OPCs (oligodendrocyte precursor cells) express golli proteins which, through regulation of Ca2+ influx, appear to be important in OPC process extension/retraction and migration. The aim of the present study was to examine further the role of golli in regulating OPC development. The effects of golli ablation and overexpression were examined in primary cultures of OPCs prepared from golli-KO (knockout) and JOE (golli J37-overexpressing) mice. In OPCs lacking golli, or overexpressing golli, differentiation induced by growth factor withdrawal was impaired. Proliferation analysis in the presence of PDGF (platelet-derived growth factor), revealed that golli enhanced the mitogen-stimulated proliferation of OPCs through activation of SOCCs (store-operated Ca2+ channels). PDGF treatment induced a biphasic increase in OPC intracellular Ca2+, and golli specifically increased Ca2+ influx during the second SOCC-dependent phase that followed the initial release of Ca2+ from intracellular stores. This store-operated Ca2+ uptake appeared to be essential for cell division, since specific SOCC antagonists completely blocked the effects of PDGF and golli on OPC proliferation. Additionally, in OPCs overexpressing golli, increased cell death was observed after mitogen withdrawal. This phenomenon could be prevented by exposure to VOCC (voltage-operated Ca2+ channel) blockers, indicating that the effect of golli on cell death involved increased Ca2+ influx through VOCCs. The results showed a clear effect of golli on OPC development and support a role for golli in modulating multiple Ca2+-regulatory events through VOCCs and SOCCs. Our results also suggest that PDGF engagement of its receptor resulting in OPC proliferation proceeds through activation of SOCCs.
Keywords: apoptosis, calcium influx, cell cycle, golli protein, oligodendrocyte, platelet-derived growth factor (PDGF)
Abbreviations: 2-APB, 2-aminoethoxydiphenyl borate; bFGF, basic fibroblast growth factor; BrdU, bromodeoxyuridine; [Ca2+]int, intracellular Ca2+ concentration; CNS, central nervous system; DIV, days in vitro; div, days in vitro for mixed cultures; DMEM, Dulbecco’s modified Eagle’s medium; FBS, fetal bovine serum; fura 2/AM, fura 2 acetoxymethyl ester; GC, galactocerebroside; GFP, green fluorescent protein; JOE, golli J37-overexpressing; KO, knockout; MBP, myelin basic protein; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide; NG2, nerve/glial antigen 2; OPC, oligodendrocyte precursor cell; OPC˜GFP, GFP-labelled OPC; PDGF, platelet-derived growth factor; Plp, proteolipid; SOCC, store-operated Ca2+ channel; Tc, cell-cycle time; VOCC, voltage-operated Ca2+ channel