GoKawiilAnimals
Experiments were performed in accordance with the EU guidelines for the care and use of laboratory animals, and with the guidelines of the UK Animals (Scientific Procedures) Act 1986 and subsequent amendments. Use of animals in this project was approved by the Animal Welfare and Ethical Review Body for the University of Cambridge and carried out under the terms of UK Home Office Licenses PP4353554, P9B1FBC4B and 70/7715, and the Animal Welfare and Ethical Review Body by the Icelandic Food and Veterinary Authority. All animals were maintained under a 12 h–12 h light–dark cycle with food and water supplied ad libitum, in individually ventilated cages at 20–24 °C and 45–65% relative humidity. All surgeries were performed aseptically under isoflurane anaesthesia, with standard pre- and post-operative analgesia and care. Animals were randomized into experimental groups. Experimenters were not blinded to experimental conditions during animal surgery and recovery, as lesioned animals, but not unlesioned controls, can develop side effects that require enhanced monitoring. However, subsequent tissue processing, imaging and quantification were performed blindly and validated by more than two experimenters. Experiments that did not require animal surgery were performed blindly at all stages of data collection and quantification.
Focal white matter demyelination in the CCP
EtBr model in rats
The CCP of 9–12-week-old female Sprague Dawley rats was bilaterally injected stereotaxically using a 10 μl Hamilton syringe. Four microlitres of EtBr (0.01% in saline) was injected at a rate of 1 μl min−1 to induce focal white matter demyelination. CCP coordinates were −7.3 mm (dorsal–ventral, DV); −2.3 mm (anterior–posterior, AP); ±2.6 mm (medium–lateral, ML) from lambda. In ageing experiments, CCP demyelinating lesions were induced in 18 month-old Sox10:dsRed rats.
EtBr model in mice
For mice, 3–5-month-old female C57Bl/6 mice were stereotaxically injected bilaterally with a mixture of EtBr and Dextran AlexaFluor 488 (Thermo Fisher Scientific; 1.50 μl, 0.01% and 2%, respectively, diluted in saline) at a rate of 0.25 μl per minute into the CCP; AP: −5.90 mm, ML: ±2.00 mm, DV −4.40 mm relative to bregma.
Lysolecithin model
Female Sprague Dawley rats (aged 9–12 weeks) were bilaterally injected with 2 µl of 1% lysolecithin (Sigma-Aldrich) in the CCP. Coordinates were AP: −2.3 mm; ML: ±2.6 mm, DV: −7.3 mm, relative to lambda.
Focal white matter hypoxic injury
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