HYPOXIA INHIBITS CYCLIC NUCLEOTIDE-STIMULATED EPITHELIAL ION-TRANSPORT - ROLE FOR NUCLEOTIDE CYCLASES AS OXYGEN SENSORS

Decreased oxygen delivery to cells (hypoxia) is prevalent in a number of important diseases. Little is known about mechanisms of oxygen sens ing at the cellular level or about whether functional correlates of ox ygen sensing exist. In this study, we examined the impact of hypoxia o n stimulated epithelial ion transport function. T84 cells, a model of intestinal epithelia, were grown on permeable supports, exposed to hyp oxia (range 1-21% O-2) for periods of time between 0 and 72 h and asse ssed for stimulated ion transport. Hypoxia evoked a specific decrease in cyclic nucleotide-stimulated (cAMP and cGMP) but not Ca++-stimulate d ion transport. Rb-86 (K+ tracer) uptake and I-125 (Cl- tracer) efflu x were reduced in hypoxic cells by >50% and >40%, respectively, fluid movement was reduced by hypoxia (>50% decrease) and reoxygenation resu lted in partial recovery of the ion transport responses. Stimulated an d basal levels of both cAMP and cGMP were decreased in response to hyp oxia, although intracellular ATP levels were unaltered under similar c onditions. Exogenous addition of cobalt, nickel or manganese, all of w hich compete far oxygen binding on heme-containing proteins, mimicked hypoxia, Because guanylate cyclase is a heme protein, we measured the influence of cobalt on activity of guanylate cyclase in purified plasm a membrane preparations and found cobalt to inhibit stimulated cGMP le vels in this cell-free system. Finally, pharmacological lowering of in tracellular cGMP (using LY83583) resulted in decreased cAMP-stimulated Cl- secretion, and direct elevation of cGMP (using 8-bromo-cGMP or di butyryl-cGMP) restored this hypoxia-induced activity. We conclude that a potential oxygen-sensing mechanism of epithelial cells involves the cooperation of heme-containing proteins such as guanylate cyclase and that biochemical cross-talk between cAMP- and cGMP-stimulated pathway s may be important in such responses.