Abstract
Various cardiorespiratory diseases (e.g. congestive heart failure, emphysema) result in systemic hypoxia and patients consequently demonstrate adaptive cellular responses which predispose them to conditions such as pulmonary hypertension and stroke. Central to many affected excitable tissues is activity of large conductance, Ca2+-activated K+ (maxiK) channels. We have studied maxiK channel activity in HEK293 cells stably co-expressing the most widely distributed of the human alpha- and beta-subunits that constitute these channel following maneuvers which mimic severe hypoxia. At all [Ca2+]i, chronic hypoxia (approximately 18 mm Hg, 72 h) increased K+ current density, most markedly at physiological [Ca2+]i K+ currents in cells cultured in normoxia showed a [Ca2+]i-dependent sensitivity to acute hypoxic inhibition ( approximately 25 mm Hg, 3 min). However, chronic hypoxia dramatically changed the Ca2+ sensitivity of this acute hypoxic inhibitory profile such that low [Ca2+]i could sustain an acute hypoxic inhibitory response. Chronic hypoxia caused no change in alpha-subunit immunoreactivity with Western blotting but evoked a 3-fold increase in beta-subunit expression. These observations were fully supported by immunocytochemistry, which also suggested that chronic hypoxia augmented alpha/beta-subunit co-localization at the plasma membrane. Using a novel nuclear run-on assay and RNase protection we found that chronic hypoxia did not alter mRNA production rates or steady-state levels, which suggests that this important environmental cue modulates maxiK channel function via post-transcriptional mechanisms.
Highlights
Crucial to the cellular and physiological response to acute perturbation of systemic and/or pulmonary O2 levels is the rapid inhibition of Kϩ channels by hypoxia
We have recently demonstrated at the single channel level that a recombinant human maxiK channel can be rapidly and reversibly inhibited by acute hypoxia; this inhibition is underlain by hypoxia-evoked depression in unitary conductance, slowed channel activation kinetics, reduced open-state probability, and altered channel sensitivity to intracellular calcium concentration ([Ca2ϩ]i) (27)
Potential Molecular Mechanism of Chronic Hypoxic Regulation—Through the use of a mammalian recombinant expression system, our whole cell patch-clamp data have demonstrated that acute hypoxia inhibits maxiK channels of known molecular identity in a manner qualitatively similar to that previously observed in excised inside-out patches (27)
Summary
Crucial to the cellular and physiological response to acute perturbation of systemic and/or pulmonary O2 levels is the rapid inhibition of Kϩ channels by hypoxia (see Ref. 1 for recent review). Hypoxic inhibition of native maxiK channel activity has been demonstrated in carotid body (4, 18, 19), pulmonary arteriolar smooth muscle (20), chromaffin cells (21), and central neurons (20, 22). Their contribution to carotid body, chromaffin cell, and central neuronal function is well supported, some controversy still surrounds their involvement in pulmonary vasoconstriction (15) and there is good evidence for both delayed rectifier (23) and tandem P domain Kϩ channels in the response (24); the latter observation is fully supported by our recent reports of O2 sensitivity of the recombinant human tandem P domain channels, hTASK1 (25), and hTASK3 (26). Individuals suffering from a variety of cardiorespiratory diseases (such as chronic obstructive pulmonary disease, apnea of sleep, emphysema, congestive heart failure, and stroke) un-
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