MEMBRANE DEPOLARIZATION IN PC-12 CELLS DURING HYPOXIA IS REGULATED BYAN O-2-SENSITIVE K+ CURRENT

The effects of hypoxia on K+ current (I-K), resting membrane potential , and cytosolic free CB2+ in rat pheochromocytoma (PC-12) cells were s tudied. Whole cell voltage- and current-clamp experiments were perform ed to measure IK and membrane potential, respectively. Cytosolic free Ca2+ level was measured using the Ca2+-sensitive fluorescent dye fura 2. Depolarizing voltage steps to +50 mV from a holding potential of -9 0 mV elicited a slowly inactivating, tetraethylammonium chloride-sensi tive, and Ca2+-insensitive IR that was reversibly inhibited by reduced O-2 tension. Graded reduction in P-O2 (from 150 to 0 mmHg) induced a graded inhibition of an O-2-sensitive I-K [I-K(O2)] up to 46% at 0 mmH g. Moreover, hypoxia induced a 19-mV membrane depolarization and a two fold increase in cytosolic free Ca2+. In Ca2+-free condition, inhibiti on I-K(O2) induced an 8-mV depolarization, suggesting that inhibition of I-K(O2) was responsible for initiating depolarization. The effect o f reduced P-O2 on the current-voltage relationship showed a reduction of out ward current and a 14-mV shift in the reversal potential compar able with the amount of depolarization measured in current clamp exper iments. Neither Ca2+-activated I-K nor inwardly rectifying I-K are res ponsible for the hypoxia-induced depolarization. In conclusion, PC-12 cells express an I-K(O2), inhibition of which leads to membrane depola rization and increased intracellular Ca2+ making the PC-12 clonal cell line a useful model for studying the molecular and biophysical mechan isms that mediate O-2 chemosensitivity.