Under anesthesia induced via intraperitoneal administration of a mixture of medetomidine (0.3 mg/kg BW, Domitor; ZENOAQ, Koriyama, Japan), Midazolam (4 mg/kg BW, Midazolam; SANDOZ, Tokyo, Japan), and butorphanol (5 mg/kg BW, Bettlefar; Meiji Seika Pharma, Tokyo, Japan), rats were perfused with 100 ml of 0.2% heparinized 0.1 M phosphate buffer (PB; pH 7.4), followed by 1,000 ml of 3% paraformaldehyde in 0.1 M PB through the ascending aorta, as previously reported (Oda et al., 2010 (link), 2018 (link)). In our experience, fixation with 3% paraformaldehyde occasionally leads to improved preservation of immunoreactivity for G-protein coupled receptors such as dopamine receptors, with structural preservation comparable to that of 4% paraformaldehyde. Following perfusion, the brains were removed and postfixed for 3 h in 3% paraformaldehyde in 0.1 M PB. Blocks of tissue specimens were immersed in 20% sucrose in 0.1 M PB overnight and then sectioned at a thickness of 60 μm with a freezing microtome. The sections were stored in tissue cryoprotective solution (25% glycerol and 30% ethylene glycol in 0.05 M PB) at −80°C until use.
Bettlefar
Manufactured by Meiji Seika Pharma
Sourced in Japan
The Bettlefar is a laboratory centrifuge designed for the separation of liquids and solids. It features high-speed rotation to generate centrifugal force, enabling efficient separation of sample components. The Bettlefar is a versatile piece of lab equipment suitable for a variety of applications, including sample preparation, cell pelleting, and other common laboratory procedures.
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2 protocols using bettlefar
1
Transcardial Perfusion and Brain Fixation
2
Rabbit Anesthesia Protocol for Ophthalmic Research
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This study was conducted in compliance with the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research and was approved by the Institutional Animal Care and Use Committee of Tokyo Medical University.
The right eyes of female New Zealand White rabbits aged 12 weeks and weighing 2.0 to 2.5 kg were used in the study. The anesthetics used were medetomidine hydrochloride (hereinafter medetomidine; Dolbene; Kyoritsu Seiyaku Corp., Tokyo, Japan), which is an adrenaline α2 receptor agonist; midazolam (hereinafter midazolam; Dormicum injection 10 mg; Astellas Pharma, Tokyo, Japan), which is a benzodiazepine sedative; and butorphanol tartrate (hereinafter butorphanol; Bettlefar; Meiji Seika Pharma Co. Ltd., Tokyo, Japan), which is an opioid κ receptor agonist. For general anesthesia, a mixture of medetomidine 0.15 mg/kg, midazolam 1.0 mg/kg, and butorphanol 1.5 mg/kg was injected intravenously.13 (link) For local anesthesia, oxybuprocaine ophthalmic solution 0.4% (hereinafter oxybuprocaine; Benoxil; Santen Pharmaceutical Co. Ltd., Oasaka, Japan) was used. In addition, tropicamide phenylephrine hydrochloride ophthalmic solution (Mydrin P; Santen Pharmaceutical Co. Ltd.) was used as a mydriatic agent. Euthanasia of rabbits was performed by intravenous administration of large doses of pentobarbital.
The right eyes of female New Zealand White rabbits aged 12 weeks and weighing 2.0 to 2.5 kg were used in the study. The anesthetics used were medetomidine hydrochloride (hereinafter medetomidine; Dolbene; Kyoritsu Seiyaku Corp., Tokyo, Japan), which is an adrenaline α2 receptor agonist; midazolam (hereinafter midazolam; Dormicum injection 10 mg; Astellas Pharma, Tokyo, Japan), which is a benzodiazepine sedative; and butorphanol tartrate (hereinafter butorphanol; Bettlefar; Meiji Seika Pharma Co. Ltd., Tokyo, Japan), which is an opioid κ receptor agonist. For general anesthesia, a mixture of medetomidine 0.15 mg/kg, midazolam 1.0 mg/kg, and butorphanol 1.5 mg/kg was injected intravenously.13 (link) For local anesthesia, oxybuprocaine ophthalmic solution 0.4% (hereinafter oxybuprocaine; Benoxil; Santen Pharmaceutical Co. Ltd., Oasaka, Japan) was used. In addition, tropicamide phenylephrine hydrochloride ophthalmic solution (Mydrin P; Santen Pharmaceutical Co. Ltd.) was used as a mydriatic agent. Euthanasia of rabbits was performed by intravenous administration of large doses of pentobarbital.
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