Ultramicropump3 4

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.

Eight- to twelve-week-old transgenic CaMKIIa-Cre mice were anesthetized with isofluorane. They were injected with AAV1.Syn.Flex.GCaMP6f.WPRE.SV40 obtained from the University of Pennsylvania Vector Core to express GCaMP6f in the pyramidal cells of layer 2/3 in S1. A total of 200 nl of the virus was injected using a stereotax at 1.1 posterior, 3.3 medial, 300 µm below the Bregma, using a 10-nl syringe (World Precision Instruments) fitted with a 33-gauge needle (NF33BL; World Precision Instruments), at a speed of 40 µl/min controlled via a microsyringe pump (UltraMicroPump 3–4; World Precision Instruments). Postop animals received an intraperitoneal injection of the analgesia Buprenorphin (0.2–0.5 mg/kg) which was continued for 48 h postsurgery every 8–12 h.

All animal procedures were approved by the Boston University Institutional Animal Care and Use Committee, and the experiments were carried out in accordance with the approved guidelines. Female C57BL/6 mice (Taconic, Hudson, NY), 8–12 weeks old, were first implanted with a custom imaging window, targeting striatum area (AP: +0.5, ML: 1.8, DV: −1.6). The custom imaging window was built with a stainless steel imaging cannula (OD: 3.17 mm, ID: 2.36 mm, height: 2 mm; Small Parts: B004TUE45E), a circular coverslip (size 0; OD: 3 mm, Warner Instruments: CS-3R-0), and an infusion guide cannula (26 gauge, C135GS4, Plastics, Roanoke, VA). The coverslip was fit onto the imaging cannula by using a UV-curable optical adhesive (Norland Products), and the infusion guide cannula was attached to the imaging cannula at 45 degree angle with its tip next to the coverslip. During the surgery, a custom aluminum head-plate was also attached to the skull. After recovery, animals were injected with 500 nl AAV9-Syn-GCaMP6f.WPRE.SV40 (titer: 6.6 × 1012 GC/ml, University of Pennsylvania Vector Core) with a 10 μl syringe (701 N; Hamilton Company, Reno, NV) and a 33 gauge needle (C135IS4; Plastics, Roanoke, VA). The injection was controlled by a microinfusion pump (UltraMicroPump3-4; World Precision Instruments, Sarasota, FL) at the rate of 100 nl/min.