Midazolam

Common marmosets were euthanized by exsanguination under deep anesthesia with a mixture of medetomizine (120 μg/kg), midazolam (600 μg/kg) and butorphanol (600 μg/kg; Meiji Seika Pharma Co., Ltd., Tokyo, Japan). Specimens of the GI tract were removed, washed with ice-cold PBS and fixed overnight with Mildform 10NM (Wako Pure Chemical Industries, Ltd., Osaka, Japan). Subsequently, tissue samples were soaked in 30% sucrose in PBS overnight at 40°C and frozen in Optimal Cutting Temperature medium (Sakura Finetek, Tokyo, Japan) at -80°C.

Male Wistar rats weighing 230 g ± 10% (7 weeks old) were purchased from Tokyo Laboratory Animals Science Co., Ltd. (Tokyo, Japan), and were maintained in a climate-controlled breeding facility under 12-h automatic ambient light cycle control at the Tokyo University of Pharmacy and Life Sciences. Treatments of experimental animals and relevant protocols complied with the Regulations on Performing Animal Experiments established by the Committee of Animal Care and Welfare of Tokyo University of Pharmacy and Life Sciences (approval number: P19-08) and the ARRIVE guidelines. All animal experiments (a total of 150 rats) were performed in accordance with the relevant guidelines and regulations. Additionally, we carried out animal anesthesia and euthanasia in accordance with American Veterinary Medical Association (AVMA) Guidelines. Specifically, rats were anesthetized with a triple anesthetic combination (propofol, midazolam, and alfentanil) as intraperitoneal anesthesia and euthanized with an overdose of Isoflurane as inhalation anesthesia. Propofol, midazolam, and alfentanil were purchased from Meiji Seika Pharma Co., Ltd. (Tokyo, Japan), FUJIFILM Wako Pure Chemical Corporation (Osaka, Japan), and Nippon Zenyaku Kogyo Co., Ltd. (Fukushima, Japan), respectively. Isoflurane was purchased from Pfizer Inc. (NY, USA).