For all surgeries, mice were anesthetized with 1–2% isoflurane, and placed in a stereotaxic apparatus (Kopf Instruments) on a heating pad (Harvard Apparatus). Fur was removed from the scalp, the incision site was cleaned with betadine and a midline incision was made. Sterile surgical techniques were used, and mice were injected with sustained-release buprenorphine for post-operative recovery. Mice were allowed to recover for at least two weeks after surgery before behavioural experiments.
For intracranial optogenetic experiments, virus was injected using a 33-gauge beveled needle and a 10-µl Nano-fil syringe (World Precision Instruments), controlled by an injection pump (Harvard Apparatus). Five hundred nanolitres of AAVdj-hSyn::iC++-eYFP or AAVdj-hSyn::eYFP (5×1011 vg ml−1) was injected at 150 nl per min and the syringe was left in place for at least 10 min before removal. The following coordinates were used (relative to Bregma): posterior insula (−0.58 (anterior–posterior (AP)), ±4.2 (medial–lateral (ML)), −3.85 (dorsal–ventral (DV)); mPFC (1.8 (AP), ±0.35 (ML), −2.9 (DV)). Optical fibres (0.39 NA, 200 µm; Thorlabs) were implanted 200 µm above virus injection coordinates. Fibres were secured to the cranium using Metabond (Parkell). Mice were allowed to recover for at least two weeks before behavioural testing.
For intracranial optogenetic experiments, virus was injected using a 33-gauge beveled needle and a 10-µl Nano-fil syringe (World Precision Instruments), controlled by an injection pump (Harvard Apparatus). Five hundred nanolitres of AAVdj-hSyn::iC++-eYFP or AAVdj-hSyn::eYFP (5×1011 vg ml−1) was injected at 150 nl per min and the syringe was left in place for at least 10 min before removal. The following coordinates were used (relative to Bregma): posterior insula (−0.58 (anterior–posterior (AP)), ±4.2 (medial–lateral (ML)), −3.85 (dorsal–ventral (DV)); mPFC (1.8 (AP), ±0.35 (ML), −2.9 (DV)). Optical fibres (0.39 NA, 200 µm; Thorlabs) were implanted 200 µm above virus injection coordinates. Fibres were secured to the cranium using Metabond (Parkell). Mice were allowed to recover for at least two weeks before behavioural testing.
Free full text: Click here
