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T₂*-weighted gradient-echo MRI images at high field (≥7T) have shown rich image contrast within and between brain regions. The source for these contrast variations has been primarily attributed to tissue magnetic susceptibility differences. In this study, the contribution of myelin to both T₂* and frequency contrasts is investigated using a mouse model of demyelination based on a cuprizone diet. The demyelinated brains showed significantly increased T₂* in white matter and a substantial reduction in gray-white matter frequency contrast, suggesting that myelin is a primary source for these contrasts. Comparison of in-vivo and in-vitro data showed that, although tissue T₂* values were reduced by formalin fixation, gray-white matter frequency contrast was relatively unaffected and fixation had a negligible effect on cuprizone-induced changes in T₂* and frequency contrasts.