INTRACEREBRAL CANNULATION
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GENERAL ITEMS ALZET Osmotic Pumps* ALZET Brain Infusion Kit* Loctite 454 Cyanoacrylate* Homeothermic pad Sterile drape Sterile gauze, cotton swabs Hair clippers (#40 blade) 70 % ethanol Betadine scrub Glass bead sterilizer Ocular lubricant SURGICAL TOOLS - AutoClip Wound Clips* - AutoClip Applier* - AutoClip Remover* - Cannula holder 1 or 2* - Scalpel handle #3 - Scalpel blade #15 - Brown Adson forceps - Hemostat (straight tip) - Stereotaxic apparatus - Cautery (fine tip, low heat) - Screwdriver - Screws (stainless steel, 1/8 inch) - Dremel tool or surgical drill - Carbide drill burs, fine round tip (e.g. HP-1, or HP-2, REF14823, S White Burs) * Available from DURECT Corp (877-922-5938) alzetcs@durect.com Pump Preparation (refer to the package insert for complete filling instructions) - Fill the empty ALZET pump with your vehicle or drug solution using a syringe and filling tube according to the procedure listed in the package insert included in your box of pumps. - Insert the flow moderator into the filled ALZET pump until the cap or flange is flush with the top of the pump. - Prime the filled pumps in sterile saline at 37°C. - Refer to your package insert for the appropriate period of priming. Most pumps require at least an overnight priming period. Anesthesia Anesthesia is required for surgical implantation of ALZET pumps. - Anesthetize the animal using either an inhalable (i.e., Isoflurane) or injectable (i e., Xylazine® and Ketamine®, or sodium pentobarbital) anesthetic. The use of inhalation anesthetics, such as Isoflurane, is highly recommended. It supplies supplemental oxygen during periods of respiratory depression and provides for rapid anesthetic recovery. Surgical Preparation - Apply ocular lubricant to the eye. - Shave the area centered over the incision site and wash the scalp. Surgical Procedure - Place the head of the animal securely and straight in the stereotaxic apparatus with the tooth bar set at an appropriate level as determined by the same stereotaxic atlas used to establish the cannula coordinates. - Starting slightly behind the eyes, make a midline, sagittal incision about 2.5 cm long and expose the skull. With the rounded end of a spatula, lightly scrape the exposed skull area and pat it dry. Scraping should remove the periosteal connective tissue adhering to the skull. This enables good adhesion of the cyanoacrylate adhesive, which is later used to secure the cannula. Remove all blood with cotton applicators and stop any remaining bleeding with the low-heat cautery device. - Identify the bone suture junctions bregma and lambda. With these as reference points, determine and mark the location for cannula placement using the stereotaxic apparatus. Drill a hole through the skull at the marked, stereotaxically correct location, being careful not to penetrate the dura. This hole will receive the cannula. - Clean and completely dry the skull area where the cannula will be placed by scraping with the scalpel blade or cotton applicators. Additional bleeding should be stopped by cauterizing blood vessels. - Insert the L-shaped cannula, which is attached by tubing to the ALZET pump, through the skull. To facilitate precise placement of the cannula, the plastic tab located on top of the cannula can be attached to the stereotaxic apparatus. Alternatively, this tab can be used to insert the cannula by hand. DURECT Corporation | 10260 Bubb road | Cupertino CA, 95014-4166 T: 408.367.4036 F: 408.865.1406 Toll Free 800.692.2990 WWW.ALZET.COM | ALZET@DURECT.COM alzet OSMOTIC PUMPS

INTRACEREBRAL CANNULATION ALZET SURGICAL INSTRUCTION SHEET placed with the delivery port pointing toward the cannula site. When the pump is properly placed, the catheter should have a generous amount of length to permit free motion of the animal's head and neck. After the cannula is firmly set in place, the plastic tab is easily removed with a heated scalpel, leaving the low- profile cannula in place. A small amount of cement can be applied over the cannula to provide an even smoother surface, conducive to wound healing (not necessary with cyanoacrylate adhesive). - After the cement has set,...