ICV Cannulation
For other stereotaxic coordinates, see also Amygdala, JA Lateral Hypothalamus Lesions.
Lateral Ventricular Cannula:
1.2 mm caudal to bregma
1.5 mm lateral to the midline
4 mm below the skull surface
Method
Each rat was stereotaxically implanted with a 22-gauge, stainless steel, guide cannula (Plastics One, Roanoke, VA) aimed towards the lateral cerebral ventricle (1.2 mm caudal to bregma, 1.5 mm lateral to the midline, and 4 mm below the skull surface; Houpt 1998 PMID 9437750). Guide cannulas were held in place with dental acrylic bonded to stainless steel screws anchored to the skull. An obdurator was inserted into each guide cannula and remained in place except during injections when it was removed and replaced with an injector that extended 1.0 mm beyond the tip of the guide cannula.
Test
To test the patency and placement of the cannula in the lateral ventricle, water-replete rats were injected with 100 ng human angiotensin II (Ang II; Sigma Chemical Co, St Louis, MO) dissolved in 5 µl volume of saline (0.15 M). The volume of all icv injections was 5 µl, delivered over 30s with a handheld 50 µl syringe (Hamilton Co, Reno, NV). Immediately after the ICV injection, rats were returned to their home cage with access to a water bottle. The latency to drink was recorded; rats that failed to drink within 2 minutes of the angiotensin II injection were dropped from the study. Cannula placements were also verified postmortem by sectioning through the brain. ICV injections of 5 µl volume were given by hand to lightly restrained rats using a 50 µl Hamilton microsyringe.
Fourth Ventricular Cannula:
3.2 mm caudal to lambda
0 mm lateral to the midline
7.2 mm below the skull surface
Method
Under isoflurane anesthesia, each rat was stereotaxically implanted with a single 22-gauge stainless steel guide cannula (model C315G; Plastics One, Inc., Roanoke, VA) aimed towards the fourth ventricle (4V). The tip of the guide cannula was positioned on the midline 3.2 mm caudal to lambda and 7.2 mm below the skull surface (stereotaxic coordinates taken from (Baird et al. 2009 PMID 19008313)). Guide cannulas were held in place with dental acrylic bonded to stainless steel screws anchored to the skull. A 26-gauge obdurator extending 1.0 mm beyond the guide cannula was inserted into each guide cannula and remained in place except during injections when it was removed and replaced with a 26-gauge injector that extended 1.0 mm beyond the tip of the guide cannula.
Test
To test the patency and placement of the cannula in the lateral ventricle, water-replete rats were injected with XXX ng bombesin (California Peptides) dissolved in 5 µl volume of saline (0.15 M). The volume of all icv injections was 5 µl, delivered by hand over 30s to lightly restrained rats using a 50 µl Hamilton microsyringe (Hamilton Co, Reno, NV). Immediately after the ICV injection, rats were returned to their home cage with access to a water bottle. The latency to initiate and duration of grooming was recorded; rats that failed to groom within 2 minutes of the bombesin injection or groomed for less than XXX minutes were dropped from the study. Cannula placements were also verified postmortem by sectioning through the brain.
Supplies
Plastics One Cannulas and Screws
- Plastics One
- 6591 Merriman Rd
- Roanoke VA 24018
- Tel: 540-772-7950
- Fax: 540-777-5900
Check prices on annual price list.
22 Gauge guide cannulas for ventricular injections:
Use C313 components, which provide 28 gauge dummy and injectors. e.g., for guide cannula(G) projecting 5 mm below skull surface, with large-cap dummy cannulas (DC) and injectors (I) projecting 1 mm beyond tip of the guide cannula:
- C313G/SPC 5 mm
- C313DC/SPC(LG) for 5 mm C313G with 1 mm projection
- C313I/SPC for 5 mm C313G with 1 mm projection
26 Gauge guide cannulas for site-specific injections
Use C315 components, which provide 33 gauge dummy and injectors. e.g., for guide cannula(G) projecting 8 mm below skull surface, with small-cap dummy cannulas (DC) and injectors (I) projecting 0.5 mm beyond tip of the guide cannula:
- C315G/SPC 8 mm
- C315DC/SPC(SM) for 8 mm C315G with 0.5 mm projection
- C315I/SPC for 8 mm C315G with 0.5 mm projection
Screws for skull
We use screws of 0-80 size with shaft length 3/32 inches (2.4 mm). These are a little long, but they are stronger and easier to work with than 1/16 inch (1.6 mm) long screws. Also need the small drill pit.
- 0-80 X 3/32 (packages of 100)
- D#56 1.18 mm Drill Bit for 0-80 screws
Other supplies needed - more detail to come
Pin Vise for hand drilling the screws
Most pin vises are nickel plated, and so cannot be autoclaved but must be sterilized e.g. with 70% EtOH.
- VWR 63041-530 Pin Vise - Double-Ended, stainless steel
Little tubing for washer around top of guide cannula?
Superglue
Size of screwdriver? source of SS screw drivers?
Trephin drill bits
Hamilton microsyringe and needle
PE-50 tubing to connect with injector
Syringe Pump
New Truliner Denture Lining Standard Kit, Pink
From either Henry Schein Dental, #2502527EZ, $105.29, or, Bosworth #0921970 $45.00
- Harry J. Bosworth Company
- 7227 North Hamlin Avenue
- Skokie, Illinois 60076 USA
- Toll-Free: 800-323-4352
- Phone: 847-679-3400
- Fax: 847-679-2080
- E-Mail: HJBinfo@bosworth.com