Requirement for nonoligodendrocyte cell signals for enhanced myelinogenic gene expression in long-term cultures of purified rat oligodendrocytes

Abstract

Comparisons have been made of myelinogenic activities in fetal rat brain mixed primary cultures and cultures of isolated oligodendrocytes of comparable age. The specific activities of the sulfatide synthesis, 2',3'-cyclic-nucleotide 3'-phosphohydrolase (2',3'-cyclic-nucleotide 3'-phosphodiesterase, EC 3.1.4.37), and accumulation of myelin basic protein, when expressed per mg of protein, were as high (or generally higher) in isolated oligodendrocyte cultures as in comparable mixed primary cultures at 29 days. However, when these data were analyzed per oligodendrocyte, it became apparent that the isolated oligodendrocytes were substantially less active than their mixed culture counterparts. The results suggest the necessity of nonologodendrocyte positive signals for the optimal expression by oligodendrocytes of myelin-related differentiated functions. The isolation method involves the selection of oligodendrocytes by shaking them from primary cultures of rat brain, followed by the lysis of other contaminating cells in a balanced salt solution at pH 7.2. More than 99% of the isolated cells are viable, at least initially divide, and can be cultured for at least 60 days. The oligodendrocytes selected in this way were characterized by: (i) morphology, (ii) immunofluorescence labeling by antibodies to myelin basic protein and galactosylceramide, and (iii) biochemical analyses for myelin basic protein, activity of 2',3'-cyclic-nucleotide 3'-phosphohydrolase, and sulfogalactosylceramide synthesis.