ATP-stimulated c-fos and zif268 mRNA expression is inhibited by chemical hypoxia in a rat brain-derived type 2...

The stimulus-transcriptional coupling during ischemia/hypoxia was examined for ATP-stimulated expression of immediate early genes (IEGs; c-fos, zif268, c-myc and nur77) in a rat brain-derived type 2 astrocyte cell line, RBA-2. Incubation of cells with 1 mM of extracellular ATP stimulated time-dependent expression of c-fos and zif268. ATP induced the largest increases in zif268 mRNA and a lesser one in c-fos mRNA. ATP also induced a slight increase in nur77 mRNA but was ineffective in inducing c-myc expression in these cells. Brief exposure of cells to potassium cyanide to simulate chemical hypoxia induced 9-fold and 7-fold transient increases in c-fos and zif268 expression, respectively, but did not affect c-myc or nur77 expression. When cyanide and ATP were added together, the expression of c-fos and zif268 expression was inhibited, and the effect was mimicked by simulating chemical hypoxia with sodium azide. To elucidate the mechanism involved, the effect of cyanide on ATP-stimulated increases in intracellular Ca(2+) concentrations, [Ca(2+)](i), and phospholipase D (PLD) activities were measured. Cyanide induced an increase in [Ca(2&plus);](i) and further enhanced the ATP-stimulated increases in [Ca(2+)](i) and PLD activities. Nevertheless, metabolic inhibitor, iodoacetate, blocked the ATP-induced c-fos and partially inhibited zif268 expression, and deprivation of cells with glucose also inhibited the ATP-induced c-fos expression. Taken together, these results demonstrate that both extracellular ATP and chemical hypoxia induce c-fos and zif268 expression in RBA-2 type 2 astrocytes. The chemical hypoxia inhibited ATP-stimulated c-fos and zif268 expression is not due to alterations in Ca(2+) and PLD signaling, and is at least partially related to metabolic disturbance in these cells.