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4 protocols using ketaved

1

Anesthetic Agent Preparation Protocol

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(-)-Nicotine hydrogen tartrate salt (MP Biomedicals) was dissolved in vehicle (50:50 propylene glycol and vegetable glycerin) for a dose of 7.5 mg/mL (free base, pH 7.4). Ketamine (KetaVed, Patterson Veterinary) and xylazine (AnaSed, Patterson Veterinary) were diluted in sterile saline and simultaneously injected intraperitoneally at a dose of 100 and 10 mg/kg, respectively, for mouse perfusions.
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2

Anesthesia and Drug Administration Protocol

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Mecamylamine hydrochloride (Tocris Bioscence) was diluted in 0.9% sterile saline and injected subcutaneously at a dose of 2 mg/kg. (-)-Nicotine hydrogen tartrate salt (MP Biomedicals) was dissolved in 0.9% sterile saline (pH 7.4), and doses of nicotine refer to the free-base form. Ketamine (KetaVed, Patterson Veterinary) and xylazine (AnaSed, Patterson Veterinary) were diluted in sterile saline and simultaneously injected intraperitoneally at a dose of 100 and 10 mg/kg, respectively, for mouse perfusions. Isoflurane (IsoFlo, Abbott Laboratories) was administered as a 1–3% mixture with oxygen via inhalation.
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3

Chronic Intermittent Ethanol Exposure in Rats

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Using standard procedures from our laboratory (Morales et al. 2018 (link)), pair-housed rats were exposed to CIE for 7 or 10 consecutive days in their home cages placed inside larger, custom-build Plexiglas chambers (Triad Plastics, Winston-Salem, NC). Ethanol vapor was pumped into the chambers beginning at 9 PM (start of the light cycle) at a constant rate of 16L/min and maintained at ∼25mg/L throughout the exposure for 12h/day. Control animals were identically housed but only exposed to room-air. All rats were weighed daily. To monitor and adjust ethanol vapor levels as necessary, tail blood samples were collected periodically throughout the CIE exposure; and, blood ethanol concentrations (BECs) were determined from plasma using an alcohol dehydrogenase/NADH enzymatic assay (Carolina Liquid Chemistries, Greensboro, NC). Average BECs in the CIE animals were 259.40 ± 9.26mg/dL. In some experiments, rats received either ketamine (10mg/kg in saline, I.P.; KetaVed, Patterson Veterinary, Devens, MA) or saline (vehicle) at the onset of withdrawal, 24h before behavioral testing and electrophysiology recording. All behavioral experiments and electrophysiology recordings were conducted 24h after the last ethanol or air exposure.
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4

Electroretinography in Dark-Adapted Mice

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Scotopic and photopic electroretinograms were performed on mice that were dark-adapted overnight. Each mouse was anesthetized using intraperitoneal (IP) injections of 100 mg/kg of ketamine and 15 mg/kg xylazine (ketamine; KetaVed from Patterson Veterinary, Greeley, CO; xylazine from Patterson Veterinary, Greeley, CO). Once anesthetized, the pupils were dilated with proparacaine (1%; Akorn Inc., Ann Arbor, MI) and tropicamide (1% Tropicamide Opthalmic Solution, USP; Akorn Inc., Ann Arbor, MI or 0.5% Tropicamide Opthalmic Solution, USP, Sandoz, Princeton, NJ) eye drops, which were administered topically. Mice were placed on a heating pad under red light and function was analyzed with Diagnosys Celeris System (Diagnosys, LLC, Lowell, MA). Full field ERGs were assessed at the following stimulus intensities (0.001, 0.005, 0.01, 1, and 10 cd s/m2). Scotopic a-, b-, and c-waves were collected. Afterwards, mice were injected with reversal agent (0.5 mg/mL atipamezole, injection volume 5 μL per gram mouse weight; Patterson Veterinary, Greeley, CO) and placed individually in cages on top of heated water pads to recover.
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