Bilateral Hypothalamic AAV Delivery in Mice

Mice were randomized to receive either bilateral injections of AAV-YFP or AAV-TrkB.FL to the hypothalamus. Mice were anaesthetized with a single dose of ketamine/xylazine (100 and 20 mg kg⁻¹; i.p.) and secured via ear bars and incisor bar on a Kopf stereotaxic frame. A mid-line incision was made through the scalp to reveal the skull and two small holes were drilled into the skull with a dental drill above the injection sites (-1.2 AP, ±0.5 ML, -6.2 DV, mm from bregma). rAAV vectors (2.5 × 10⁹ genomic particles per site) were injected bilaterally into the hypothalamus at a rate of 0.1 μl minute⁻¹ using a 10 μl Hamilton syringe attached to Micro4 Micro Syringe Pump Controller (World Precision Instruments, Sarasota, FL). At the end of infusion, the syringe was slowly raised from the brain and the scalp was sutured. Animals were placed back into a clean cage and carefully monitored until recovery from anesthesia.