For stereotaxis surgeries, mice were anesthetized with 1.5%–2.0% isoflurane, and placed in a stereotaxic apparatus (Kopf Instruments) on a heating pad. The fur was cut from the scalp and a midline incision was made. 3% hydrogen peroxide was applied to the skull, and a craniotomy was made above the injection site. Virus was injected using a 33-gauge beveled needle and a 10 μL Hamilton syringe (World Precision Instruments), controlled by an injection pump (Harvard Apparatus). 1,000 nL of virus was injected at 150 nL min−1. For FLiCRE recordings, a 400-μm core diameter, 0.39-NA optical fiber (Thorlabs) was implanted above the VTA, mPFC, or NAc. Implants were secured to the skull using dental adhesive (Parkell, C&B metabond). The following coordinates (in mm relative to bregma) were used for viral injections: VTA −3.2 A/P, 0.35 M/L, −4.6 D/V; mPFC +1.98 A/P, 0.35 M/L, −2.25 D/V; NAc +1.15 A/P, 1.6 M/L, −4.6 D/V. Fiber implants were placed at the same A/P and M/L locations, but 0.2 mm more dorsally. The virus titers, dilutions, and volumes used for each experiment are listed in Table S2 .
0.39 na optical fiber
Manufactured by Thorlabs
The 0.39 NA optical fiber is a multimode fiber with a numerical aperture of 0.39. It is designed to efficiently transmit light over short to medium distances. The fiber features a core diameter of 105 microns and a cladding diameter of 125 microns. This product is suitable for general-purpose optical applications where high light collection and transmission are required.
Automatically generated - may contain errors
4 protocols using 0.39 na optical fiber
1
Stereotactic Surgery Protocol for Mice
2
Inhibition of Cerebellar Interneurons
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CKit-Cre mice (generous gift from J. Christie) were used for in vivo inhibition of molecular layer interneurons. Mice were injected with 250 nl of AAV9-Ef1a-DIO eNpHR 3.0-EYFP (Addgene) at nine sites in the cerebellum and in vivo recordings were performed 2-3 weeks later. A 647 nm laser (Opto Engine, Midvale, UT) delivered via a 400 μm 0.39 NA optical fiber (Thorlabs, Newton, NJ) was focused to a spot of 2 mm2 centered over the recording site at an estimated power density of (40 mW/mm2). A silicon probe was lowered into the brain (P-series, Cambridge Neurotech, Cambridge, England), to find pairs of neighboring PCs within Halo-expressing regions. Once a pair was located, the posterior vermis was illuminated for 5 s on and 5s off for at least 30 minutes. Recorded PC pairs were screened for an increase in activity in response to inhibition of MLIs. For analysis, complex spikes were sorted into laser on trials and laser off trials, and otherwise analyzed as previously discussed.
3
Fabrication of Implantable Optical Ferrules
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Custom-made implantable optical ferrules were constructed from 2.5 mm long 270 μm Ø ceramic stick ferrules (Thorlabs) based on a previously described design65 (link). Briefly, a 200 μm Ø 0.39 NA optical fiber (Thorlabs) was stripped of its protective coating and cleaved. Next, cyanoacrylate adhesive (Loctite) was applied to the concave end of the ferrule through which the cleaved fiber segment was threaded. After wiping off the excess glue, the ferrules were allowed to cure for 30 min at room temperature. The ferrule connector was polished using a polishing disk and increasingly fine grades of polishing paper (Thorlabs), with frequent inspection to ensure transmission quality. Once polished, the free end of the fiber was scored and cleaved to 5 mm in length.
4
Inhibition of Cerebellar Interneurons
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