Figure 3.

Developmental Profile of Oligodendrocytes in Cocultures and Effects of P2-Receptor Activation The state of differentiation of oligodendrocytes in cocultures was determined by staining with antibodies against markers of OPC differentiation after 7, 9, 11, and 14 days in coculture with DRG neurons in the presence and absence of 100 μM 2MeSATP, and cell proliferation was measured by BrdU incorporation (magenta). Hoechst nuclear stain was used to identify all cells in the culture (blue) and to calculate the percentage of cells identified with each marker. Mean percentage and SEM of cells stained positively with each marker are indicated in the lower right corner of each figure (n = 40 microscope fields in each). Immunocytochemical staining for a marker of early oligodendrocytes O4 is shown in green; a marker for intermediate stage oligodendrocytes, myelin basic protein (MBP), is shown in red; and the myelin oligodendrocyte glycoprotein (MOG) expressed by mature oligodendrocytes is shown in yellow. The results show that OPCs have progressed into a postmitotic phase and begun to express the O4 antigen in large numbers by 7 days in culture, the time when treatment with 2MeSATP was begun. ATP receptor activation with 2MeSATP had no effects on maturation or proliferation of oligodendrocytes when applied after 7 days in coculture, but it did increase the number of cells expressing the mature oligodendrocyte marker MOG after 11 and 14 days (4-7 days 2MeSATP treatment; p < 0.001). The results show that ATP receptor activation increases myelination by increasing development of oligodendrocytes at the mature MOG positive stage, rather than acting on cells at the progenitor stage.