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Key points Endothelial function in resistance vessels entails Ca2+ and electrical signalling to promote vasodilatation and increase tissue blood flow. Whether membrane potential (V-m) governs intracellular calcium concentration ([Ca2+](i)) of the endothelium remains controversial. [Ca2+](i) and V-m were evaluated simultaneously during intracellular current injection using intact endothelial tubes freshly isolated from mouse skeletal muscle resistance arteries. [Ca2+](i) did not change during hyperpolarization or depolarization under resting conditions. However in the presence of 100nM ACh (approximate to EC50), [Ca2+](i) increased during hyperpolarization and decreased during depolarization. These responses required extracellular Ca2+ and were attenuated by half with genetic ablation of TRPV4 channels. In native microvascular endothelium, half-maximal stimulation of muscarinic receptors enables V-m to govern [Ca2+](i) by activating Ca2+-permeable channels in the plasma membrane. This effect of V-m is absent at rest and can be masked during maximal receptor stimulation. AbstractIn resistance arteries, coupling a rise of intracellular calcium concentration ([Ca2+](i)) to endothelial cell hyperpolarization underlies smooth muscle cell relaxation and vasodilatation, thereby increasing tissue blood flow and oxygen delivery. A controversy persists as to whether changes in membrane potential (V-m) alter endothelial cell [Ca2+](i). We tested the hypothesis that V-m governs [Ca2+](i) in endothelium of resistance arteries by performing Fura-2 photometry while recording and controlling V-m of intact endothelial tubes freshly isolated from superior epigastric arteries of C57BL/6 mice. Under resting conditions, [Ca2+](i) did not change when V-m shifted from baseline (approximate to-40mV) via exposure to 10M NS309 (hyperpolarization to approximate to-80mV), via equilibration with 145mm [K+](o) (depolarization to approximate to-5mV), or during intracellular current injection (+/- 0.5 to 5nA, 20s pulses) while V-m changed linearly between approximate to-80mV and +10mV. In contrast, during the plateau (i.e. Ca2+ influx) phase of the [Ca2+](i) response to approximately half-maximal stimulation with 100nm ACh (approximate to EC50), [Ca2+](i) increased as V-m hyperpolarized below -40mV and decreased as V-m depolarized above -40mV. The magnitude of [Ca2+](i) reduction during depolarizing current injections correlated with the amplitude of the plateau [Ca2+](i) response to ACh. The effect of hyperpolarization on [Ca2+](i) was abolished following removal of extracellular Ca2+, was enhanced subtly by raising extracellular [Ca2+] from 2mm to 10mm and was reduced by half in endothelium of TRPV4(-/-) mice. Thus, during submaximal activation of muscarinic receptors, V-m can modulate Ca2+ entry through the plasma membrane in accord with the electrochemical driving force.
- 出版日期2015-10-15
