Animal models are used to study demyelination and, importantly, recovery from demyelinating insult. Generally, recovery is assessed by morphological techniques which are dauntingly labor intensive if the results are to be both quantitative and to represent various regions of the CNS. A biochemical methodology which allows quantitation of the rate and extent of remyelination following demyelinating insult will be developed and calibrated. Proposed experiments will define, in vivo during development (8-35 days of age), the stoichiometry for incorporation of radioactivity from 3[H]-water into the myelin-specific lipid, cerebroside, and demonstrate that this correlates with myelination. The same assay will next be used to quantitate remyelination, subsequent to the massive demyelination induced by feeding of adult mice with the copper chelating agent, cuprizone. These detailed data concerning time-course of cuprizone-induced demyelination/remyelination will be correlated with expression of mRNA coding for myelin structural proteins as well as other genes involved in myelin assembly. If preliminary results are verified, there will be upregulation of mRNA for myelin genes long prior to remyelination. Since myelin gene upregulation does not correlate with remyelination, the applicants will use this convenient system to determine at what level (i.e., whether translational or post-translational) assembly of proteins into myelin is controlled. Also assayed will be levels of mRNA for proteins involved in cholesterol trafficking. These data will be informative concerning involvement extracellular lipoproteins in regeneration of neural membranes. Additionally, levels of mRNA for certain cytokines and growth factors will be assayed. Data obtained will be used to test hypotheses concerning sequential involvement of different cell types in initiating demyelination and, eventually, remyelination. Such information is relevant to understanding of human demyelinating disorders and may be of interest with respect to suggesting therapies for such disorders.
