This work has been supported by Ministero dell&#39;Istruzione, dell&#39;Universit&#224; e della Ricerca - MIUR project Dipartimenti di Eccellenza 2018-2022 and 2023-2027 to Department of Neuroscience Rita Levi Montalcini; Cavalieri-Ottolenghi Foundation, Orbassano, Italy. BB was fellow of INFRA-P, Piedmont Region (n.378-35) (2022-2023) and PRIN 2020 - 20203AMKTW. We thank Fondazione per la Ricerca Biomedica Onlus (FORB) for the support. The publication fees have been supported by the kind donation of Distretto Rotaract 2031, and particularly, Rotaract Club Torino Nord-Est. We thank Elaine Miller for the proofreading of our manuscript.

Induction and Evaluation of Experimental Autoimmune Encephalomyelitis in Mouse

<ol>
	<li>Thompson, A. J., Baranzini, S. E., Geurts, J., Hemmer, B., Ciccarelli, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Multiple+sclerosis.">Multiple sclerosis.</a> Lancet. 391 (10130), 1622-1636 (2018).</li><li>Gold, S. M., Willing, A., Leypoldt, F., Paul, F., Friese, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sex+differences+in+autoimmune+disorders+of+the+central+nervous+system.">Sex differences in autoimmune disorders of the central nervous system.</a> Semin immunopathol. 41 (2), 177-188 (2019).</li><li>Smith, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Animal+models+of+multiple+sclerosis.">Animal models of multiple sclerosis.</a> Curr Protoc. 1 (6), 185 (2021).</li><li>Procaccini, C., De Rosa, V., Pucino, V., Formisano, L., Matarese, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Animal+models+of+Multiple+Sclerosis.">Animal models of Multiple Sclerosis.</a> Eur J Pharmacol. 759, 182-191 (2015).</li><li>Torre-Fuentes, L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+models+of+demyelination+and+remyelination.">Experimental models of demyelination and remyelination.</a> Neurologia. 35 (1), 32-39 (2020).</li><li>Kipp, M., Nyamoya, S., Hochstrasser, T., Amor, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Multiple+sclerosis+animal+models:+a+clinical+and+histopathological+perspective.">Multiple sclerosis animal models: a clinical and histopathological perspective.</a> Brain Pathol. 27 (2), 123-137 (2017).</li><li>Racke, M. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+autoimmune+encephalomyelitis+(EAE).">Experimental autoimmune encephalomyelitis (EAE).</a> Curr Protoc Neurosci. , (2001).</li><li>Constantinescu, C. S., Farooqi, N., O'Brien, K., Gran, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+autoimmune+encephalomyelitis+(EAE)+as+a+model+for+multiple+sclerosis+(MS).">Experimental autoimmune encephalomyelitis (EAE) as a model for multiple sclerosis (MS).</a> Br J Pharmacol. 164 (4), 1079-1106 (2011).</li><li>Montarolo, F., Perga, S., Martire, S., Bertolotto, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nurr1+reduction+influences+the+onset+of+chronic+EAE+in+mice.">Nurr1 reduction influences the onset of chronic EAE in mice.</a> Inflamm Res. 64 (11), 841-844 (2015).</li><li>Montarolo, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+isoxazolo-pyridinone+7e,+a+potent+activator+of+the+Nurr1+signaling+pathway,+on+experimental+autoimmune+encephalomyelitis+in+mice.">Effects of isoxazolo-pyridinone 7e, a potent activator of the Nurr1 signaling pathway, on experimental autoimmune encephalomyelitis in mice.</a> PLoS One. 9 (9), 108791 (2014).</li><li>Furlan, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Analysis+of+the+gadolinium+retention+in+the+experimental+autoimmune+encephalomyelitis+(EAE)+murine+model+of+multiple+sclerosis.">Analysis of the gadolinium retention in the experimental autoimmune encephalomyelitis (EAE) murine model of multiple sclerosis.</a> J Trace Elem Med Biol. 68, 126831 (2021).</li><li>Desole, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Engineering,+characterization,+and+biological+evaluation+of+an+antibody+targeting+the+HGF+receptor.">Engineering, characterization, and biological evaluation of an antibody targeting the HGF receptor.</a> Front Immunol. 12, 775151 (2021).</li><li>Voskuhl, R. R., MacKenzie-Graham, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chronic+experimental+autoimmune+encephalomyelitis+is+an+excellent+model+to+study+neuroaxonal+degeneration+in+multiple+sclerosis.">Chronic experimental autoimmune encephalomyelitis is an excellent model to study neuroaxonal degeneration in multiple sclerosis.</a> Front Mol Neurosci. 15, 1024058 (2022).</li><li>McCombe, P. A., Greer, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+biological+sex+and+pregnancy+in+experimental+autoimmune+encephalomyelitis:+It's+complicated.">Effects of biological sex and pregnancy in experimental autoimmune encephalomyelitis: It's complicated.</a> Front Immunol. 13, 1059833 (2022).</li><li>Ascherio, A., Munger, K. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epidemiology+of+multiple+sclerosis:+from+risk+factors+to+prevention-an+update.">Epidemiology of multiple sclerosis: from risk factors to prevention-an update.</a> Semin Neurol. 36 (2), 103-114 (2016).</li><li>Spence, R. D., Voskuhl, R. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neuroprotective+effects+of+estrogens+and+androgens+in+CNS+inflammation+and+neurodegeneration.">Neuroprotective effects of estrogens and androgens in CNS inflammation and neurodegeneration.</a> Front Neuroendocrinol. 33 (1), 105-115 (2012).</li><li>Laffont, S., Garnier, L., L&#233;lu, K., Gu&#233;ry, J. -. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Estrogen-mediated+protection+of+experimental+autoimmune+encephalomyelitis:+Lessons+from+the+dissection+of+estrogen+receptor-signaling+in+vivo.">Estrogen-mediated protection of experimental autoimmune encephalomyelitis: Lessons from the dissection of estrogen receptor-signaling in vivo.</a> Biomed J. 38 (3), 194-205 (2015).</li><li>Avila, M., Bansal, A., Culberson, J., Peiris, A. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+role+of+sex+hormones+in+multiple+sclerosis.">The role of sex hormones in multiple sclerosis.</a> Eur Neurol. 80 (1-2), 93-99 (2018).</li><li>Collongues, N., Patte-Mensah, C., De Seze, J., Mensah-Nyagan, A. -. G., Derfuss, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Testosterone+and+estrogen+in+multiple+sclerosis:+from+pathophysiology+to+therapeutics.">Testosterone and estrogen in multiple sclerosis: from pathophysiology to therapeutics.</a> Expert Rev Neurother. 18 (6), 515-522 (2018).</li><li>Kilkenny, C., Browne, W. J., Cuthill, I. C., Emerson, M., Altman, D. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Improving+bioscience+research+reporting:+the+ARRIVE+guidelines+for+reporting+animal+research.">Improving bioscience research reporting: the ARRIVE guidelines for reporting animal research.</a> PLoS Biol. 8 (6), 1000412 (2010).</li><li>Shaw, M. K., Zhao, X., Tse, H. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Overcoming+unresponsiveness+in+experimental+autoimmune+encephalomyelitis+(EAE)+resistant+mouse+strains+by+adoptive+transfer+and+antigenic+challenge.">Overcoming unresponsiveness in experimental autoimmune encephalomyelitis (EAE) resistant mouse strains by adoptive transfer and antigenic challenge.</a> J Vis Exp. (62), e3778 (2012).</li><li>Bittner, S., Afzali, A. M., Wiendl, H., Meuth, S. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Myelin+oligodendrocyte+glycoprotein+(MOG35-55)+induced+experimental+autoimmune+encephalomyelitis+(EAE)+in+C57BL/6+mice.">Myelin oligodendrocyte glycoprotein (MOG35-55) induced experimental autoimmune encephalomyelitis (EAE) in C57BL/6 mice.</a> J Vis Exp. (86), (2014).</li><li>Downton, P., Early, J. O., Gibbs, J. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Circadian+rhythms+in+adaptive+immunity.">Circadian rhythms in adaptive immunity.</a> Immunology. 161 (4), 268-277 (2020).</li><li>McLean, A. C., Valenzuela, N., Fai, S., Bennett, S. A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Performing+vaginal+lavage,+crystal+violet+staining,+and+vaginal+cytological+evaluation+for+mouse+estrous+cycle+staging+identification.">Performing vaginal lavage, crystal violet staining, and vaginal cytological evaluation for mouse estrous cycle staging identification.</a> J Vis Exp. (67), e4389 (2012).</li><li>Rahn, E. J., Iannitti, T., Donahue, R. R., Taylor, B. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sex+differences+in+a+mouse+model+of+multiple+sclerosis:+neuropathic+pain+behavior+in+females+but+not+males+and+protection+from+neurological+deficits+during+proestrus.">Sex differences in a mouse model of multiple sclerosis: neuropathic pain behavior in females but not males and protection from neurological deficits during proestrus.</a> Biol Sex Differ. 5 (1), 4 (2014).</li><li>Bonaldo, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+perinatal+exposure+to+bisphenol+A+or+S+in+EAE+model+of+multiple+sclerosis.">Effects of perinatal exposure to bisphenol A or S in EAE model of multiple sclerosis.</a> Cell Tissue Res. 392 (2), 467-480 (2023).</li><li>vanden Berg, R., Laman, J. D., van Meurs, M., Hintzen, R. Q., Hoogenraad, C. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rotarod+motor+performance+and+advanced+spinal+cord+lesion+image+analysis+refine+assessment+of+neurodegeneration+in+experimental+autoimmune+encephalomyelitis.">Rotarod motor performance and advanced spinal cord lesion image analysis refine assessment of neurodegeneration in experimental autoimmune encephalomyelitis.</a> J Neurosci Methods. 262, 66-76 (2016).</li><li>Bolton, C., Smith, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Defining+and+regulating+acute+inflammatory+lesion+formation+during+the+pathogenesis+of+multiple+sclerosis+and+experimental+autoimmune+encephalomyelitis.">Defining and regulating acute inflammatory lesion formation during the pathogenesis of multiple sclerosis and experimental autoimmune encephalomyelitis.</a> CNS Neurol Disord Drug Targets. 14 (7), 915-935 (2015).</li><li>Glaser, J. R., Glaser, E. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neuron+imaging+with+Neurolucida--a+PC-based+system+for+image+combining+microscopy.">Neuron imaging with Neurolucida--a PC-based system for image combining microscopy.</a> Comput Med Imaging Graph. 14 (5), 307-317 (1990).</li><li>Kennedy, H. S., Puth, F., Van Hoy, M., Le Pichon, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+method+for+removing+the+brain+and+spinal+cord+as+one+unit+from+adult+mice+and+rats.">A method for removing the brain and spinal cord as one unit from adult mice and rats.</a> Lab Anim (NY). 40 (2), 53-57 (2011).</li><li>Watson, C., Paxinos, G., Kayalioglu, G., Heise, C., Watson, C., Paxinos, G., Kayalioglu, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chapter+16+-+Atlas+of+the+mouse+spinal+cord.">Chapter 16 - Atlas of the mouse spinal cord.</a> The Spinal. , 308-379 (2009).</li><li>Khan, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Suppression+of+TRPV1/TRPM8/P2Y+nociceptors+by+withametelin+via+downregulating+MAPK+signaling+in+mouse+model+of+vincristine-induced+neuropathic+pain.">Suppression of TRPV1/TRPM8/P2Y nociceptors by withametelin via downregulating MAPK signaling in mouse model of vincristine-induced neuropathic pain.</a> Int J Mol Sci. 22 (11), 6084 (2021).</li><li>Bergamaschi, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prognostic+factors+in+multiple+sclerosis.">Prognostic factors in multiple sclerosis.</a> Int Rev Neurobiol. 79, 423-447 (2007).</li><li>Harbo, H. F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genes+in+the+HLA+class+I+region+may+contribute+to+the+HLA+class+II-associated+genetic+susceptibility+to+multiple+sclerosis.">Genes in the HLA class I region may contribute to the HLA class II-associated genetic susceptibility to multiple sclerosis.</a> Tissue Antigens. 63 (3), 237-247 (2004).</li><li>Ryan, L., Mills, K. H. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sex+differences+regulate+immune+responses+in+experimental+autoimmune+encephalomyelitis+and+multiple+sclerosis.">Sex differences regulate immune responses in experimental autoimmune encephalomyelitis and multiple sclerosis.</a> Eur J Immunol. 52 (1), 24-33 (2022).</li><li>Lasrado, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanisms+of+sex+hormones+in+autoimmunity:+focus+on+EAE.">Mechanisms of sex hormones in autoimmunity: focus on EAE.</a> Biol Sex Differ. 11 (1), 50 (2020).</li><li>Hofstetter, H. H., Shive, C. L., Forsthuber, T. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pertussis+toxin+modulates+the+immune+response+to+neuroantigens+injected+in+incomplete+Freund's+adjuvant:+induction+of+Th1+cells+and+experimental+autoimmune+encephalomyelitis+in+the+presence+of+high+frequencies+of+Th2+cells.">Pertussis toxin modulates the immune response to neuroantigens injected in incomplete Freund's adjuvant: induction of Th1 cells and experimental autoimmune encephalomyelitis in the presence of high frequencies of Th2 cells.</a> J Immunol. 169 (1), 117-125 (2002).</li><li>Maria, Z., Turner, E., Agasing, A., Kumar, G., Axtell, R. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pertussis+toxin+inhibits+encephalitogenic+T-cell+infiltration+and+promotes+a+B-cell-driven+disease+during+Th17-EAE.">Pertussis toxin inhibits encephalitogenic T-cell infiltration and promotes a B-cell-driven disease during Th17-EAE.</a> Int J Mol Sci. 22 (6), 2924 (2021).</li><li>Krementsov, D. N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Studies+in+experimental+autoimmune+encephalomyelitis+do+not+support+developmental+bisphenol+a+exposure+as+an+environmental+factor+in+increasing+multiple+sclerosis+risk.">Studies in experimental autoimmune encephalomyelitis do not support developmental bisphenol a exposure as an environmental factor in increasing multiple sclerosis risk.</a> Toxicol Sci. 135 (1), 91-102 (2013).</li><li>Kummari, E., Nichols, J. M., Yang, E. -. J., Kaplan, B. L. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neuroinflammation+and+B-cell+phenotypes+in+cervical+and+lumbosacral+regions+of+the+spinal+cord+in+experimental+autoimmune+encephalomyelitis+in+the+absence+of+pertussis+toxin.">Neuroinflammation and B-cell phenotypes in cervical and lumbosacral regions of the spinal cord in experimental autoimmune encephalomyelitis in the absence of pertussis toxin.</a> Neuroimmunomodulation. 26 (4), 198-207 (2019).</li><li>Huntemann, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+optimized+and+validated+protocol+for+inducing+chronic+experimental+autoimmune+encephalomyelitis+in+C57BL/6J+mice.">An optimized and validated protocol for inducing chronic experimental autoimmune encephalomyelitis in C57BL/6J mice.</a> J Neurosci Methods. 367, 109443 (2022).</li></ol>

None of the authors have any conflicts of interest to declare with respect to the research, authorship, and/or publication of this article.

The MOG35-55-induced EAE protocol that we described led to the development of a chronic form of MS in C57BL/6J mice7,8,13. In these representative results, we reported that the animals of both sexes that underwent the immunization procedure developed a chronic form of the disease (i.e., they do not fully recover after the disease onset, they accumulate deficits, and maintain a CS at least of 1.5 in the chronic phase)....

An erratum was issued for:&#160;<a href="https://www.jove.com/t/65778">Modeling Multiple Sclerosis in the Two Sexes: MOG35-55-Induced Experimental Autoimmune Encephalomyelitis</a>. The Authors section was updated. An additional affiliation (School of Pharmacy, Pharmacology Unit, University of Camerino) was added for author Antonino Casile.

An erratum was issued for:&#160;<a href="https://www.jove.com/t/65778">Modeling Multiple Sclerosis in the Two Sexes: MOG35-55-Induced Experimental Autoimmune Encephalomyelitis</a>. The Authors section was updated.

Erratum: Modeling Multiple Sclerosis in the Two Sexes: MOG35-55-Induced Experimental Autoimmune Encephalomyelitis

Multiple Sclerosis (MS) is a chronic autoimmune inflammatory disease affecting the central nervous system (CNS). It shows the presence of perivascular infiltration of inflammatory cells, demyelination, axonal loss, and gliosis1. Its etiology remains unknown, and its clinical aspects, radiographic, and pathological features suggest remarkable heterogeneity in the disease2.
Due to its unknown etiology and complexity, at present, no animal model recapitulates all the clinical and radiological features displayed in human MS3,4. However, various animal models are employed to study different aspects of MS3,4. In these models, disease initiation is typically extremely artificial, and the timeframe of the onset of clinical signs is different between humans and mice. For example, in humans, the pathophysiological processes underlying the disease are undetected for years before the onset of clinical manifestations. Conversely, the experimenters can detect symptoms in animal models within weeks or even days after MS induction4.
Three basic animal models produce the features of demyelination that are characteristic of MS: those that are virus-induced (e.g., Theiler&#39;s murine encephalomyelitis virus), those that are induced by toxic agents (e.g., cuprizone, lysolecithin), and the different variants of experimental autoimmune encephalomyelitis (EAE)5. Each model helps study some specific facets of the disease but none replicates all the features of MS6. Thus, it is critical to choose the correct model considering the specific experimental needs and the scientific questions to be addressed.
Thanks to immunization procedures against myelin-derived antigens, EAE is induced by triggering an autoimmune response to CNS components in susceptible mice. The interplay between a wide range of immunopathological and neuropathological mechanisms causes the development of principal pathological traits of MS (i.e., inflammation, demyelination, axonal loss, and gliosis) in the immunized mice7,8. Mice begin to show clinical symptoms around the second week after immunization and generally show ascending paralysis from the tail to the limb and forelimb. The clinical score (i.e., quantification of the accumulation of disease-related deficits) is generally assessed using a 5-point scale7.
Active immunization with protein or peptide or passive transfer of encephalitogenic T cells can be used to induce EAE in mice with different genetic backgrounds (e.g., SJL/J, C57BL/6, and non-obese-diabetic (NOD) mice). Myelin proteolipid protein (PLP), myelin basic protein (MBP), and myelin oligodendrocyte glycoprotein (MOG) are examples of self-CNS proteins from which immunogens are usually produced. Particularly, SJL/J mice immunized with the immunodominant epitope of PLP (PLP139&#8722;151) develop a relapsing-remitting (RR) disease course, while C57BL/6J mice immunized with the immunodominant MOG35-55 peptide show EAE of a chronic nature1. Despite some limitations, such as providing very little information about MS progression, the role of B cells in the disease, the inside-out mechanisms, or difficulties in studying remyelination, the EAE models have hugely contributed to the understanding of autoimmune and neuroinflammatory processes, increasing the knowledge in the MS field and thus allowing the development of novel therapeutic approaches for this disease4,6.
In the present work, we focused on a particular form of active EAE, the myelin oligodendrocyte glycoprotein peptide 35-55 (MOG35-55)-induced form9,10,11,12. The MOG35-55-induced EAE models a chronic form of MS. After immunization, the mice undergo an asymptomatic phase within the first week after immunization, then the disease typically arises during the second week after immunization, while between the third and fourth weeks after immunization, the disease becomes chronic, with no possibility of full recovery from the accumulated deficits7,8,13. Interestingly, no differences between males and females in incidence, disease onset, course, or progression are observed in most of the studies present in the literature14, even if fewer studies compare the disease in males and females.
In contrast, in humans, these parameters are known to be strongly sexually dimorphic2. MS affects more women than men; however, men generally develop a more aggressive form of the disease2. This evidence has suggested an essential, as well as complex, role of the gonadal hormones15; nevertheless, the role and the mechanism of action of sex hormones in the pathology remain unclear. Moreover, data from animal models support the idea that both estrogens and androgens exert positive effects on different tracts of the pathology in a sex-specific manner16,17.
Some studies also suggest neuroprotective, promyelinating, and anti-inflammatory effects of progesterone18 and, although evidence in MS patients is scarce18, neuroactive steroids (i.e., de novo synthetized steroids by the nervous system, such as pregnenolone, tetrahydroprogesterone, and dihydroprogesterone) might also affect the pathological course19. Collectively, these data support the idea that sex hormones produced both peripherally and inside the CNS have an important and sex-specific role in disease onset and progression. Therefore, in the present work, we urge the collection of separate data from both male and female animals.
From the histopathological point of view, the white matter of the spinal cord serves as the principal site of CNS injury in this model, which is characterized by multifocal, confluent regions of mononuclear inflammatory infiltration and demyelination8. Thus, in describing this protocol for the induction of MOG35-55-induced EAE in C57BL/6J mice, we will take into account the disease outcome in the two sexes and provide some histopathological insights regarding the spinal cords.

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			<td>18 G x 1 &#189;&#8220; 1.2 x 40 mm needle for the glass syringe&#160;</td>
			<td>Terumo</td>
			<td>TER-HYP-18G-112-PIN</td>
			<td></td>
		</tr>
		<tr>
			<td>Digital camera connected to the optical microscope</td>
			<td>NIKON DS-U1 digital camera</td>
			<td></td>
			<td></td>
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		<tr>
			<td>Electronic precision balance</td>
			<td>Merck</td>
			<td>Mod. Kern-440-47N, resolution 0.1 g</td>
			<td></td>
		</tr>
		<tr>
			<td>Eosin Y</td>
			<td>Sigma-Aldrich</td>
			<td>HT110216</td>
			<td></td>
		</tr>
		<tr>
			<td>Glass syringe pipet &#8220;ultra asept&#8221; 10 ml</td>
			<td>Sacco System&#160;</td>
			<td>L003465</td>
			<td></td>
		</tr>
		<tr>
			<td>Glassware (i.e., becker to prepare the emulsion)</td>
			<td>VWR</td>
			<td>213-1170, 213-1172</td>
			<td></td>
		</tr>
		<tr>
			<td>Hematoxylin (Mayer&#8217;s)</td>
			<td>Sigma-Aldrich</td>
			<td>MHS32</td>
			<td>Filter before using it.&#160;</td>
		</tr>
		<tr>
			<td>Image analysis Software</td>
			<td>Fiji</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Incomplete Freund&#8217;s adjuvant (IFA)</td>
			<td>Sigma-Aldrich</td>
			<td>F5506</td>
			<td>Store at +4 &#176;C.&#160;</td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Wellona Pharma</td>
			<td></td>
			<td>This drug is used as inhalational anaesthetic.</td>
		</tr>
		<tr>
			<td>Male and female C57BL/6J mice</td>
			<td>Jackson Laboratory, Envigo</td>
			<td></td>
			<td>Age 8-10 weeks, optimal body weight of ~20 g.&#160;</td>
		</tr>
		<tr>
			<td>Microtome</td>
			<td>Leica HistoCore BIOCUT R</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Mounting Medium&#160;</td>
			<td>Merck</td>
			<td>107961</td>
			<td></td>
		</tr>
		<tr>
			<td>Mouse Rotarod</td>
			<td>Ugo Basile&#160;</td>
			<td>#47600</td>
			<td></td>
		</tr>
		<tr>
			<td>Mycobacterium tuberculosis (MT), strain H37Ra&#160;</td>
			<td>Difco Laboratories Inc.&#160;</td>
			<td>231141</td>
			<td>Store at +4 &#176;C.</td>
		</tr>
		<tr>
			<td>Myelin oligodendrocyte glycoprotein peptide 35-55 (MOG35-55)</td>
			<td>Espikem</td>
			<td>EPK1</td>
			<td>Store at -80 &#176;C diluted (2 mg/mL) in physiological solution; prepare it on the day of the immunization to avoid, as much as possible, alterations or contaminations.&#160;</td>
		</tr>
		<tr>
			<td>Optical microscope</td>
			<td>NIKON eclipse 90i</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Paraformaldehyde (PFA)</td>
			<td>Sigma-Aldrich</td>
			<td>158127</td>
			<td>Store at +4 &#176;C once diluted (4%) in phosphate buffer.&#160;</td>
		</tr>
		<tr>
			<td>Pertussis toxin (PT)</td>
			<td>Duotech&#160;</td>
			<td>PT.181</td>
			<td>Store at -80&#176;C diluted (concentration 5 &#181;g/mL) in physiological solution&#160;</td>
		</tr>
		<tr>
			<td>Physiological solution (sodium chloride 0.9% solution)</td>
			<td>B. Eurospital</td>
			<td>A 032182038</td>
			<td>Store at +4 &#176;C once opened.</td>
		</tr>
		<tr>
			<td>Saline phosphate buffer (PBS)</td>
			<td>Thermo Scientific</td>
			<td>J61196.AP</td>
			<td></td>
		</tr>
		<tr>
			<td>Software for image acquisition</td>
			<td>&#160;NIS-Element AR 2.10</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Syringes U-100 0.5 mL with 30 G x 5/16&#8221; (0.30 x 8 mm) in fixed needle</td>
			<td>&#160;Nipro</td>
			<td>SYMS-0.5U100-3008B-EC</td>
			<td></td>
		</tr>
		<tr>
			<td>Syringes U-100 1 mL with 26G x &#189;&#8221; (0.45 x 12.7 mm) in needle</td>
			<td>PIC</td>
			<td>20,71,26,03,00,354</td>
			<td></td>
		</tr>
		<tr>
			<td>Vet ointment for eyes</td>
			<td>Lacrilube, Lacrigel Europhta</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Xylazine</td>
			<td>Rompun</td>
			<td></td>
			<td>This mixture of drug is used as injectable anaesthetic and sedative.&#160;</td>
		</tr>
		<tr>
			<td>Zolazepam and Tiletamine</td>
			<td>Zoletil&#160; 100</td>
			<td></td>
			<td>This drug is used as injectable anaesthetic, sedative, muscle relaxer, and analgesic</td>
		</tr>
	</tbody>
</table>

modeling multiple sclerosis in the two sexes: mog35-55-induced experimental autoimmune encephalomyelitis

Multiple Sclerosis (MS) is a chronic autoimmune inflammatory disease affecting the central nervous system (CNS). It is characterized by different prevalence in the sexes, affecting more women than men, and different outcomes, showing more aggressive forms in men than in women. Furthermore, MS is highly heterogeneous in terms of clinical aspects, radiological, and pathological features. Thus, it is necessary to take advantage of experimental animal models that allow the investigation of as many aspects of the pathology as possible. Experimental autoimmune encephalomyelitis (EAE) represents one of the most used models of MS in mice, modeling different disease features, from the activation of the immune system to CNS damage. Here we describe a protocol for the induction of EAE in both male and female C57BL/6J mice using myelin oligodendrocyte glycoprotein peptide 35-55 (MOG35-55) immunization, which leads to the development of a chronic form of the disease. We also report the evaluation of the daily clinical score and motor performance of these mice for 28 days post immunization (28 dpi). Lastly, we illustrate some basic histological analysis at the CNS level, focusing on the spinal cord as the primary site of disease-induced damage.

EAE follow-up after immunization 
This was assessed as described below.
Body weight and food intake 
The two-way analysis of variance (ANOVA) (sex and time as independent variables) shows a decrease in the BW of EAE animals of both sexes, especially within the second week post induction (F(1,57) = 4.952, p &#60; 0.001; Figure 2A). However, the sexual dimorphism in BW is always maintained (<st...

Watch this Scientific Journal Video about Creating a Versatile Experimental Autoimmune Encephalomyelitis Model Relevant for Both Male and Female Mice at JoVE.com

Author Spotlight: Creating a Versatile Experimental Autoimmune Encephalomyelitis Model Relevant for Both Male and Female Mice 

Experimental autoimmune encephalomyelitis is one of the most widely used murine models of multiple sclerosis. In the current protocol, C57BL/6J mice of both sexes are immunized with myelin oligodendrocyte glycoprotein peptide, resulting mainly in ascending paresis of the tail and limbs. Here we discuss the protocol of EAE induction and evaluation.

Modeling Multiple Sclerosis in the Two Sexes: MOG35-55-Induced Experimental Autoimmune Encephalomyelitis

Multiple Sclerosis (MS) is a chronic autoimmune inflammatory disease affecting the central nervous system (CNS). It is characterized by different ...

modeling-multiple-sclerosis-two-sexes-mog35-55-induced-experimental

Research

JoVE Journal

Neuroscience

1.2K Views.  Neuroscience Institute Cavalieri Ottolenghi (NICO). Experimental autoimmune encephalomyelitis is one of the most widely used murine models of multiple sclerosis. In the current protocol, C57BL/6J mice of both sexes are immunized with myelin oligodendrocyte glycoprotein peptide, resulting mainly in ascending paresis of the tail and limbs. Here we discuss the protocol of EAE induction and evaluation. 

Video: Author Spotlight: Creating a Versatile Experimental Autoimmune Encephalomyelitis Model Relevant for Both Male and Female Mice 

Automated Sholl Analysis of Digitized Neuronal Morphology at Multiple Scales

Neuronal morphology plays a significant role in determining how neurons function and communicate1-3. Specifically, it affects the ability of neurons to receive inputs from other cells2 and contributes to the propagation of action potentials4,5. The morphology of the neurites also affects how information is processed. The diversity of dendrite morphologies facilitate local and long range signaling and allow individual neurons or groups of neurons to carry out specialized functions within the neuronal network6,7. Alterations in dendrite morphology, including fragmentation of dendrites and changes in branching patterns, have been observed in a number of disease states, including Alzheimer's disease8, schizophrenia9,10, and mental retardation11. The ability to both understand the factors that shape dendrite morphologies and to identify changes in dendrite morphologies is essential in the understanding of nervous system function and dysfunction.
Neurite morphology is often analyzed by Sholl analysis and by counting the number of neurites and the number of branch tips. This analysis is generally applied to dendrites, but it can also be applied to axons. Performing this analysis by hand is both time consuming and inevitably introduces variability due to experimenter bias and inconsistency. The Bonfire program is a semi-automated approach to the analysis of dendrite and axon morphology that builds upon available open-source morphological analysis tools. Our program enables the detection of local changes in dendrite and axon branching behaviors by performing Sholl analysis on subregions of the neuritic arbor. For example, Sholl analysis is performed on both the neuron as a whole as well as on each subset of processes (primary, secondary, terminal, root, etc.) Dendrite and axon patterning is influenced by a number of intracellular and extracellular factors, many acting locally. Thus, the resulting arbor morphology is a result of specific processes acting on specific neurites, making it necessary to perform morphological analysis on a smaller scale in order to observe these local variations12.
The Bonfire program requires the use of two open-source analysis tools, the NeuronJ plugin to ImageJ and NeuronStudio. Neurons are traced in ImageJ, and NeuronStudio is used to define the connectivity between neurites. Bonfire contains a number of custom scripts written in MATLAB (MathWorks) that are used to convert the data into the appropriate format for further analysis, check for user errors, and ultimately perform Sholl analysis. Finally, data are exported into Excel for statistical analysis. A flow chart of the Bonfire program is shown in Figure 1.

We have developed a computer program to analyze neuronal morphology. In combination with two existing open source analysis tools, our program performs Sholl analysis and determines the number of neurites, branch points, and neurite tips. The analyses are performed so that local changes in neurite morphology can be observed.

Neuronal morphology plays a significant role in determining how neurons function and communicate1-3.  Specifically, it affects the ability of neurons to ...

Hyponeophagia: A Measure of Anxiety in the Mouse

Before the present day, when fast-acting and potent rodenticides such as alpha-chloralose were not yet in use, the work of pest controllers was often hampered by a phenomenon known as "bait shyness". Mice and rats cannot vomit, due to the tightness of the cardiac sphincter of the stomach, so to overcome the problem of potential food toxicity they have evolved a strategy of first ingesting only very small amounts of novel substances. The amounts ingested then gradually increase until the animal has determined whether the substance is safe and nutritious. So the old rat-catchers would first put a palatable substance such as oatmeal, which was to be the vehicle for the toxin, in the infested area. Only when large amounts were being readily consumed would they then add the poison, in amounts calculated not to affect the taste of the vehicle. The poisoned bait, which the animals were now readily eating in large amounts, would then swiftly perform its function.
Bait shyness is now used in the behavioural laboratory as a way of measuring anxiety. A highly palatable but novel substance, such as sweet corn, nuts or sweetened condensed milk, is offered to the mice (or rats) in a novel situation, such as a new cage. The latency to consume a defined amount of the new food is then measured.
Robert M.J. Deacon can be reach at <a href="mailto:robert.deacon@psy.ox.ac.uk">robert.deacon@psy.ox.ac.uk</a>

Mice and rats, due to their innate cautiousness, are initially slow in consuming a novel food, particularly in a novel place. This hyponeophagia can readily be measured in the laboratory, even though laboratory animals are much less anxious than their wild counterparts

Before the present day, when fast-acting and potent rodenticides such as alpha-chloralose were not yet in use, the work of pest controllers was often ...

Simultaneous Two-photon In Vivo Imaging of Synaptic Inputs and Postsynaptic Targets in the Mouse Retrosplenial Cortex

This video shows the craniotomy procedure that allows chronic imaging of neurons in the mouse retrosplenial cortex (RSC) using in vivo two-photon microscopy in Thy1-GFP transgenic mouse line. This approach creates a possibility to investigate the correlation of behavioural manipulations with changes in neuronal morphology in vivo.
The cranial window implantation procedure was considered to be limited only to the easily accessible cortex regions such as the barrel field. Our approach allows visualization of neurons in the highly vascularized RSC. RSC is an important element of the brain circuit responsible for spatial memory, previously deemed to be problematic for in vivo two-photon imaging.
The cranial window implantation over the RSC is combined with an injection of mCherry-expressing recombinant adeno-associated virus (rAAVmCherry) into the dorsal hippocampus. The expressed mCherry spreads out to axonal projections from the hippocampus to RSC, enabling the visualization of changes in both presynaptic axonal boutons and postsynaptic dendritic spines in the cortex. 
This technique allows long-term monitoring of experience-dependent structural plasticity in RSC.

Simultaneous Two-photon In Vivo Imaging of Synaptic Inputs and Postsynaptic Targets in the Mouse Retrosplenial Cortex

This video shows the craniotomy procedure that allows chronic imaging of neurons in mouse retrosplenial cortex using in vivo two photon microscopy in Thy1-GFP transgenic line. This approach is combined with injection of mCherry-expressing adeno-associated virus into dorsal hippocampus. These techniques allow long-term monitoring of experience-dependent structural plasticity in RSC.

This video shows the craniotomy procedure that allows chronic imaging of neurons in the mouse retrosplenial cortex (RSC) using in vivo two-photon ...

Measuring Progressive Neurological Disability in a Mouse Model of Multiple Sclerosis

After intracerebral infection with the Theiler's Murine Encephalomyelitis Virus (TMEV), susceptible SJL mice develop a chronic-progressive demyelinating disease, with clinical features similar to the progressive forms of multiple sclerosis (MS). The mice show progressive disability with loss of motor and sensory functions, which can be assessed with multiple apparatuses and protocols. Among them, the Rotarod performance test is a very common behavioral test, its advantage being that it provides objective measurements, but it is often used assuming that it is straightforward and simple. In contrast to visual scoring systems used in some models of MS, which are highly subjective, the Rotarod test generates an objective, measurable, continuous variable (i.e., length of time), allowing almost perfect inter-rater concordances. However, inter-laboratory reliability is only achieved if the various testing parameters are replicated. In this manuscript, recommendations of specific testing parameters, such as size, speed, and acceleration of the rod; amount of training given to the animals; and data processing, are presented for the Rotarod test.

An optimized testing protocol is presented in this paper for the Rotarod performance test, used for measuring progressive neurological disability in TMEV-infected mice.

After intracerebral infection with the Theiler's Murine Encephalomyelitis Virus (TMEV), susceptible SJL mice develop a chronic-progressive demyelinating ...

Sagittal Plane Kinematic Gait Analysis in C57BL/6 Mice Subjected to MOG35-55 Induced Experimental Autoimmune Encephalomyelitis

Kinematic gait analysis in the sagittal plane has frequently been used to characterize motor deficits in multiple sclerosis (MS). We describe the application of these techniques to identify gait deficits in a mouse model of MS, known as experimental autoimmune encephalomyelitis (EAE). Paralysis and motor deficits in mice subjected to EAE are typically assessed using a clinical scoring scale. However, this scale yields only ordinal data that provides little information about the precise nature of the motor deficits. EAE disease severity has also been assessed by rotarod performance, which provides a measure of general motor coordination. By contrast, kinematic gait analysis of the hind limb in the sagittal plane generates highly precise information about how movement is impaired. To perform this procedure, reflective markers are placed on a hind limb to detect joint movement while a mouse is walking on a treadmill. Motion analysis software is used to measure movement of the markers during walking. Kinematic gait parameters are then derived from the resultant data. We show how these gait parameters can be used to quantify impaired movements of the hip, knee, and ankle joints in EAE. These techniques may be used to better understand disease mechanisms and identify potential treatments for MS and other neurodegenerative disorders that impair mobility.

Kinematic gait analysis in the sagittal plane yields highly precise information about how movement is executed. We describe the application of these techniques to identify gait deficits for mice subjected to autoimmune-mediated demyelination. These methods may also be used to characterize gait deficits for other mouse models featuring impaired locomotion.

Kinematic gait analysis in the sagittal plane has frequently been used to characterize motor deficits in multiple sclerosis (MS). We describe the ...

A Simple Approach to Induce Experimental Autoimmune Neuritis in C57BL/6 Mice for Functional and Neuropathological Assessments

Experimental autoimmune neuritis (EAN) is a well-appreciated experimental model of autoimmune peripheral demyelinating diseases. EAN disease is induced by immunizing mice with neurogenic peptides to direct an inflammatory attack toward components of the peripheral nervous system (PNS). Recent advances have enabled the induction of EAN in the relatively resistant C57BL/6 mouse line using myelin protein zero (P0)106-125 or P0180-199 peptides delivered in adjuvant combined with the injection of pertussis toxin. The ability to induce EAN in the C57BL/6 strain allows for the use of the numerous genetic tools that exist on this mouse background, and thus allows the sophisticated study of disease pathogenesis and interrogation of the mechanistic action of novel therapeutics in combination with transgenic approaches. In this study, we demonstrate a simple approach to successfully induce EAN using the P0180-199 peptide in C57BL/6 mice. We also outline a protocol for the assessment of functional deficits that occur in this model, accompanied by an array of neuropathological features. Thus, this model is a powerful experimental model to study the pathogenesis of human peripheral demyelinating neuropathies, and to determine the efficacy of potential therapies that aim to promote myelin repair and protect against nerve damage in autoimmune neuritis.

This report outlines a simple approach to successfully induce experimental autoimmune neuritis (EAN) using the myelin protein zero (P0)180-199 peptide in combination with Freund&#39;s complete adjuvant and pertussis toxin. We present a sophisticated paradigm capable of accurately assessing the extent of functional deficits and neuropathology that occur in this EAN.

Experimental autoimmune neuritis (EAN) is a well-appreciated experimental model of autoimmune peripheral demyelinating diseases. EAN disease is induced by ...

Comprehensive Autopsy Program for Individuals with Multiple Sclerosis

We describe a rapid tissue donation program for individuals with multiple sclerosis (MS) that requires scientists and technicians to be on-call 24/7, 365 days a year. Participants consent to donate their brain and spinal cord. Most patients were followed by neurologists at the Cleveland Clinic Mellen Center for MS Treatment and Research. Their clinical courses and neurological disabilities are well-characterized. Soon after death, the body is transported to the MS Imaging Center, where the brain is scanned in situ by 3 T magnetic resonance imaging (MRI). The body is then transferred to the autopsy room, where the brain and spinal cord are removed. The brain is divided into two hemispheres. One hemisphere is immediately placed in a slicing box and alternate 1 cm-thick slices are either fixed in 4% paraformaldehyde for two days or rapidly frozen in dry ice and 2-methylbutane. The short-fixed brain slices are stored in a cryopreservation solution and used for histological analyses and immunocytochemical detection of sensitive antigens. Frozen slices are stored at -80 &#176;C and used for molecular, immunocytochemical, and in situ hybridization/RNA scope studies. The other hemisphere is placed in 4% paraformaldehyde for several months, placed in the slicing box, re-scanned in the 3 T magnetic resonance (MR) scanner and sliced into centimeter-thick slices. Postmortem in situ MR images (MRIs) are co-registered with 1 cm-thick brain slices to facilitate MRI-pathology correlations. All brain slices are photographed and brain white-matter lesions are identified. The spinal cord is cut into 2 cm segments. Alternate segments are fixed in 4% paraformaldehyde or rapidly frozen. The rapid procurement of postmortem MS tissues allows pathological and molecular analyses of MS brains and spinal cords and pathological correlations of brain MRI abnormalities. The quality of these rapidly-processed postmortem tissues (usually within 6 h of death) is of great value to MS research and has resulted in many high-impact discoveries.

Multiple sclerosis is an inflammatory demyelinating disease with no cure. Analysis of brain tissue provides important clues to understanding the pathogenesis of the disease. Here we discuss the methodology and downstream analysis of MS brain tissue collected through a unique rapid autopsy program in operation at the Cleveland Clinic.

We describe a rapid tissue donation program for individuals with multiple sclerosis (MS) that requires scientists and technicians to be on-call 24/7, 365 ...

Transpupillary Two-Photon In Vivo Imaging of the Mouse Retina

The retina transforms light signals from the environment into electrical signals that are propagated to the brain. Diseases of the retina are prevalent and cause visual impairment and blindness. Understanding how such diseases progress is critical to formulating new treatments. In vivo&#160;microscopy in animal models of disease is a powerful tool for understanding neurodegeneration and has led to important progress towards treatments of conditions ranging from Alzheimer&#39;s disease to stroke. Given that the retina is the only central nervous system structure inherently accessible by optical approaches, it naturally lends itself towards in vivo imaging. However, the native optics of the lens and cornea present some challenges for effective imaging access.
This protocol outlines methods for in vivo two-photon imaging of cellular cohorts and structures in the mouse retina at cellular resolution, applicable for both acute- and chronic-duration imaging experiments. It presents examples of retinal ganglion cell (RGC), amacrine cell, microglial, and vascular imaging using a suite of labeling techniques including adeno-associated virus (AAV) vectors, transgenic mice, and inorganic dyes. Importantly, these techniques extend to all cell types of the retina, and suggested methods for accessing other cellular populations of interest are described. Also detailed are example strategies for manual image postprocessing for display and quantification. These techniques are directly applicable to studies of retinal function in health and disease.

In vivo imaging is a powerful tool for the study of biology in health and disease. This protocol describes transpupillary imaging of the mouse retina with a standard two-photon microscope. It also demonstrates different in vivoimaging methods to fluorescently label multiple cellular cohorts of the retina.

The retina transforms light signals from the environment into electrical signals that are propagated to the brain. Diseases of the retina are prevalent ...

Positron Emission Tomography Imaging for In Vivo Measuring of Myelin Content in the Lysolecithin Rat Model of Multiple Sclerosis

Multiple sclerosis (MS) is a neuroinflammatory disease with expanding axonal and neuronal degeneration and demyelination in the central nervous system, leading to motor dysfunctions, psychical disability, and cognitive impairment during MS progression. Positron emission tomography (PET) is an imaging technique able to quantify in vivo cellular and molecular alterations.
Radiotracers with affinity to intact myelin can be used for in vivo imaging of myelin content changes over time. It is possible to detect either an increase or decrease in myelin content, what means this imaging technique can detect demyelination and remyelination processes of the central nervous system. In this protocol we demonstrate how to use PET imaging to detect myelin changes in the lysolecithin rat model, which is a model of focal demyelination lesion (induced by stereotactic injection) (i.e., a model of multiple sclerosis disease). 11C-PIB PET imaging was performed at baseline, and 1 week and 4 weeks after stereotaxic injection of lysolecithin 1% in the right striatum (4 &#181;L) and corpus callosum (3 &#181;L) of the rat brain, allowing quantification of focal demyelination (injection site after 1 week) and the remyelination process (injection site at 4 weeks).
Myelin PET imaging is an interesting tool for monitoring in vivo changes in myelin content which could be useful for monitoring demyelinating disease progression and therapeutic response.

Positron Emission Tomography Imaging for In Vivo Measuring of Myelin Content in the Lysolecithin Rat Model of Multiple Sclerosis

This protocol has the aim of monitoring in vivo myelin changes (demyelination and remyelination) by positron emission tomography (PET) imaging in an animal model of multiple sclerosis.

Multiple sclerosis (MS) is a neuroinflammatory disease with expanding axonal and neuronal degeneration and demyelination in the central nervous system, ...

Modeling Neonatal Intraventricular Hemorrhage Through Intraventricular Injection of Hemoglobin

Neonatal intraventricular hemorrhage (IVH) is a common consequence of premature birth and leads to brain injury, posthemorrhagic hydrocephalus (PHH), and lifelong neurological deficits. While PHH can be treated by temporary and permanent cerebrospinal fluid (CSF) diversion procedures (ventricular reservoir and ventriculoperitoneal shunt, respectively), there are no pharmacological strategies to prevent or treat IVH-induced brain injury and hydrocephalus. Animal models are needed to better understand the pathophysiology of IVH and test pharmacological treatments. While there are existing models of neonatal IVH, those that reliably result in hydrocephalus are often limited by the necessity for large-volume injections, which may complicate modeling of the pathology or introduce variability in the clinical phenotype observed.
Recent clinical studies have implicated hemoglobin and ferritin in causing ventricular enlargement after IVH. Here, we develop a straightforward animal model that mimics the clinical phenotype of PHH utilizing small-volume intraventricular injections of the blood breakdown product hemoglobin. In addition to reliably inducing ventricular enlargement and hydrocephalus, this model results in white matter injury, inflammation, and immune cell infiltration in periventricular and white matter regions. This paper describes this clinically relevant, simple method for modeling IVH-PHH in neonatal rats using intraventricular injection and presents methods for quantifying ventricle size post injection.

We present a model of neonatal intraventricular hemorrhage using rat pups that mimics the pathology seen in humans.

Neonatal intraventricular hemorrhage (IVH) is a common consequence of premature birth and leads to brain injury, posthemorrhagic hydrocephalus (PHH), and ...

Modeling Multiple Sclerosis in the Two Sexes: MOG_35-55-Induced Experimental Autoimmune Encephalomyelitis

Summary

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Chapters in this video

Automated Sholl Analysis of Digitized Neuronal Morphology at Multiple Scales

Hyponeophagia: A Measure of Anxiety in the Mouse

Simultaneous Two-photon In Vivo Imaging of Synaptic Inputs and Postsynaptic Targets in the Mouse Retrosplenial Cortex

Measuring Progressive Neurological Disability in a Mouse Model of Multiple Sclerosis

Sagittal Plane Kinematic Gait Analysis in C57BL/6 Mice Subjected to MOG35-55 Induced Experimental Autoimmune Encephalomyelitis

A Simple Approach to Induce Experimental Autoimmune Neuritis in C57BL/6 Mice for Functional and Neuropathological Assessments

Comprehensive Autopsy Program for Individuals with Multiple Sclerosis

Transpupillary Two-Photon In Vivo Imaging of the Mouse Retina

Positron Emission Tomography Imaging for In Vivo Measuring of Myelin Content in the Lysolecithin Rat Model of Multiple Sclerosis

Modeling Neonatal Intraventricular Hemorrhage Through Intraventricular Injection of Hemoglobin