Anoxia differentially modulates multiple K+ currents and depolarizes neonatal rat adrenal chromaffin cells

摘要

class="enumerated" style="list-style-type:decimal">Using perforated-patch, whole cell recording, we investigated the membrane mechanisms underlying O2 chemosensitivity in neonatal rat adrenomedullary chromaffin cells (AMC) bathed in extracellular solution containing tetrodotoxin (TTX; 0.5–1 μm), with or without blockers of calcium entry.Under voltage clamp, low PO2 (0–15 mmHg) caused a graded and reversible suppression in macroscopic outward K⁺ current. The suppression during anoxia (PO2 = 0 mmHg) was ∼35% (voltage step from −60 to +30 mV) and was due to a combination of several factors: (i) suppression of a cadmium-sensitive, Ca²⁺-dependent K⁺ current, IK(CaO2); (ii) suppression of a Ca²⁺-insensitive, delayed rectifier type K⁺ current, IK(VO2); (iii) activation of a glibenclamide- (and Ca²⁺)-sensitive current, IK(ATP).During normoxia (PO2 = 150 mmHg), application of pinacidil (100 μm), an ATP-sensitive potassium channel (KATP) activator, increased outward current density by 45.0 ± 7.0 pA pF⁻¹ (step from −60 to + 30 mV), whereas the KATP blocker glibenclamide (50 μm) caused only a small suppression by 6.3 ± 4.0 pA pF⁻¹. In contrast, during anoxia the presence of glibenclamide resulted in a substantial reduction in outward current density by 24.9 ± 7.9 pA pF⁻¹, which far exceeded that seen in its absence. Thus, activation of IK(ATP) by anoxia appears to reduce the overall K⁺ current suppression attributable to the combined effects of IK(CaO₂₎ and I_K(VO2).Pharmacological tests revealed that I_K(CaO2) was carried predominantly by maxi-K⁺ or BK potassium channels, sensitive to 50–100 nm iberiotoxin; this current also accounted for the major portion (∼60%) of the anoxic suppression of outward current. Tetraethylammonium (TEA; 10–20 mm) blocked all of the anoxia-sensitive K⁺ currents recorded under voltage clamp, i.e. I_K(CaO2), I_K(VO2) and I_K(ATP).Under current clamp, anoxia depolarized neonatal AMC by 10–15 mV from a resting potential of ∼-55 mV. At least part of this depolarization persisted in the presence of either TEA, Cd²⁺, 4-aminopyridine or charybdotoxin, suggesting the presence of anoxia-sensitive mechanisms additionalto those revealed under voltage clamp. In Na⁺/Ca²⁺-free solutions, the membrane hyperpolarized, though at least a portion of the anoxia-induced depolarization persisted.In the presence of glibenclamide, the anoxia-induced depolarization increased significantly to ∼25 mV, suggesting that activation of K_ATP channels may function to attenuate the anoxia-induced depolarization or receptor potential.