EFFECTS OF HYPOXIA ON MEMBRANE-POTENTIAL AND INTRACELLULAR CALCIUM IN RAT NEONATAL CAROTID-BODY TYPE-I CELLS

1. We have studied the effects of hypoxia on membrane potential and [Ca2+](i) in enzymically isolated type I cells of the neonatal rat carotid body (the principal respiratory O-2 chemosensor). Isolated cells were maintained in short term culture (3-36 h) before use. [Ca2+](i) was measured using the Ca2+-sensitive fluoroprobe indo-1. Indo-1 was loaded into cells using the esterified form indo-1 AM. Membrane potential was measured (and clamped) in single isolated type I cells using the perforated-patch (amphotericin B) whole-cell recording technique. 2. Graded reductions in P-O2 from 160 Torr to 38, 19, 8, 5 and 0 Torr induced a graded rise of [Ca2+](i) in both single and clumps of type I cells. 3. The rise of [Ca2+](i) in response to anoxia was 98 % inhibited by removal of external Ca2+ (+ 1 mM EGTA), indicating the probable involvement of Ca2+ influx from the external medium in mediating the anoxic [Ca2+](i) response. 4. The L-type Ca2+ channel antagonist nicardipine (10 mu M) inhibited the anoxic [Ca2+](i) response by 67 %, and the non-selective Ca2+ channel antagonist Ni2+ (2 mM) inhibited the response by 77 %. 5. Under voltage recording conditions, anoxia induced a reversible membrane depolarization (or receptor potential) accompanied, in many cases, by trains of action potentials. These electrical events were coincident with a rapid rise of [Ca2+](i). When cells were voltage clamped close to their resting potential (-40 to -60 mV), the [Ca2+](i) response to anoxia was greatly reduced and its onset was much slower. Under voltage clamp conditions, anoxia also induced a small inward shift in holding current. 6. We conclude that anoxia promotes a rise of [Ca2+](i) in type I cells, principally through voltage-gated Ca2+ entry which occurs in response to the receptor potential and/or concomitant electrical activity. We propose that this mechanism forms the basis for hypoxic chemotransduction in the carotid body.