Materials

Sterilize the following tools via autoclave (D1 cycle, 121°C, 20 minutes): • Small scissors
• Fine tipped tweezers • Drill bits (0.7 mm and 0.9mm)
• Scalpel handle
• 2 butterfly clips

Other tools and materials (these are not sterilized): • Opthalamic ointment
• Sterile scalpel blade
• Betadyne (single use pads or liquid)
• Alcohol pads
• Cotton swabs
• Metabond (note we only use the following parts from the kit):

• Dental acrylic and liquid

Anesthetize mouse

• Place mouse in anesthetizing box until limbs lose rigidity. DO NOT leave mouse unattended; too long in the box can kill the mouse. • Quickly move mouse to stereotax and place in ear bars. IF THE MOUSE WAKES UP BEFORE SECURELY IN THE BITE BAR, use the isoflurane box to re-anesthetize. • At this point, the level of isoflurane can be lowered to 1.0-2.0. Leave him like this for ~1-2 minutes for the anesthesia level to equilibrate.
• Continuously monitor the mouse throughout surgery to ensure proper anesthesia.

Surgery part 1 - prepare skull for electrodes

• Prior to making the initial incision, pinch the mouse’s toes and tail with tweezers to confirm deep anesthesia. If a pain reflex is still present, increase isoflurane level, wait a minute and test again. • Make a single incision (1.5 cm long) in the center of the scalp, moving rostral -> caudal. If the initial incision is not long enough to expose bregma, lambda, and the target area, extend the length of the incision using surgical scissors.
• Using butterfly clips, clamp the sides of the scalp flanking the incision open to expose the skull. • Drill the craniotomy for the array. Use the foot pedal to power the drill while lowering it with the stereotax arm. Continue drilling until the drill pierces the skull, but stop before breaking through the meninges. Move the drill bit around to remove ~2mm x 2mm craniotomy, centered on your target coordinates. For dorsomedial striatum, this is -1.5mm ML, +0.5mm AP, and the array will be positioned at ~2.6mm below brain. • Drill 3 more holes at this point with the 0.9mm drill bit, 2 for skull screws, and 1 for a ground wire.

• If you are also infusing a virus in this surgery, do it at this point, into the center of the craniotomy. • To confirm that the craniotomy is clear, use a bent 27G needle tip and check for bone around the entire inside edge of the hole. Also use this needle to remove the dura, as the array wires cannot penetrate it and will get damaged if you try. If this results in bleeding, apply slight pressure on the hole with a cotton swab and wash with saline. • Insert screws into the two screw holes. This takes a little pressure to make the screws grip the holes. The bent forceps are easiest for grabbing the screws for this part.
• Cover the surface of the skull, and especially around the base of the screws with metabond. Wait for it to dry.

Surgery part 1 - Implant electrodes

• To mount the array in the stereotax arm, first connect it fully into a Plexon headstage. Attach the headstage to an electrode holder with metal clamp, and position the array wires ~1cm above the craniotomy.

• Manipulate the ground wire so it makes a “Z”, and trim it so that when the array is lowered it will enter the ground wire hole easily.

• Lower the array to the surface of the brain (watch through the scope), zero the stereotax, and then lower it ~1mm into the brain. View through the eyepiece:

• Over the next ~5-10 minutes, lower the array to 100um above your final depth in 100micron increments. Use the surgical scope to watch the wires enter the brain – if they are bending or buckling you need to remove the array and reposition it! • Once the array is 100um above your final depth, apply pink dental cement to the array, and let it run into the wires. Be careful not to actually touch the wires with your spatula. A good technique here is to apply somewhat runny cement to the side of the array connector, and let gravity pull it down into the wires. Shaving the end of a Q-tip with the scalpel blade makes a nice applicator

• Built up the cement hat until it fully supports the array, and then lower the stereotax arm the final 100um. At this point do not touch the mouse - wait until the cement is fully cured before moving to the next step. It will turn somewhat transparent when it is fully cured. • Remove the mouse from the stereotax, and turn off the isoflurane. • Put an additional layer of cement, making a smooth surface around the entire hat.
• If necessary put sutures behind or in front of the hat. • Leave the headstage in place until the very end. It is not necessary to remove it earlier, and this leaves the maximal amount of time for the cement to cure before putting strain on the connector when you remove the headstage.
• When you are ready to remove the headstage, grasp the array connector with pliers and carefully remove the headstage.

Post operative care

• Spread antibiotic cream on the wound using a cotton swab. • Transfer the mouse to the recovery cage on the heating pad. • Once the mouse’s pain reflex begins to reappear (test via tail and foot pinch), inject analgesic between the shoulder blades.
• Continue to monitor the mouse until he becomes ambulatory, at which point you can return him to the colony rack.