, 2010.[https://pubmed.ncbi.nlm.nih.gov/20537899/]

Nunnink M, Davenport RA, Ortega B, Houpt TA. d-Cycloserine enhances conditioned taste aversion learning in rats. Pharmacology Biochemistry and Behavior 87(3): 321-330, 2007. [https://www.sciencedirect.com/science/article/pii/S0091305707001517?via%3Dihub]

=People=
Marena Bass

[[Category:CTA Experiments]]
[[Category:Taste_Aversion]]

MBF Selenate Lying on Belly

2023-07-05T20:58:09Z

MBass: /* Agenda */

Selenate modulation of LiCl-induced Lying-on-Belly

=Background=

[[Selenium|Sodium Selenate]] (Na2Se04) activates PP2A (1).

[[CTA and Selenate]] experiments show that male Sprague Dawley rats given systemic injections of 0.5mg/kg sodium selenate two hours before taste aversion conditioning form attenuated taste aversions that extinguish rapidly. Selenate also decreases LiCl-induced phospho-MAP kinase in the NTS and lateral PBN.

Results suggest that selenate interferes with the regulation of learning-induced intracellular signaling by PP2A, and prevents consolidation of memories acquired during CTA learning. However, an alternative explanation is that sodium selenate affects LiCl-induced malaise and prevents the rats from associating the conditioned stimulus (saccharin taste) with LiCl by dampening the effects of LiCl toxicity.

Rats typically display lying-on-belly (LOB) behavior in response to LiCl-induced malaise. The time to onset of LOB behavior has been shown to decrease as the dose of LiCl increases (2), so the time to onset of LOB can be used to infer the magnitude of LiCl-induced malaise. This experiment aims to determine if sodium selenate pretreatment weakens the toxic effects of LiCl by measuring the time to onset of LOB after LiCl (0.15M, 20 ml/kg, i.p.) administration in selenate-pretreated vs control rats.

{| class="wikitable"
|+ Drug Injection Summary
|-
! Subject !! Day1 (6ml/kg) !! Day2 (12ml/kg) !! Day3 (20ml/kg) !! Day4 (12ml/kg) !! Day5 (12ml/kg) !! Day6 (12ml/kg)
|-
| MBF01 || NaCl/NaCl || Sel/NaCl || Sel/LiCl || NaCl/LiCl || Sel/LiCl || NaCl/NaCl
|-
| MBF02 || Sel/NaCl || Sel/LiCl || NaCl/LiCl || NaCl/NaCl || NaCl/LiCl || Sel/NaCl
|-
| MBF03 || Sel/LiCl || NaCl/LiCl || Sel/NaCl || NaCl/NaCl || Sel/LiCl || Sel/NaCl
|-
| MBF04 || NaCl/LiCl || NaCl/NaCl || NaCl/NaCl || Sel/LiCl || NaCl/LiCl || Sel/NaCl
|-
| MBF05 || NaCl/LiCl || Sel/LiCl || NaCl/NaCl || Sel/NaCl || NaCl/NaCl || NaCl/LiCl
|-
| MBF06 || Sel/LiCl || Sel/NaCl || Sel/NaCl || NaCl/LiCl || NaCl/NaCl || Sel/LiCl
|}
=Agenda=

=Results=

=References=
Corcoran NM, Martin D, Hutter-Paier B, Windisch M, Nguyen T, Nheu L, Sundstrom LE, Costello AJ, Hovens CM. Sodium selenate specifically activates PP2A phosphatase, dephosphorylates tau and reverses memory deficits in an Alzheimer’s disease model. Journal of Clinical Neuroscience 17: 1025- 1033, 2010.[https://pubmed.ncbi.nlm.nih.gov/20537899/]

Nunnink M, Davenport RA, Ortega B, Houpt TA. d-Cycloserine enhances conditioned taste aversion learning in rats. Pharmacology Biochemistry and Behavior 87(3): 321-330, 2007. [https://www.sciencedirect.com/science/article/pii/S0091305707001517?via%3Dihub]

=People=
Marena Bass

[[Category:CTA Experiments]]
[[Category:Taste_Aversion]]

CTA and Selenate

2023-07-05T20:47:59Z

MBass: /* Behavioral Experiments */

== Behavioral Experiments ==

* [[JB Systemic Sodium Selenate]]

* [[JC & JD Systemic Sodium Selenate]]

* Selenate-induced CTA in Rats (round 2) (expt JE)

* [[JF Systemic Sodium Selenate]] (includes polycose retest)

* [[MBG Selenate Lying on Belly]] Selenate modulation of LiCl-induced Lying-on-Belly

* [[MBF Selenate Lying on Belly]] Selenate modulation of LiCl-induced Lying-on-Belly

== Immunohistochemical Experiments ==

* Selenate-induced c-Fos

# JB c-fos
# JC c-fos
# JD c-fos
# HHA01-06

* Selenate modulation of LiCl-induced c-Fos and pMAPK

# JG attenuation of c-Fos and pMAPK by selenate
# JI attenuation of c-Fos and pMAPK by selenate

[[Category:CTA_Experiments]]
[[Category:Taste_Aversion]]

MBF Selenate Lying on Belly

2023-07-05T20:46:12Z

MBass: Created page with "Selenate modulation of LiCl-induced Lying-on-Belly =Background= Sodium Selenate (Na2Se04) activates PP2A (1). CTA and Selenate experiments show that male S..."

CTA and Selenate

2023-06-29T17:27:27Z

MBass: added link to MBG

== Behavioral Experiments ==

* [[JB Systemic Sodium Selenate]]

* [[JC & JD Systemic Sodium Selenate]]

* Selenate-induced CTA in Rats (round 2) (expt JE)

* [[JF Systemic Sodium Selenate]] (includes polycose retest)

* [[MBG Selenate Lying on Belly]] Selenate modulation of LiCl-induced Lying-on-Belly

== Immunohistochemical Experiments ==

* Selenate-induced c-Fos

# JB c-fos
# JC c-fos
# JD c-fos
# HHA01-06

* Selenate modulation of LiCl-induced c-Fos and pMAPK

# JG attenuation of c-Fos and pMAPK by selenate
# JI attenuation of c-Fos and pMAPK by selenate

[[Category:CTA_Experiments]]
[[Category:Taste_Aversion]]

MBG Selenate Lying on Belly

2023-06-29T17:23:49Z

MBass: this page is mostly complete now

Selenate modulation of LiCl-induced Lying-on-Belly

=Background=

[[Selenium|Sodium Selenate]] (Na2Se04) activates PP2A (1).

[[CTA and Selenate]] experiments show that male Sprague Dawley rats given systemic injections of 0.5mg/kg sodium selenate two hours before taste aversion conditioning form attenuated taste aversions that extinguish rapidly. Selenate also decreases LiCl-induced phospho-MAP kinase in the NTS and lateral PBN.

Results suggest that selenate interferes with the regulation of learning-induced intracellular signaling by PP2A, and prevents consolidation of memories acquired during CTA learning. However, an alternative explanation is that sodium selenate affects LiCl-induced malaise and prevents the rats from associating the conditioned stimulus (saccharin taste) with LiCl by dampening the effects of LiCl toxicity.

Rats typically display lying-on-belly (LOB) behavior in response to LiCl-induced malaise. The time to onset of LOB behavior has been shown to decrease as the dose of LiCl increases (2), so the time to onset of LOB can be used to infer the magnitude of LiCl-induced malaise. This experiment aims to determine if sodium selenate pretreatment weakens the toxic effects of LiCl by measuring the time to onset of LOB after LiCl (0.15M, 20 ml/kg, i.p.) administration in selenate-pretreated vs control rats.

=Agenda=
The first set of LOB experiments (MBF01-06) determined that a 20 ml/kg i.p. injection of 0.15 LiCl reliably induces LOB behavior in male Sprague Dawley rats within 30 minutes.
*The low dose of LiCl (0.15M, 6 ml/kg, i.p.) did not induce LOB behavior
*The moderate dose of LiCl (0.15M, 12 ml/kg, i.p.) reliably induced pica but did not induce LOB in all of the rats.

MBG01-08
*A 0.5mg/ml solution of sodium selenate in 0.15M NaCl was prepared the day before the experiment.
*Male Sprague Dawley rats (n=8/group) were given selenate (1 ml/kg, i.p.) or NaCl (0.15M, 1 ml/kg, i.p.) two hours before receiving LiCl (0.15M, 20 ml/kg, i.p.) or NaCl (0.15M, 20 ml/kg, i.p.).
*Immediately after the LiCl injections, rats were placed in a test cage and video recorded for 30 minutes.
*Recordings were stopped after 30 minutes and the rats were returned to their home cages.
*Videos were scored for the time to onset of LOB for all rats.

=Results=
*Selenate pretreatment did not affect the time to onset of LOB.
*Two-way ANOVAs with pretreatment (vehicle or selenate) and LiCl condition (LiCl or NaCl) as factors revealed a significant effect of LiCl condition, but not pretreatment, on time to LOB onset (F[1,28] = 35.15, p<1e-05).

=References=
Corcoran NM, Martin D, Hutter-Paier B, Windisch M, Nguyen T, Nheu L, Sundstrom LE, Costello AJ, Hovens CM. Sodium selenate specifically activates PP2A phosphatase, dephosphorylates tau and reverses memory deficits in an Alzheimer’s disease model. Journal of Clinical Neuroscience 17: 1025- 1033, 2010.[https://pubmed.ncbi.nlm.nih.gov/20537899/]

Nunnink M, Davenport RA, Ortega B, Houpt TA. d-Cycloserine enhances conditioned taste aversion learning in rats. Pharmacology Biochemistry and Behavior 87(3): 321-330, 2007. [https://www.sciencedirect.com/science/article/pii/S0091305707001517?via%3Dihub]

=People=
Marena Bass

[[Category:CTA Experiments]]
[[Category:Taste_Aversion]]

MBG Selenate Lying on Belly

2023-06-29T15:46:16Z

MBass: added to background

MBG Selenate Lying on Belly

2023-06-29T15:33:39Z

MBass: Creating wiki page for LOB experiments

Selenate modulation of LiCl-induced Lying-on-Belly

=Background=

[[Selenium|Sodium Selenate]] (Na2Se04) activates PP2A (1).

* [[CTA and Selenate]] experiments show that male Sprague Dawley rats given systemic injections of 0.5mg/kg sodium selenate two hours before taste aversion conditioning form attenuated taste aversions that extinguish rapidly. Selenate also decreases LiCl-induced phospho-MAP kinase in the NTS and lateral PBN.

=Agenda=

=Results=

=References=

Corcoran NM, Martin D, Hutter-Paier B, Windisch M, Nguyen T, Nheu L, Sundstrom LE, Costello AJ, Hovens CM. Sodium selenate specifically activates PP2A phosphatase, dephosphorylates tau and reverses memory deficits in an Alzheimer’s disease model. Journal of Clinical Neuroscience 17: 1025- 1033, 2010.[https://pubmed.ncbi.nlm.nih.gov/20537899/]

=People=
Marena Bass

[[Category:CTA Experiments]]
[[Category:Taste_Aversion]]

Electronic Esophagus

2022-05-22T13:03:24Z

MBass: /* Assembly */

Electronic Esophagus

2022-05-22T12:36:35Z

MBass: /* Electronics */

Selection, construction and programming of the apparatus hardware was modified from open-source syringe pump manuscripts by Longley et al (2017) and Wijen et al (2014). A capacitive touch sensor (Adafruit MPR121) is connected to the rats’ sipper tubes and programmed to detect each lick from the spout with millisecond accuracy. The capacitive touch sensor relays its information to an Arduino- compatible microcontroller (Teensy-LC) which subsequently signals a stepper motor driver (Pololu DRV8825) to turn a stepper motor (NEMA-17, 200 steps/rev, 12V, 350mA), resulting in the release of 4–8 μl liquid per lick from a 20 ml syringe connected to the stepper motor (Figure 4). The syringe pumps liquid into the rat’s chronic gastric catheter at a rate of 1 ml/min over 10 min. This infusion rate is slow enough that the stomach easily accommodates the fill and allows for gastric emptying into the intestine at a normal rate.

Using a protocol adapted from Elizalde & Sclafani (1990), unconditioned stimuli (glucose or saline) will be infused intragastrically as the animals orally sample distinct flavored solutions. With this "Electronic Esophagus" preparation utilized by Sclafani and others, animals form robust and persistent preferences for the flavored solution that is paired with glucose infusions.

=Parts=

==Electronics==
[https://www.pjrc.com/teensy/teensyLC.html Teensy-LC]

[https://www.adafruit.com/product/1982 MPR121 Capacitive Touch Sensor]

[https://www.pololu.com/product/2133/ DRV8825 Stepper Motor Driver]

[https://www.adafruit.com/product/324 Stepper motor, NEMA-17, 200 steps/rev, 12V 350mA]

==Hardware==
1 [https://www.adafruit.com/product/1176 Aluminum Flex Shaft Coupler - 5mm to 8mm]

1 [https://www.adafruit.com/product/1181 Linear Ball Bearing - 8mm diameter]

1 M8 x 250mm Fully Threaded Rod, 304 Stainless Steel, Right Hand Threads [https://www.amazon.com/dp/B078H2ZM3C?ref=ppx_yo2_dt_b_product_details&th=1 like this one]

1 Linear Motion Rod 8mmx 250mm [https://www.amazon.com/dp/B08JFZQG2H?ref=ppx_yo2_dt_b_product_details&th=1 like these]

1 M8-1.25 Hex Nut [https://www.amazon.com/Persberg-M8-1-25-Height-Stainless-120153/dp/B089PR8DVP/ref=sr_1_1_sspa?crid=29DW29A9AWQEP&keywords=m8+hex+nut&qid=1642699078&s=industrial&sprefix=m8+he%2Cindustrial%2C87&sr=1-1-spons&psc=1&spLa=ZW5jcnlwdGVkUXVhbGlmaWVyPUEyUEtNOVlXSlkyR00wJmVuY3J5cHRlZElkPUEwMzc5MzU5MjJSWVcyRDg5MDY4TSZlbmNyeXB0ZWRBZElkPUEwNzUzNTQ3M0Q4TEZKWTJZRDdCViZ3aWRnZXROYW1lPXNwX2F0ZiZhY3Rpb249Y2xpY2tSZWRpcmVjdCZkb05vdExvZ0NsaWNrPXRydWU= like these]

==3D Printed Parts==
The 4 parts are currently available as STL and 3MF files.
[https://drive.google.com/drive/folders/1_GCll42MNhbK981q99WKfd3auBnq7ENU?usp=sharing Download here]

==Software==
[https://learn.adafruit.com/adafruit-mpr121-12-key-capacitive-touch-sensor-breakout-tutorial/wiring Adafruit Tutorial]

[https://www.arduino.cc/en/software Arduino IDE]

[https://www.pjrc.com/teensy/teensyduino.html Teensyduino add-on]

[https://circuitjournal.com/arduino-serial-to-spreadsheet ArduSpreadsheet add-on]

=Assembly=

==Electronics==
The MPR121 capacitive touch sensor can be connected to up to 12 capacitive electrodes (e.g., 12 sipper tubes). Its GND, 3.3V, SCL and SDA pins are connected to the corresponding Teensy-LC pins. The 5V and GND Teensy-LC pins provide the logic power supply to the DRV8825 motor driver, which is connected to a 12V DC power supply with a 100 uF decoupling capacitor to power to the stepper motor. The direction (DIR), step (STP) and enable (ENA) pins of the motor driver receive signals from the Teensy-LC digital pins D0, D1 and D2 respectively. Because the reset (RES) and sleep (SLP) motor driver pins are active low pins, they are connected to the logic power supply to ensure they remain inactive. [https://drive.google.com/drive/folders/1_GCll42MNhbK981q99WKfd3auBnq7ENU Fritzing schematic of electronic esophagus circuit.]

==Hardware and 3D Printed Parts==

=Code=
Arduino code is written in C/C++

//include relevant libraries
#include <Wire.h>
#include "Adafruit_MPR121.h"
#include <AccelStepper.h>

//define MPR121 info
#ifndef _BV
#define _BV(bit) (1 << (bit))
#endif
Adafruit_MPR121 cap = Adafruit_MPR121();
uint16_t lasttouched = 0;
uint16_t currtouched = 0;

//define DRV8825 stuff:
#define dirPin 2
#define stepPin 3
#define motorInterfaceType 1

// Create a new instance of the AccelStepper class:
AccelStepper stepper = AccelStepper(motorInterfaceType, stepPin, dirPin);

//setup code here, to run once:
void setup() {
stepper.setMaxSpeed(1000); //set max stepper speed in steps per second
Serial.begin(9600); //start serial monitor
while (!Serial) { // needed to keep from starting too fast!
delay(10);
}
Serial.println("Electronic Esophagus test start");
// Default address is 0x5A, if tied to 3.3V its 0x5B
// If tied to SDA its 0x5C and if SCL then 0x5D
if (!cap.begin(0x5A)) {
Serial.println("MPR121 not found, check wiring?");
while (1);
}
Serial.println("MPR121 found!");
// to change threshold sensitivites (touched, released). Defaults are (12,6). Valuess from 0-255 = less-more sensitive.
cap.setThresholds(24, 12);
}

// main code here, to run repeatedly:
void loop() {
// Get the currently touched pads
currtouched = cap.touched();

for (uint8_t i=0; i<12; i++) {
// it if *is* touched and *wasnt* touched before, alert!
if ((currtouched & _BV(i)) && !(lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" touched");
// set the position to 0:
stepper.setCurrentPosition(0);
// Run the motor forward at 400 steps/second until the motor reaches 12 steps (~0.125 revolutions):
while(stepper.currentPosition() != 12)
{
stepper.setSpeed(400);
stepper.runSpeed();
}
delayMicroseconds(500);
}

// if it *was* touched and now *isnt*, alert!
if (!(currtouched & _BV(i)) && (lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" released");
stepper.setCurrentPosition(0);
stepper.stop(); // Stop as fast as possible: sets new target
stepper.runToPosition(); // Now stopped after quickstop
delayMicroseconds(500);
}
}
// reset our state
lasttouched = currtouched;
// comment out "return" line for detailed data from the sensor!
return;
// debugging info, what
Serial.print("\t\t\t\t\t\t\t\t\t\t\t\t\t 0x"); Serial.println(cap.touched(), HEX);
Serial.print("Filt: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.filteredData(i)); Serial.print("\t");
}
Serial.println();
Serial.print("Base: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.baselineData(i)); Serial.print("\t");
}
Serial.println();
// put a delay so it isn't overwhelming
delay(100);
}

=References=
Dr. D-Flo. (2017, Feb 20). DIY Syringe Pump (Food 3D Printer - Part 1) [Video]. YouTube. [https://www.youtube.com/watch?v=UHa-OKb_CiM&t=212s]

Elizalde G & Sclafani A, Flavor preferences conditioned by intragastric polycose infusions: A detailed analysis using an electronic esophagus preparation, Physiology & Behavior 47: 63-67, 1990. [https://pubmed.ncbi.nlm.nih.gov/2109327/]

Longley M et al. An open source device for operant licking in rats. PeerJ 5:e2981, 2017. [https://peerj.com/articles/2981/]

Wijen B et al. Open-source syringe pump library. PLOS ONE 9(9): e107216, 2014. [https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0107216]

=People=
Marena Bass

[[Category:Flavor Preference]]

Electronic Esophagus

2022-05-22T12:28:09Z

MBass: typo

Selection, construction and programming of the apparatus hardware was modified from open-source syringe pump manuscripts by Longley et al (2017) and Wijen et al (2014). A capacitive touch sensor (Adafruit MPR121) is connected to the rats’ sipper tubes and programmed to detect each lick from the spout with millisecond accuracy. The capacitive touch sensor relays its information to an Arduino- compatible microcontroller (Teensy-LC) which subsequently signals a stepper motor driver (Pololu DRV8825) to turn a stepper motor (NEMA-17, 200 steps/rev, 12V, 350mA), resulting in the release of 4–8 μl liquid per lick from a 20 ml syringe connected to the stepper motor (Figure 4). The syringe pumps liquid into the rat’s chronic gastric catheter at a rate of 1 ml/min over 10 min. This infusion rate is slow enough that the stomach easily accommodates the fill and allows for gastric emptying into the intestine at a normal rate.

Using a protocol adapted from Elizalde & Sclafani (1990), unconditioned stimuli (glucose or saline) will be infused intragastrically as the animals orally sample distinct flavored solutions. With this "Electronic Esophagus" preparation utilized by Sclafani and others, animals form robust and persistent preferences for the flavored solution that is paired with glucose infusions.

=Parts=

==Electronics==
[https://www.pjrc.com/teensy/teensyLC.html Teensy-LC]

[https://www.adafruit.com/product/1982 MPR121 Capacitive Touch Sensor]

[https://www.pololu.com/product/2133/ DRV8825 Stepper Motor Driver]

[https://www.adafruit.com/product/324 Stepper motor, NEMA-17, 200 steps/rev, 12V 350mA]

==Hardware==
1 [https://www.adafruit.com/product/1176 Aluminum Flex Shaft Coupler - 5mm to 8mm]

1 [https://www.adafruit.com/product/1181 Linear Ball Bearing - 8mm diameter]

1 M8 x 250mm Fully Threaded Rod, 304 Stainless Steel, Right Hand Threads [https://www.amazon.com/dp/B078H2ZM3C?ref=ppx_yo2_dt_b_product_details&th=1 like this one]

1 Linear Motion Rod 8mmx 250mm [https://www.amazon.com/dp/B08JFZQG2H?ref=ppx_yo2_dt_b_product_details&th=1 like these]

1 M8-1.25 Hex Nut [https://www.amazon.com/Persberg-M8-1-25-Height-Stainless-120153/dp/B089PR8DVP/ref=sr_1_1_sspa?crid=29DW29A9AWQEP&keywords=m8+hex+nut&qid=1642699078&s=industrial&sprefix=m8+he%2Cindustrial%2C87&sr=1-1-spons&psc=1&spLa=ZW5jcnlwdGVkUXVhbGlmaWVyPUEyUEtNOVlXSlkyR00wJmVuY3J5cHRlZElkPUEwMzc5MzU5MjJSWVcyRDg5MDY4TSZlbmNyeXB0ZWRBZElkPUEwNzUzNTQ3M0Q4TEZKWTJZRDdCViZ3aWRnZXROYW1lPXNwX2F0ZiZhY3Rpb249Y2xpY2tSZWRpcmVjdCZkb05vdExvZ0NsaWNrPXRydWU= like these]

==3D Printed Parts==
The 4 parts are currently available as STL and 3MF files.
[https://drive.google.com/drive/folders/1_GCll42MNhbK981q99WKfd3auBnq7ENU?usp=sharing Download here]

==Software==
[https://learn.adafruit.com/adafruit-mpr121-12-key-capacitive-touch-sensor-breakout-tutorial/wiring Adafruit Tutorial]

[https://www.arduino.cc/en/software Arduino IDE]

[https://www.pjrc.com/teensy/teensyduino.html Teensyduino add-on]

[https://circuitjournal.com/arduino-serial-to-spreadsheet ArduSpreadsheet add-on]

=Assembly=

==Electronics==

==Hardware and 3D Printed Parts==

=Code=
Arduino code is written in C/C++

//include relevant libraries
#include <Wire.h>
#include "Adafruit_MPR121.h"
#include <AccelStepper.h>

//define MPR121 info
#ifndef _BV
#define _BV(bit) (1 << (bit))
#endif
Adafruit_MPR121 cap = Adafruit_MPR121();
uint16_t lasttouched = 0;
uint16_t currtouched = 0;

//define DRV8825 stuff:
#define dirPin 2
#define stepPin 3
#define motorInterfaceType 1

// Create a new instance of the AccelStepper class:
AccelStepper stepper = AccelStepper(motorInterfaceType, stepPin, dirPin);

//setup code here, to run once:
void setup() {
stepper.setMaxSpeed(1000); //set max stepper speed in steps per second
Serial.begin(9600); //start serial monitor
while (!Serial) { // needed to keep from starting too fast!
delay(10);
}
Serial.println("Electronic Esophagus test start");
// Default address is 0x5A, if tied to 3.3V its 0x5B
// If tied to SDA its 0x5C and if SCL then 0x5D
if (!cap.begin(0x5A)) {
Serial.println("MPR121 not found, check wiring?");
while (1);
}
Serial.println("MPR121 found!");
// to change threshold sensitivites (touched, released). Defaults are (12,6). Valuess from 0-255 = less-more sensitive.
cap.setThresholds(24, 12);
}

// main code here, to run repeatedly:
void loop() {
// Get the currently touched pads
currtouched = cap.touched();

for (uint8_t i=0; i<12; i++) {
// it if *is* touched and *wasnt* touched before, alert!
if ((currtouched & _BV(i)) && !(lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" touched");
// set the position to 0:
stepper.setCurrentPosition(0);
// Run the motor forward at 400 steps/second until the motor reaches 12 steps (~0.125 revolutions):
while(stepper.currentPosition() != 12)
{
stepper.setSpeed(400);
stepper.runSpeed();
}
delayMicroseconds(500);
}

// if it *was* touched and now *isnt*, alert!
if (!(currtouched & _BV(i)) && (lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" released");
stepper.setCurrentPosition(0);
stepper.stop(); // Stop as fast as possible: sets new target
stepper.runToPosition(); // Now stopped after quickstop
delayMicroseconds(500);
}
}
// reset our state
lasttouched = currtouched;
// comment out "return" line for detailed data from the sensor!
return;
// debugging info, what
Serial.print("\t\t\t\t\t\t\t\t\t\t\t\t\t 0x"); Serial.println(cap.touched(), HEX);
Serial.print("Filt: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.filteredData(i)); Serial.print("\t");
}
Serial.println();
Serial.print("Base: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.baselineData(i)); Serial.print("\t");
}
Serial.println();
// put a delay so it isn't overwhelming
delay(100);
}

=References=
Dr. D-Flo. (2017, Feb 20). DIY Syringe Pump (Food 3D Printer - Part 1) [Video]. YouTube. [https://www.youtube.com/watch?v=UHa-OKb_CiM&t=212s]

Elizalde G & Sclafani A, Flavor preferences conditioned by intragastric polycose infusions: A detailed analysis using an electronic esophagus preparation, Physiology & Behavior 47: 63-67, 1990. [https://pubmed.ncbi.nlm.nih.gov/2109327/]

Longley M et al. An open source device for operant licking in rats. PeerJ 5:e2981, 2017. [https://peerj.com/articles/2981/]

Wijen B et al. Open-source syringe pump library. PLOS ONE 9(9): e107216, 2014. [https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0107216]

=People=
Marena Bass

[[Category:Flavor Preference]]

Electronic Esophagus

2022-04-27T14:59:31Z

MBass: Added references and other headers

Selection, construction and programming of the apparatus hardware was modified from open-source syringe pump manuscripts by Longley et al (2017) and Wijen et al (2014). A capacitive touch sensor (Adafruit MPR121) is connected to the rats’ sipper tubes and programmed to detect each lick from the spout with millisecond accuracy. The capacitive touch sensor relays its information to an Arduino- compatible microcontroller (Teensy-LC) which subsequently signals a stepper motor driver (Pololu DRV8825) to turn a stepper motor (NEMA-17, 200 steps/rev, 12V, 350mA, resulting in the release of 4–8 μl liquid per lick from a 20 ml syringe connected to the stepper motor (Figure 4). The syringe pumps liquid into the rat’s chronic gastric catheter at a rate of 1 ml/min over 10 min. This infusion rate is slow enough that the stomach easily accommodates the fill and allows for gastric emptying into the intestine at a normal rate.

Using a protocol adapted from Elizalde & Sclafani (1990), unconditioned stimuli (glucose or saline) will be infused intragastrically as the animals orally sample distinct flavored solutions. With this "Electronic Esophagus" preparation utilized by Sclafani and others, animals form robust and persistent preferences for the flavored solution that is paired with glucose infusions.

=Parts=

==Electronics==
[https://www.pjrc.com/teensy/teensyLC.html Teensy-LC]

[https://www.adafruit.com/product/1982 MPR121 Capacitive Touch Sensor]

[https://www.pololu.com/product/2133/ DRV8825 Stepper Motor Driver]

[https://www.adafruit.com/product/324 Stepper motor, NEMA-17, 200 steps/rev, 12V 350mA]

==Hardware==
1 [https://www.adafruit.com/product/1176 Aluminum Flex Shaft Coupler - 5mm to 8mm]

1 [https://www.adafruit.com/product/1181 Linear Ball Bearing - 8mm diameter]

1 M8 x 250mm Fully Threaded Rod, 304 Stainless Steel, Right Hand Threads [https://www.amazon.com/dp/B078H2ZM3C?ref=ppx_yo2_dt_b_product_details&th=1 like this one]

1 Linear Motion Rod 8mmx 250mm [https://www.amazon.com/dp/B08JFZQG2H?ref=ppx_yo2_dt_b_product_details&th=1 like these]

1 M8-1.25 Hex Nut [https://www.amazon.com/Persberg-M8-1-25-Height-Stainless-120153/dp/B089PR8DVP/ref=sr_1_1_sspa?crid=29DW29A9AWQEP&keywords=m8+hex+nut&qid=1642699078&s=industrial&sprefix=m8+he%2Cindustrial%2C87&sr=1-1-spons&psc=1&spLa=ZW5jcnlwdGVkUXVhbGlmaWVyPUEyUEtNOVlXSlkyR00wJmVuY3J5cHRlZElkPUEwMzc5MzU5MjJSWVcyRDg5MDY4TSZlbmNyeXB0ZWRBZElkPUEwNzUzNTQ3M0Q4TEZKWTJZRDdCViZ3aWRnZXROYW1lPXNwX2F0ZiZhY3Rpb249Y2xpY2tSZWRpcmVjdCZkb05vdExvZ0NsaWNrPXRydWU= like these]

==3D Printed Parts==
The 4 parts are currently available as STL and 3MF files.
[https://drive.google.com/drive/folders/1_GCll42MNhbK981q99WKfd3auBnq7ENU?usp=sharing Download here]

==Software==
[https://learn.adafruit.com/adafruit-mpr121-12-key-capacitive-touch-sensor-breakout-tutorial/wiring Adafruit Tutorial]

[https://www.arduino.cc/en/software Arduino IDE]

[https://www.pjrc.com/teensy/teensyduino.html Teensyduino add-on]

[https://circuitjournal.com/arduino-serial-to-spreadsheet ArduSpreadsheet add-on]

=Assembly=

==Electronics==

==Hardware and 3D Printed Parts==

=Code=
Arduino code is written in C/C++

//include relevant libraries
#include <Wire.h>
#include "Adafruit_MPR121.h"
#include <AccelStepper.h>

//define MPR121 info
#ifndef _BV
#define _BV(bit) (1 << (bit))
#endif
Adafruit_MPR121 cap = Adafruit_MPR121();
uint16_t lasttouched = 0;
uint16_t currtouched = 0;

//define DRV8825 stuff:
#define dirPin 2
#define stepPin 3
#define motorInterfaceType 1

// Create a new instance of the AccelStepper class:
AccelStepper stepper = AccelStepper(motorInterfaceType, stepPin, dirPin);

//setup code here, to run once:
void setup() {
stepper.setMaxSpeed(1000); //set max stepper speed in steps per second
Serial.begin(9600); //start serial monitor
while (!Serial) { // needed to keep from starting too fast!
delay(10);
}
Serial.println("Electronic Esophagus test start");
// Default address is 0x5A, if tied to 3.3V its 0x5B
// If tied to SDA its 0x5C and if SCL then 0x5D
if (!cap.begin(0x5A)) {
Serial.println("MPR121 not found, check wiring?");
while (1);
}
Serial.println("MPR121 found!");
// to change threshold sensitivites (touched, released). Defaults are (12,6). Valuess from 0-255 = less-more sensitive.
cap.setThresholds(24, 12);
}

// main code here, to run repeatedly:
void loop() {
// Get the currently touched pads
currtouched = cap.touched();

for (uint8_t i=0; i<12; i++) {
// it if *is* touched and *wasnt* touched before, alert!
if ((currtouched & _BV(i)) && !(lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" touched");
// set the position to 0:
stepper.setCurrentPosition(0);
// Run the motor forward at 400 steps/second until the motor reaches 12 steps (~0.125 revolutions):
while(stepper.currentPosition() != 12)
{
stepper.setSpeed(400);
stepper.runSpeed();
}
delayMicroseconds(500);
}

// if it *was* touched and now *isnt*, alert!
if (!(currtouched & _BV(i)) && (lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" released");
stepper.setCurrentPosition(0);
stepper.stop(); // Stop as fast as possible: sets new target
stepper.runToPosition(); // Now stopped after quickstop
delayMicroseconds(500);
}
}
// reset our state
lasttouched = currtouched;
// comment out "return" line for detailed data from the sensor!
return;
// debugging info, what
Serial.print("\t\t\t\t\t\t\t\t\t\t\t\t\t 0x"); Serial.println(cap.touched(), HEX);
Serial.print("Filt: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.filteredData(i)); Serial.print("\t");
}
Serial.println();
Serial.print("Base: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.baselineData(i)); Serial.print("\t");
}
Serial.println();
// put a delay so it isn't overwhelming
delay(100);
}

=References=
Dr. D-Flo. (2017, Feb 20). DIY Syringe Pump (Food 3D Printer - Part 1) [Video]. YouTube. [https://www.youtube.com/watch?v=UHa-OKb_CiM&t=212s]

Elizalde G & Sclafani A, Flavor preferences conditioned by intragastric polycose infusions: A detailed analysis using an electronic esophagus preparation, Physiology & Behavior 47: 63-67, 1990. [https://pubmed.ncbi.nlm.nih.gov/2109327/]

Longley M et al. An open source device for operant licking in rats. PeerJ 5:e2981, 2017. [https://peerj.com/articles/2981/]

Wijen B et al. Open-source syringe pump library. PLOS ONE 9(9): e107216, 2014. [https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0107216]

=People=
Marena Bass

[[Category:Flavor Preference]]

Electronic Esophagus

2022-04-27T14:09:19Z

MBass:

Electronic Esophagus

2022-04-24T12:56:23Z

MBass: added more hardware

Electronic Esophagus

2022-04-14T20:07:21Z

MBass: Added description and more links to parts

Electronic Esophagus

2022-04-13T19:28:29Z

MBass: Typo fix

=Parts=

==Electronics==
[https://www.pjrc.com/teensy/teensyLC.html Teensy-LC]

[https://www.adafruit.com/product/1982 MPR121 Capacitive Touch Sensor]

[https://www.pololu.com/product/2133/ DRV8825 Stepper Motor Driver]

[https://www.adafruit.com/product/324 Stepper motor, NEMA-17, 200 steps/rev, 12V 350mA]

==Hardware==

==3D Printed Parts==

==Software==
[https://learn.adafruit.com/adafruit-mpr121-12-key-capacitive-touch-sensor-breakout-tutorial/wiring Adafruit Tutorial]

[https://www.arduino.cc/en/software Arduino IDE]

[https://www.pjrc.com/teensy/teensyduino.html Teensyduino add-on]

[https://circuitjournal.com/arduino-serial-to-spreadsheet ArduSpreadsheet add-on]

=Code=

//include relevant libraries
#include <Wire.h>
#include "Adafruit_MPR121.h"
#include <AccelStepper.h>

//define MPR121 info
#ifndef _BV
#define _BV(bit) (1 << (bit))
#endif
Adafruit_MPR121 cap = Adafruit_MPR121();
uint16_t lasttouched = 0;
uint16_t currtouched = 0;

//define DRV8825 stuff:
#define dirPin 2
#define stepPin 3
#define motorInterfaceType 1

// Create a new instance of the AccelStepper class:
AccelStepper stepper = AccelStepper(motorInterfaceType, stepPin, dirPin);

//setup code here, to run once:
void setup() {
stepper.setMaxSpeed(1000); //set max stepper speed in steps per second
Serial.begin(9600); //start serial monitor
while (!Serial) { // needed to keep from starting too fast!
delay(10);
}
Serial.println("Electronic Esophagus test start");
// Default address is 0x5A, if tied to 3.3V its 0x5B
// If tied to SDA its 0x5C and if SCL then 0x5D
if (!cap.begin(0x5A)) {
Serial.println("MPR121 not found, check wiring?");
while (1);
}
Serial.println("MPR121 found!");
// to change threshold sensitivites (touched, released). Defaults are (12,6). Valuess from 0-255 = less-more sensitive.
cap.setThresholds(24, 12);
}

// main code here, to run repeatedly:
void loop() {
// Get the currently touched pads
currtouched = cap.touched();

for (uint8_t i=0; i<12; i++) {
// it if *is* touched and *wasnt* touched before, alert!
if ((currtouched & _BV(i)) && !(lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" touched");
// set the position to 0:
stepper.setCurrentPosition(0);
// Run the motor forward at 400 steps/second until the motor reaches 12 steps (~0.125 revolutions):
while(stepper.currentPosition() != 12)
{
stepper.setSpeed(400);
stepper.runSpeed();
}
delayMicroseconds(500);
}

// if it *was* touched and now *isnt*, alert!
if (!(currtouched & _BV(i)) && (lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" released");
stepper.setCurrentPosition(0);
stepper.stop(); // Stop as fast as possible: sets new target
stepper.runToPosition(); // Now stopped after quickstop
delayMicroseconds(500);
}
}
// reset our state
lasttouched = currtouched;
// comment out "return" line for detailed data from the sensor!
return;
// debugging info, what
Serial.print("\t\t\t\t\t\t\t\t\t\t\t\t\t 0x"); Serial.println(cap.touched(), HEX);
Serial.print("Filt: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.filteredData(i)); Serial.print("\t");
}
Serial.println();
Serial.print("Base: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.baselineData(i)); Serial.print("\t");
}
Serial.println();
// put a delay so it isn't overwhelming
delay(100);
}

=People=
Marena Bass

[[Category:Flavor Preference]]

Electronic Esophagus

2022-04-13T19:21:27Z

MBass: edit headers

=Parts=

==Electronics==
[https://www.pjrc.com/teensy/teensyLC.html Teensy-LC]

[https://www.adafruit.com/product/1982 MPR121 Capacitive Touch Sensor]

[https://www.pololu.com/product/2133/ DRV8825 Stepper Motor Driver]

[https://www.adafruit.com/product/324 Stepper motor, NEMA-17, 200 steps/rev, 12V 350mA]

==Hardware==

==3D Printed Parts==

==Software==
[https://learn.adafruit.com/adafruit-mpr121-12-key-capacitive-touch-sensor-breakout-tutorial/wiring Adafruit Tutorial]

[https://www.arduino.cc/en/software Arduino IDE]

[https://www.pjrc.com/teensy/teensyduino.html Teensyduino add-on]

[https://circuitjournal.com/arduino-serial-to-spreadsheet ArduSpreadsheet add-on]

=Code=

//include relevant libraries
#include <Wire.h>
#include "Adafruit_MPR121.h"
#include <AccelStepper.h>

//define MPR121 info
#ifndef _BV
#define _BV(bit) (1 << (bit))
#endif
Adafruit_MPR121 cap = Adafruit_MPR121();
uint16_t lasttouched = 0;
uint16_t currtouched = 0;

//define DRV8825 stuff:
#define dirPin 2
#define stepPin 3
#define motorInterfaceType 1

// Create a new instance of the AccelStepper class:
AccelStepper stepper = AccelStepper(motorInterfaceType, stepPin, dirPin);

//setup code here, to run once:
void setup() {
stepper.setMaxSpeed(1000); //set max stepper speed in steps per second
Serial.begin(9600); //start serial monitor
while (!Serial) { // needed to keep from starting too fast!
delay(10);
}
Serial.println("Electronic Esophagus test start");
// Default address is 0x5A, if tied to 3.3V its 0x5B
// If tied to SDA its 0x5C and if SCL then 0x5D
if (!cap.begin(0x5A)) {
Serial.println("MPR121 not found, check wiring?");
while (1);
}
Serial.println("MPR121 found!");
// to change threshold sensitivites (touched, released). Defaults are (12,6). Valuess from 0-255 = less-more sensitive.
cap.setThresholds(24, 12);
}

// main code here, to run repeatedly:
void loop() {
// Get the currently touched pads
currtouched = cap.touched();

for (uint8_t i=0; i<12; i++) {
// it if *is* touched and *wasnt* touched before, alert!
if ((currtouched & _BV(i)) && !(lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" touched");
// set the position to 0:
stepper.setCurrentPosition(0);
// Run the motor forward at 400 steps/second until the motor reaches 24 steps (~0.125 revolutions):
while(stepper.currentPosition() != 12)
{
stepper.setSpeed(400);
stepper.runSpeed();
}
delayMicroseconds(500);
}

// if it *was* touched and now *isnt*, alert!
if (!(currtouched & _BV(i)) && (lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" released");
stepper.setCurrentPosition(0);
stepper.stop(); // Stop as fast as possible: sets new target
stepper.runToPosition(); // Now stopped after quickstop
delayMicroseconds(500);
}
}
// reset our state
lasttouched = currtouched;
// comment out "return" line for detailed data from the sensor!
return;
// debugging info, what
Serial.print("\t\t\t\t\t\t\t\t\t\t\t\t\t 0x"); Serial.println(cap.touched(), HEX);
Serial.print("Filt: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.filteredData(i)); Serial.print("\t");
}
Serial.println();
Serial.print("Base: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.baselineData(i)); Serial.print("\t");
}
Serial.println();
// put a delay so it isn't overwhelming
delay(100);
}

=People=
Marena Bass

[[Category:Flavor Preference]]

Electronic Esophagus

2022-04-13T19:15:39Z

MBass: /* Code */

=Raspberry PI Lickometer=

==Parts==

===Electronics===
[https://www.pjrc.com/teensy/teensyLC.html Teensy-LC]

[https://www.adafruit.com/product/1982 MPR121 Capacitive Touch Sensor]

[https://www.pololu.com/product/2133/ DRV8825 Stepper Motor Driver]

[https://www.adafruit.com/product/324 Stepper motor, NEMA-17, 200 steps/rev, 12V 350mA]

===Hardware===

===3D Printed Parts===

===Software===
[https://learn.adafruit.com/adafruit-mpr121-12-key-capacitive-touch-sensor-breakout-tutorial/wiring Adafruit Tutorial]

[https://www.arduino.cc/en/software Arduino IDE]

[https://www.pjrc.com/teensy/teensyduino.html Teensyduino add-on]

[https://circuitjournal.com/arduino-serial-to-spreadsheet ArduSpreadsheet add-on]

==Code==

//include relevant libraries
#include <Wire.h>
#include "Adafruit_MPR121.h"
#include <AccelStepper.h>

//define MPR121 info
#ifndef _BV
#define _BV(bit) (1 << (bit))
#endif
Adafruit_MPR121 cap = Adafruit_MPR121();
uint16_t lasttouched = 0;
uint16_t currtouched = 0;

//define DRV8825 stuff:
#define dirPin 2
#define stepPin 3
#define motorInterfaceType 1

// Create a new instance of the AccelStepper class:
AccelStepper stepper = AccelStepper(motorInterfaceType, stepPin, dirPin);

//setup code here, to run once:
void setup() {
stepper.setMaxSpeed(1000); //set max stepper speed in steps per second
Serial.begin(9600); //start serial monitor
while (!Serial) { // needed to keep from starting too fast!
delay(10);
}
Serial.println("Electronic Esophagus test start");
// Default address is 0x5A, if tied to 3.3V its 0x5B
// If tied to SDA its 0x5C and if SCL then 0x5D
if (!cap.begin(0x5A)) {
Serial.println("MPR121 not found, check wiring?");
while (1);
}
Serial.println("MPR121 found!");
// to change threshold sensitivites (touched, released). Defaults are (12,6). Valuess from 0-255 = less-more sensitive.
cap.setThresholds(24, 12);
}

// main code here, to run repeatedly:
void loop() {
// Get the currently touched pads
currtouched = cap.touched();

for (uint8_t i=0; i<12; i++) {
// it if *is* touched and *wasnt* touched before, alert!
if ((currtouched & _BV(i)) && !(lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" touched");
// set the position to 0:
stepper.setCurrentPosition(0);
// Run the motor forward at 400 steps/second until the motor reaches 24 steps (~0.125 revolutions):
while(stepper.currentPosition() != 12)
{
stepper.setSpeed(400);
stepper.runSpeed();
}
delayMicroseconds(500);
}

// if it *was* touched and now *isnt*, alert!
if (!(currtouched & _BV(i)) && (lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" released");
stepper.setCurrentPosition(0);
stepper.stop(); // Stop as fast as possible: sets new target
stepper.runToPosition(); // Now stopped after quickstop
delayMicroseconds(500);
}
}
// reset our state
lasttouched = currtouched;
// comment out "return" line for detailed data from the sensor!
return;
// debugging info, what
Serial.print("\t\t\t\t\t\t\t\t\t\t\t\t\t 0x"); Serial.println(cap.touched(), HEX);
Serial.print("Filt: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.filteredData(i)); Serial.print("\t");
}
Serial.println();
Serial.print("Base: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.baselineData(i)); Serial.print("\t");
}
Serial.println();
// put a delay so it isn't overwhelming
delay(100);
}

==People==
Marena Bass

[[Category:Flavor Preference]]

Electronic Esophagus

2022-04-13T19:13:24Z

MBass: /* Code */

=Raspberry PI Lickometer=

==Parts==

===Electronics===
[https://www.pjrc.com/teensy/teensyLC.html Teensy-LC]

[https://www.adafruit.com/product/1982 MPR121 Capacitive Touch Sensor]

[https://www.pololu.com/product/2133/ DRV8825 Stepper Motor Driver]

[https://www.adafruit.com/product/324 Stepper motor, NEMA-17, 200 steps/rev, 12V 350mA]

===Hardware===

===3D Printed Parts===

===Software===
[https://learn.adafruit.com/adafruit-mpr121-12-key-capacitive-touch-sensor-breakout-tutorial/wiring Adafruit Tutorial]

[https://www.arduino.cc/en/software Arduino IDE]

[https://www.pjrc.com/teensy/teensyduino.html Teensyduino add-on]

[https://circuitjournal.com/arduino-serial-to-spreadsheet ArduSpreadsheet add-on]

==Code==

//include relevant libraries

#include <Wire.h>
#include "Adafruit_MPR121.h"
#include <AccelStepper.h>

//define MPR121 info
#ifndef _BV
#define _BV(bit) (1 << (bit))
#endif

Adafruit_MPR121 cap = Adafruit_MPR121();
uint16_t lasttouched = 0;
uint16_t currtouched = 0;

//define DRV8825 stuff:
#define dirPin 2
#define stepPin 3
#define motorInterfaceType 1

// Create a new instance of the AccelStepper class:
AccelStepper stepper = AccelStepper(motorInterfaceType, stepPin, dirPin);

//setup code here, to run once:
void setup() {
stepper.setMaxSpeed(1000); //set max stepper speed in steps per second
Serial.begin(9600); //start serial monitor
while (!Serial) { // needed to keep from starting too fast!
delay(10);
}
Serial.println("Electronic Esophagus test start");
// Default address is 0x5A, if tied to 3.3V its 0x5B
// If tied to SDA its 0x5C and if SCL then 0x5D
if (!cap.begin(0x5A)) {
Serial.println("MPR121 not found, check wiring?");
while (1);
}
Serial.println("MPR121 found!");
// to change threshold sensitivites (touched, released). Defaults are (12,6). Valuess from 0-255 = less-more sensitive.
cap.setThresholds(24, 12);
}

// main code here, to run repeatedly:
void loop() {
// Get the currently touched pads
currtouched = cap.touched();

for (uint8_t i=0; i<12; i++) {
// it if *is* touched and *wasnt* touched before, alert!
if ((currtouched & _BV(i)) && !(lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" touched");
// set the position to 0:
stepper.setCurrentPosition(0);
// Run the motor forward at 400 steps/second until the motor reaches 24 steps (~0.125 revolutions):
while(stepper.currentPosition() != 12)
{
stepper.setSpeed(400);
stepper.runSpeed();
}
delayMicroseconds(500);
}

// if it *was* touched and now *isnt*, alert!
if (!(currtouched & _BV(i)) && (lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" released");
stepper.setCurrentPosition(0);
stepper.stop(); // Stop as fast as possible: sets new target
stepper.runToPosition(); // Now stopped after quickstop
delayMicroseconds(500);
}
}
// reset our state
lasttouched = currtouched;
// comment out "return" line for detailed data from the sensor!
return;
// debugging info, what
Serial.print("\t\t\t\t\t\t\t\t\t\t\t\t\t 0x"); Serial.println(cap.touched(), HEX);
Serial.print("Filt: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.filteredData(i)); Serial.print("\t");
}
Serial.println();
Serial.print("Base: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.baselineData(i)); Serial.print("\t");
}
Serial.println();
// put a delay so it isn't overwhelming
delay(100);
}

==People==
Marena Bass

[[Category:Flavor Preference]]

Electronic Esophagus

2022-04-13T19:11:33Z

MBass: /* Code */

=Raspberry PI Lickometer=

==Parts==

===Electronics===
[https://www.pjrc.com/teensy/teensyLC.html Teensy-LC]

[https://www.adafruit.com/product/1982 MPR121 Capacitive Touch Sensor]

[https://www.pololu.com/product/2133/ DRV8825 Stepper Motor Driver]

[https://www.adafruit.com/product/324 Stepper motor, NEMA-17, 200 steps/rev, 12V 350mA]

===Hardware===

===3D Printed Parts===

===Software===
[https://learn.adafruit.com/adafruit-mpr121-12-key-capacitive-touch-sensor-breakout-tutorial/wiring Adafruit Tutorial]

[https://www.arduino.cc/en/software Arduino IDE]

[https://www.pjrc.com/teensy/teensyduino.html Teensyduino add-on]

[https://circuitjournal.com/arduino-serial-to-spreadsheet ArduSpreadsheet add-on]

===Code===

//include relevant libraries

#include <Wire.h>
#include "Adafruit_MPR121.h"
#include <AccelStepper.h>

//define MPR121 info
#ifndef _BV
#define _BV(bit) (1 << (bit))
#endif

Adafruit_MPR121 cap = Adafruit_MPR121();
uint16_t lasttouched = 0;
uint16_t currtouched = 0;

//define DRV8825 stuff:
#define dirPin 2
#define stepPin 3
#define motorInterfaceType 1

// Create a new instance of the AccelStepper class:
AccelStepper stepper = AccelStepper(motorInterfaceType, stepPin, dirPin);

//setup code here, to run once:
void setup() {
stepper.setMaxSpeed(1000); //set max stepper speed in steps per second
Serial.begin(9600); //start serial monitor
while (!Serial) { // needed to keep from starting too fast!
delay(10);
}
Serial.println("Electronic Esophagus test start");
// Default address is 0x5A, if tied to 3.3V its 0x5B
// If tied to SDA its 0x5C and if SCL then 0x5D
if (!cap.begin(0x5A)) {
Serial.println("MPR121 not found, check wiring?");
while (1);
}
Serial.println("MPR121 found!");
// to change threshold sensitivites (touched, released). Defaults are (12,6). Valuess from 0-255 = less-more sensitive.
cap.setThresholds(24, 12);
}

// main code here, to run repeatedly:
void loop() {
// Get the currently touched pads
currtouched = cap.touched();

for (uint8_t i=0; i<12; i++) {
// it if *is* touched and *wasnt* touched before, alert!
if ((currtouched & _BV(i)) && !(lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" touched");
// set the position to 0:
stepper.setCurrentPosition(0);
// Run the motor forward at 400 steps/second until the motor reaches 24 steps (~0.125 revolutions):
while(stepper.currentPosition() != 12)
{
stepper.setSpeed(400);
stepper.runSpeed();
}
delayMicroseconds(500);
}

// if it *was* touched and now *isnt*, alert!
if (!(currtouched & _BV(i)) && (lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" released");
stepper.setCurrentPosition(0);
stepper.stop(); // Stop as fast as possible: sets new target
stepper.runToPosition(); // Now stopped after quickstop
delayMicroseconds(500);
}
}
// reset our state
lasttouched = currtouched;
// comment out "return" line for detailed data from the sensor!
return;
// debugging info, what
Serial.print("\t\t\t\t\t\t\t\t\t\t\t\t\t 0x"); Serial.println(cap.touched(), HEX);
Serial.print("Filt: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.filteredData(i)); Serial.print("\t");
}
Serial.println();
Serial.print("Base: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.baselineData(i)); Serial.print("\t");
}
Serial.println();
// put a delay so it isn't overwhelming
delay(100);
}

==People==
Marena Bass

[[Category:Flavor Preference]]

Electronic Esophagus

2022-04-13T19:10:21Z

MBass: /* Software */

=Raspberry PI Lickometer=

==Parts==

===Electronics===
[https://www.pjrc.com/teensy/teensyLC.html Teensy-LC]

[https://www.adafruit.com/product/1982 MPR121 Capacitive Touch Sensor]

[https://www.pololu.com/product/2133/ DRV8825 Stepper Motor Driver]

[https://www.adafruit.com/product/324 Stepper motor, NEMA-17, 200 steps/rev, 12V 350mA]

===Hardware===

===3D Printed Parts===

===Software===
[https://learn.adafruit.com/adafruit-mpr121-12-key-capacitive-touch-sensor-breakout-tutorial/wiring Adafruit Tutorial]

[https://www.arduino.cc/en/software Arduino IDE]

[https://www.pjrc.com/teensy/teensyduino.html Teensyduino add-on]

[https://circuitjournal.com/arduino-serial-to-spreadsheet ArduSpreadsheet add-on]

===Code===

//include relevant libraries

#include <Wire.h>
#include "Adafruit_MPR121.h"

#include <AccelStepper.h>

//define MPR121 info
#ifndef _BV
#define _BV(bit) (1 << (bit))
#endif

Adafruit_MPR121 cap = Adafruit_MPR121();

uint16_t lasttouched = 0;
uint16_t currtouched = 0;

//define DRV8825 stuff:
#define dirPin 2
#define stepPin 3
#define motorInterfaceType 1

// Create a new instance of the AccelStepper class:
AccelStepper stepper = AccelStepper(motorInterfaceType, stepPin, dirPin);

//setup code here, to run once:
void setup() {
stepper.setMaxSpeed(1000); //set max stepper speed in steps per second
Serial.begin(9600); //start serial monitor
while (!Serial) { // needed to keep from starting too fast!
delay(10);
}
Serial.println("Electronic Esophagus test start");
// Default address is 0x5A, if tied to 3.3V its 0x5B
// If tied to SDA its 0x5C and if SCL then 0x5D
if (!cap.begin(0x5A)) {
Serial.println("MPR121 not found, check wiring?");
while (1);
}
Serial.println("MPR121 found!");

// to change threshold sensitivites (touched, released). Defaults are (12,6). Valuess from 0-255 = less-more sensitive.
cap.setThresholds(24, 12);
}

// main code here, to run repeatedly:
void loop() {
// Get the currently touched pads
currtouched = cap.touched();

for (uint8_t i=0; i<12; i++) {
// it if *is* touched and *wasnt* touched before, alert!
if ((currtouched & _BV(i)) && !(lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" touched");
// set the position to 0:
stepper.setCurrentPosition(0);
// Run the motor forward at 400 steps/second until the motor reaches 24 steps (~0.125 revolutions):
while(stepper.currentPosition() != 12)
{
stepper.setSpeed(400);
stepper.runSpeed();
}
delayMicroseconds(500);
}

// if it *was* touched and now *isnt*, alert!
if (!(currtouched & _BV(i)) && (lasttouched & _BV(i)) ) {
Serial.print(i); Serial.print('\t'); Serial.println(" released");
stepper.setCurrentPosition(0);
stepper.stop(); // Stop as fast as possible: sets new target
stepper.runToPosition(); // Now stopped after quickstop
delayMicroseconds(500);
}
}

// reset our state
lasttouched = currtouched;

// comment out "return" line for detailed data from the sensor!
return;

// debugging info, what
Serial.print("\t\t\t\t\t\t\t\t\t\t\t\t\t 0x"); Serial.println(cap.touched(), HEX);
Serial.print("Filt: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.filteredData(i)); Serial.print("\t");
}
Serial.println();
Serial.print("Base: ");
for (uint8_t i=0; i<12; i++) {
Serial.print(cap.baselineData(i)); Serial.print("\t");
}
Serial.println();

// put a delay so it isn't overwhelming
delay(100);
}

==People==
Marena Bass

[[Category:Flavor Preference]]

Electronic Esophagus

2022-04-13T18:55:46Z

MBass: /* Electronics */

Electronic Esophagus

2022-04-13T18:53:39Z

MBass: Created page with "=Raspberry PI Lickometer= ==Parts== ===Electronics=== [https://www.pjrc.com/teensy/teensyLC.html Teensy-LC] [https://www.adafruit.com/product/1982gclid=CjwKCAjwtfqKBhBoEiwA..."

=Raspberry PI Lickometer=

==Parts==

===Electronics===
[https://www.pjrc.com/teensy/teensyLC.html Teensy-LC]

[https://www.adafruit.com/product/1982gclid=CjwKCAjwtfqKBhBoEiwAZuesiAnHL0G0795N8D4g5lvkHXnFm5FSEbK9cAypaqVWGE8EsfFCxR6VwxoCJogQAvD_BwE MPR121 Capacitive Touch Sensor]

[https://www.pololu.com/product/2133/ DRV8825 Stepper Motor Driver]

[https://www.adafruit.com/product/324 Stepper motor, NEMA-17, 200 steps/rev, 12V 350mA]

===Hardware===

===3D Printed Parts===

===Software===
[https://learn.adafruit.com/adafruit-mpr121-12-key-capacitive-touch-sensor-breakout-tutorial/wiring Adafruit Tutorial]

[https://www.arduino.cc/en/software Arduino IDE]

[https://www.pjrc.com/teensy/teensyduino.html Teensyduino add-on]

[https://circuitjournal.com/arduino-serial-to-spreadsheet ArduSpreadsheet add-on]

==People==
Marena Bass

[[Category:Flavor Preference]]

MBD Selenate and CFP 4

2021-07-10T17:53:49Z

MBass: link edit

This is the fourth replicate of selenate pre-Rx before glucose-induced flavor preference conditioning

==Data==

[https://houptlab.org/bartab/expt/?id=MBD Raw Data Link]

==People==

Marena Bass

[[Category:Flavor Preference]]

MBD Selenate and CFP 4

2021-07-10T17:50:51Z

MBass: Creating MBD Selenate and CFP 4

raw data link: https://houptlab.org/bartab/expt/?id=MBD

[[Category:Flavor Preference]]

MBC Selenate and CFP 3

2021-06-18T16:08:51Z

MBass: /* Agenda */ added 2-bottle test dates

This is the third replicate of selenate pre-Rx before glucose-induced flavor preference conditioning

== Selenate Dosage ==

To test selenate attenuation of CFNP, rats will be injected with 1 ml/kg of 0.5 mg/ml Na2SeO4 in 0.15 M NaCl, (so 0.5 mg/kg selenate), 2 hours before conditioning.

* Sodium Selenate Na2SeO4 from 2021-2-16 from [[https://www.sigmaaldrich.com/catalog/product/sial/71950?lang=en&region=US&gclid=EAIaIQobChMI0Lqo_9S38AIVWW5vBB0u2g1xEAAYASAAEgJ-9fD_BwE Millipore-Sigma S8295-25G]

* MW 188.95, CAS Number 13410-01-0, [https://pubchem.ncbi.nlm.nih.gov/substance/24899787 PubChem Substance ID 24899787]

* 0.5 mg/ml Na2Se04 is 2.65 mM, or 7.95 mOsm (assuming complete dissociation into 2 Na+ and 1 SeO4- ions)

* So OK to mix 0.5 mg Na2SeO4 in 1 ml of 0.15 M NaCl, resulting in a 308 mOsm solution that is still near isotonic (300mOsM).

==Data==

[https://houptlab.org/bartab/expt/?id=MBC Raw Data Link]

==People==

Marena Bass

==Agenda==

'''Day 0 (04/29/2021)''': At 5pm, adult male Sprague-Dawley rats (N=16) were weighed to determine their initial body weight, and their food and water was removed to begin the food and water restriction schedule.

'''Days 1-8 (04/30/2021-05/07/2021)''': Food/water restriction and training
* The rats were trained on a restricted food and water schedule for 8 days before conditioning.
* Each day at 1pm, the animals were weighed and given 23-25 grams of chow to maintain 90% body weight.
* After being weighed, animals were given access to two bottles, 0.05% saccharin and water. Initially, they were given 1hr to drink the solutions, and this was gradually reduced to 30 mins per day. Bottle positions were alternated each day.

'''Day 9 (05/08/2021)''': CS+ Conditioning
* 11 am: Animals injected (IP) with 1 mL per kg body weight of 0.15M NaCl or 0.5mg/mL sodium selenate
* 1pm: Animals given 30min access to CS+ containing 4% glucose, 0.05% saccharin and 0.05% grape or cherry flavored Kool-Aid (flavors counterbalanced)
* 23-25g food ration given after CS+ removed

'''Day 10 (05/09/2021)''': CS- Conditioning
* 11 am: Animals injected (IP) with 1 mL per kg body weight of 0.15M NaCl or 0.5mg/mL sodium selenate
* 1pm: Animals given 30min access to CS- containing, 0.05% saccharin and 0.05% Kool-Aid (whichever flavor the animal did not receive yesterday)
* Ad. lib. water returned to cages and 23-25g food ration given after CS- removed

'''Days 11-20 (05/10/2021-05/20/2021)''': 10 days of 2-bottle preference testing
* Each day, animals were given ad. lib. food and 24 hr access to both the CS- and the CS+ (without glucose) solutions. Bottle positions were alternated each day (i.e., CS+ on right for day 11 and CS+ on left for day 12, etc.)

[[Category:Flavor Preference]]

MBC Selenate and CFP 3

2021-05-11T17:14:25Z

MBass: updated agenda for days 9-11

This is the third replicate of selenate pre-Rx before glucose-induced flavor preference conditioning

== Selenate Dosage ==

To test selenate attenuation of CFNP, rats will be injected with 1 ml/kg of 0.5 mg/ml Na2SeO4 in 0.15 M NaCl, (so 0.5 mg/kg selenate), 2 hours before conditioning.

* Sodium Selenate Na2SeO4 from 2021-2-16 from [[https://www.sigmaaldrich.com/catalog/product/sial/71950?lang=en&region=US&gclid=EAIaIQobChMI0Lqo_9S38AIVWW5vBB0u2g1xEAAYASAAEgJ-9fD_BwE Millipore-Sigma S8295-25G]

* MW 188.95, CAS Number 13410-01-0, [https://pubchem.ncbi.nlm.nih.gov/substance/24899787 PubChem Substance ID 24899787]

* 0.5 mg/ml Na2Se04 is 2.65 mM, or 7.95 mOsm (assuming complete dissociation into 2 Na+ and 1 SeO4- ions)

* So OK to mix 0.5 mg Na2SeO4 in 1 ml of 0.15 M NaCl, resulting in a 308 mOsm solution that is still near isotonic (300mOsM).

==Data==

[https://houptlab.org/bartab/expt/?id=MBC Raw Data Link]

==People==

Marena Bass

==Agenda==

'''Day 0 (04/29/2021)''': At 5pm, adult male Sprague-Dawley rats (N=16) were weighed to determine their initial body weight, and their food and water was removed to begin the food and water restriction schedule.

'''Days 1-8 (04/30/2021-05/07/2021)''': Food/water restriction and training
* The rats were trained on a restricted food and water schedule for 8 days before conditioning.
* Each day at 1pm, the animals were weighed and given 23-25 grams of chow to maintain 90% body weight.
* After being weighed, animals were given access to two bottles, 0.05% saccharin and water. Initially, they were given 1hr to drink the solutions, and this was gradually reduced to 30 mins per day. Bottle positions were alternated each day.

'''Day 9 (05/08/2021)''': CS+ Conditioning
* 11 am: Animals injected (IP) with 1 mL per kg body weight of 0.15M NaCl or 0.5mg/mL sodium selenate
* 1pm: Animals given 30min access to CS+ containing 4% glucose, 0.05% saccharin and 0.05% grape or cherry flavored Kool-Aid (flavors counterbalanced)
* 23-25g food ration given after CS+ removed

'''Day 10 (05/09/2021)''': CS- Conditioning
* 11 am: Animals injected (IP) with 1 mL per kg body weight of 0.15M NaCl or 0.5mg/mL sodium selenate
* 1pm: Animals given 30min access to CS- containing, 0.05% saccharin and 0.05% Kool-Aid (whichever flavor the animal did not receive yesterday)
* Ad. lib. water returned to cages and 23-25g food ration given after CS- removed

'''Days 11-? (05/10/2021-?)''': 2-bottle preference testing
* Each day, animals were given ad. lib. food and 24 hr access to both the CS- and the CS+ (without glucose) solutions. Bottle positions were alternated each day (i.e., CS+ on right for day 11 and CS+ on left for day 12, etc.)

[[Category:Flavor Preference]]

MBC Selenate and CFP 3

2021-05-07T15:56:08Z

MBass: /* Selenate Dosage */ corrected links/calculations for selenate (was selenite before)

MBC Selenate and CFP 3

2021-05-04T18:20:07Z

MBass: added the agenda for days 0-8

This is the third replicate of selenate pre-Rx before glucose-induced flavor preference conditioning

==Data==

[https://houptlab.org/bartab/expt/?id=MBC Raw Data Link]

==People==

Marena Bass

==Agenda==

Day 0 (04/29/2021): At 5pm Rats were weighed to determine their initial body weight, and their food and water was removed to begin the food and water restriction schedule.

Days 1-8 (04/30/2021-05/07/2021) Food/water restriction and training: Adult male Sprague-Dawley rats (N=16) were trained on a restricted food and water schedule for 8 days before conditioning. Each day at 1pm, the animals were weighed and given 23-25 grams of chow to maintain 90% body weight. After being weighed, animals were given access to two bottles, 0.05% saccharin and water. Initially, they were given 1hr to drink the solutions, and this was gradually reduced to 30 mins per day. Bottle positions were alternated each day.

Day 9 (05/08/2021): CS+ Conditioning

Day 10 (05/09/2021): CS- Conditioning

Days 11-? (05/10/2021-?): 2-bottle preference testing

[[Category:Flavor Preference]]

MBC Selenate and CFP 3

2021-04-29T13:27:50Z

MBass: /* People */

This is the third replicate of selenate pre-Rx

==Data==

[https://houptlab.org/bartab/expt/?id=MBC Raw Data Link]

==People==

Marena Bass

[[Category:Flavor Preference]]