Pentobarbital sodium

All animal experiments in this study were approved by Research Ethics Committee of Sun Yat-sen University and conformed to Guide for the Care and Use of Laboratory Animals (NIH Publication No. 85-23, revised 1996). SD rats (male, weighting 185-225g, SPF grade, Certification No. 44008500011930) were from the Experimental Animal Center of Sun Yat-sen University (Guangzhou, China). Sodium pentobarbital (40 mg/kg, Merck) was used for rat anesthetization before surgery, and then connected the noninvasive tracheal intubation to the ventilator under sterile conditions. The abdominal aorta above the kidney was exposed through a midline incision of the abdomen, and sutured twice with 6-0 silk sutures at the suprarenal level, tightly surrounding the aorta and a 22-gauge needle, the needle was then extracted to produce a 65-75% contraction. A similar procedure was performed in the Sham group, but without the aortic banding. 3-Aminobenzamide (3AB) (MedChem Express) was injected intraperitoneally (20 mg/kg, twice daily) starting the week after AAC surgery for 7 weeks. At the end of experiment, all rats were euthanized by injecting Sodium pentobarbital (150 mg/kg, Merck) intraperitoneally for deep anaesthesia, followed by decapitation.

The in vivo study was performed in accordance with the standard animal study guideline of Zhejiang University, and all experimental protocols were approved by the Zhejiang University Ethics Committee (ZJU2014-1-05-093). This study applied 16 New Zealand white rabbits of 2.8 ± 0.2 kg in weight (half males and half females). The procedure was similar to our previous study2 . Surgery was performed under general anesthesia by intravenously injecting of pentobarbital sodium (30 mg/kg, Sigma). The animal was placed in a prone position and the cranium was shaved and disinfected with povodine iodine. A longitudinal incision was made along the midline from nasal bone to the occipital protuberance and the skin flap was elevated to expose the cranial region. Once the periosteum was removed, four circular bone defects (8 mm in diameter) were created with a dental trephine bur (Fig. 1D). The defects were at least 5 mm away from each other. The four circular defects were randomly filled with four groups of scaffolds and the incision was closed with suture. The rabbits were euthanized with injection of pentobarbital sodium overdose (120 mg/kg, Sigma) 6 or 12 weeks postoperatively. The cranial bones were harvested for further analyses.

All experimental procedures involving animals were performed in compliance with Institutional Animal Care guideline of Nantong University, and were ethically approved by the Administration Committee of Experimental Animals, Jiangsu Province, China. Thirty adult, female Sprague-Dawley (SD) rats (~200 g) were underwent surgery of sciatic nerve crush injury as previously described^{25 (link)}. Briefly, the rats were anaesthetized by an intraperitoneal injection of complex narcotics (42 mg/kg magnesium sulfate (MilliporeSigma, Burlington, MA, USA), 85 mg/kg tri-chloroacetaldehyde monohydrate (MilliporeSigma) and 17 mg/kg sodium pentobarbital (MilliporeSigma), and the sciatic nerve was exposed by a small incision. The left sciatic nerve at 10 mm above the bifurcation into the tibial and common fibular nerves was crushed with a pair of forceps at a force of 54 N for 3 times (a period of 10 s for each time). The animals were then randomly divided into 6 groups, and the lumbar (L)4–5 DRGs were collected at 0 hour (h), 3 h, 9 h, 1 day (d), 4 d and 7 d after surgery respectively, and stored at −80 °C.

Institute of Cancer Research (ICR) mice (male; 8 weeks old) were provided by Shanghai SLAC Laboratory Animal Co. (Shanghai, China) and provided with free access to food and water in a controlled temperature of 23–25 °C. All animal protocols were approved by the Ethics Committee of the Experimental Animals of Shanghai Seventh People’s Hospital (approval no. 2020-AR-053). Mice were anesthetized using sodium pentobarbital (i.p., 30 mg/kg; MilliporeSigma, Burlington, MA) and then subjected to a single intratracheal dose of BLM (3 mg/kg; Selleck Chemicals, Houston, TX) diluted in 50 µL sterile saline. Mice that had been intratracheally administrated 50 µL sterile saline served as the control. In the baicalein group, mice received baicalein orally (100 mg/kg/day; MilliporeSigma, Burlington, MA); the dose was based on a previous study, which showed that baicalein (p.o. 100 mg/kg/day) markedly mitigated BLM-induced lung fibrosis (Gao et al. 2013 (link)).

Animal procedures were approved by the Shanghai Jiao Tong University Institutional Review Board, and conformed to the Association for Research in Vision and Ophthalmology Statement for the Use of Animals in Ophthalmic and Vision Research. Wild-type and Cyp4v3 knockout animals were general anesthetized with 1% sodium pentobarbital (Millipore Sigma, Burlington, MA, USA) intraperitoneally, and were topically anesthetized with benoxinate HCl 0.4% (Santen Pharmaceuticals, Osaka, Japan) drops. We took fundus images and OCT images with a Micron IV System (Phoenix Research Laboratories, Pleasanton, CA, USA).