All animal procedures were approved by the Boston University Institutional Animal Care and Use Committee, and the methods were carried out in accordance with the approved guidelines. Female C57BL/6 mice, 8–12 week old at the start of the experiments, were used in all studies (Taconic; Hudson, NY). Mice were first injected with AAV9-Syn-GCaMP6f.WPRE.SV40 virus obtained from the University of Pennsylvania Vector Core (titer ~6e12 GC/ml). 250 nL of virus was stereotaxically injected into the CA1 region (AP: –2 mm, ML: 1.4 mm, DV: –1.6 mm) using a 10 nL syringe (World Precision Instruments, Sarasota, FL) fitted with a 33 gauge needle (NF33BL; World Precision Instruments, Sarasota, FL), at a speed of 40 µl/min controlled via a microsyringe pump (UltraMicroPump3–4; World Precision Instruments, Sarasota, FL).
Upon complete recovery, animals were surgically implanted with custom imaging windows, that consisted of a stainless steel cannula (OD: 0.317 in., ID: 0.236 in., height 2 mm), adhered to a circular coverslip (size 0; OD: 3 mm) using a UV-curable optical adhesive (Norland Products). After careful aspiration of the overlying cortical tissue, using the corpus callosum as an anatomical guide, the imaging window was placed above the CA1 viral injection site. During the same surgery, a custom aluminum head-plate was attached to the skull anterior to the imaging cannula.
Upon complete recovery, animals were surgically implanted with custom imaging windows, that consisted of a stainless steel cannula (OD: 0.317 in., ID: 0.236 in., height 2 mm), adhered to a circular coverslip (size 0; OD: 3 mm) using a UV-curable optical adhesive (Norland Products). After careful aspiration of the overlying cortical tissue, using the corpus callosum as an anatomical guide, the imaging window was placed above the CA1 viral injection site. During the same surgery, a custom aluminum head-plate was attached to the skull anterior to the imaging cannula.