83 TSPO in neurodegeneration S4 gives an overview of the EAE mice used in this study, including a score of neurological signs (0 = normal, 1 = flaccid tail, 2 = impaired righting reflex, 3 = partial hindlimb paresis, 4 = complete hindlimb paresis, 5 = moribund). Spinal cord was collected from acute (aEAE)51 in the young mice, and progressive EAE (PEAE) in the 12 month old mice. Animal procedures complied with national and institutional guidelines (UK Animals Scientific Procedures Act 1986) and adhered to the 3R guidelines53. Marmoset EAE EAE was induced by subcutaneous immunization with 0.2 g of white matter homogenate emulsified in CFA in 3 adult common marmosets (Callithrix jacchus) at 4 dorsal sites adjacent to inguinal and axillary lymph nodes. Animals were monitored daily for clinical symptoms of EAE progression and assigned clinical EAE scores weekly based on extent of disability. Neurological exams were performed by a neurologist prior to each MRI scan. All animals discussed in this study are shown in Table S5. Animal #8 was treated with prednisolone for 5 days as part of a concurrent study (primary results not yet published). These animals were the first within their twin pair that showed three or more brain lesions by in vivo MRI and received corticosteroid treatment with the goal to reduce the severity of inflammation and potentially allow longer-term evaluation of the lesions. MRI analyses were performed according to previously published marmoset imaging protocols using T1, T2, T2*, and PDweighted sequences on a Bruker 7T animal magnet54. Marmosets were scanned biweekly over the course of the EAE study. Following the completion of EAE studies, the brains, spinal cords, and optic nerves excised from euthanized animals were scanned by MRI for postmortem characterization of brain lesions and previously uncharacterized spinal lesions and optic nerve lesions. Animal procedures complied with national and institutional guidelines (NIH, Bethesda, USA) SOD1G93A Female hemizygous transgenic SOD1G93A mice on 129SvHsd genetic background (n=10) and corresponding non transgenic littermates (n=9) were used. This mouse line was raised at the Mario Negri Institute for Pharmacological Research-IRCCS, Milan, Italy, derived from the line (B6SJL-TgSOD1G93A-1Gur, originally purchased from Jackson Laboratories, USA) and maintained on a 129S2/SvHsd background55. The thoracic segments of spinal cord were collected from 10- and 16-week-old mice and processed as previously described56. Briefly, anaesthetised mice were transcardially perfused with 0.1M PBS followed by 4% PFA. The spinal cord was quickly dissected out and left PFA overnight at 4°C, rinsed, and stored 24 h in 10% sucrose with 0.1% sodium azide in 0.1 M PBS at 4°C for cryoprotection, before mounting in optimal cutting temperature compound (OCT) and stored at-80°C. Procedures involving animals and their care were conducted in conformity with the following laws, regulations, and policies governing the care and use of laboratory animals: Italian Governing Law (D.lgs 26/2014; Authorization 19/2008-A issued 6 March, 2008 by Ministry of Health); Mario Negri Institutional Regulations and Policies providing internal authorization for persons conducting animal experiments; the National Institutes of Health’s Guide for the Care and Use of Laboratory Animals (2011 edition), and European Union directives and guidelines (EEC Council Directive, 2010/63/UE). APPNL-G-F For the APPNL-G-F model of AD, male and female brain tissue was obtained from 11 homozygous
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