Many studies indicate that hypoxic inhibition of some K+ channels in the membrane of the pulmonary arterial smooth muscle cells (PASMCs) plays a part in initiating hypoxic pulmonary vasoconstriction. The sensitivity of the K+ current (I(k)), resting membrane potential (E(m)), and intracellular Ca2+ concentration ([Ca2+]i) of PASMCs to different levels of hypoxia in these cells has not been explored fully. Reducing PO2 levels gradually inhibited steady-state I(k) of rat resistance PASMCs and depolarized the cell membrane. The block of I(k) by hypoxia was voltage dependent in that low O2 tensions (3 and 0% O2) inhibited I(k) more at 0 and -20 mV than at 50 mV. As expected, the hypoxia-sensitive I(k) was also 4-aminopyridine sensitive. Fura 2-loaded PASMCs showed a graded increase in [Ca2+]i as PO2 levels declined. This increase was reduced markedly by nifedipine and removal of extracellular Ca2+. We conclude that, as in the carotid body type I cells, PC-12 pheochromocytoma cells, and cortical neurons, increasing severity of hypoxia causes a proportional decrease in I(k) and E(m) and an increase of [Ca2+]i.