Oxidative-reductive modulation of guinea pig splenic cell guanylate cyclase activity.

The modulation of cyclic GMP metabolism by oxidativereductive-related processes was studied in intact guinea pig splenic cells and with respect to the activities of soluble and particulate guanylate cyclase. In intact cells the oxidant dehydroascorbic acid produced timeand concentration-dependent increases in cGMP steady state levels which were potentiated by the phosphodiesterase inhibitor 3-isobutyl-lmethylxanthine. Thiol-containing reductants, such as cysteine or dithiothreitol, decreased cellular cGMP levels 50 to 70% and exposure of cells to sulfhydryl reactive reagents, such as N-ethylmaleimide, p-hydroxymercuribenzoate, or 5,5’-dithiobis(Z-nitrobenzoic acid), or to the transition metal chelator 4.7-diphenyl-l,lO-phenanthroline also caused striking lowering of cGMP levels. The relatively stable dehydroascorbic acid-induced elevation of cGMP in intact cells, which persisted after removal of the oxidant, could be reversed upon reduction by dithiothreitol, and the stable depression of cGMP steady state levels induced by dithiothreitol could be reversed by exposure of the cells to the oxidant, dehydroascorbic acid. The effects produced by dehydroascorbic acid, dithiothreitol, and N-ethylmaleimide on soluble and particulate fractions of guanylate cyclase from these cells corresponded to the alterations each produced on cGMP steady state levels in the intact cells. Dehydroascorbic acid (0.1 to 10 rn& activated while dithiothreitol (0.1 to 5 mM) or N-ethylmaleimide (0.5 to 5 mM) inhibited guanylate cyclase activity. Dehydroasrorbic acid activation of the soluble enzyme was rapid and additive with the slower activation that occurred spontaneously in air; dithiothreitol could prevent or reverse dehydroascorbic acid as well as spontaneous air activation. Activation or inhibition by dehydroascorbic acid or dithiothreitol, respectively, of the particulate enzyme persisted upon removal of the effecters but this stable effect produced by the oxidant could be reversed by the reductant and vice versa. Membrane-associated basal and fluoride-stimulated adenylate cyclase activity was suppressed (50 to 70%) by