Oligodendrocytes are the myelin-forming cells in the central nervous system (CNS) and their loss or dysfunction is a hallmark of CNS demyelinating diseases, such as multiple sclerosis (MS), hypoxic-ischemic demyelination, or spinal cord injury. In the rodent CNS, oligodendrocyte progenitor cells (OPCs) arise in multiple ventral and dorsal locations of the forebrain during late embryogenesis and early postnatal periods. OPCs migrate out from these germinal zones and disperse throughout the CNS, to populate the developing white and gray matter. There, OPCs can begin to mature through a series of intermediate states characterized by the expression of stage-specific proteins until completely differentiated into postmitotic myelinating oligodendrocytes. Elucidating the cellular and molecular mechanisms that control oligodendrocyte maturation requires isolating OPCs and premyelinating oligodendrocytes by rapid and reliable methods that provide high yield, pure and viable culture, being a powerful tool to characterize their differentiation and potential capacity for myelin repair after injury. This chapter describes in detail two simple and efficient protocols for the preparation of highly enriched rat OPC populations and immature oligodendrocytes derived from mixed glial cultures and optic nerves, respectively. Functional oligodendrocytes obtained with these protocols can be cocultured with primary neurons to study myelination.
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