title: Membrane potential governs calcium influx into microvascular endothelium: integral role for muscarinic receptor activation.
abstract: In resistance arteries, coupling a rise of intracellular calcium concentration ([Ca(2+)]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 [Ca(2+)]i. We tested the hypothesis that V(m) governs [Ca(2+)]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, [Ca(2+)]i did not change when V(m) shifted from baseline (∼-40 mV) via exposure to 10 μM NS309 (hyperpolarization to ∼-80 mV), via equilibration with 145 mm [K(+)]o (depolarization to ∼-5 mV), or during intracellular current injection (±0.5 to 5 nA, 20 s pulses) while V(m) changed linearly between ∼-80 mV and +10 mV. In contrast, during the plateau (i.e. Ca(2+) influx) phase of the [Ca(2+)]i response to approximately half-maximal stimulation with 100 nm ACh (∼EC50), [Ca(2+)]i increased as V(m) hyperpolarized below -40 mV and decreased as V(m) depolarized above -40 mV. The magnitude of [Ca(2+)]i reduction during depolarizing current injections correlated with the amplitude of the plateau [Ca(2+)]i response to ACh. The effect of hyperpolarization on [Ca(2+)]i was abolished following removal of extracellular Ca(2+), was enhanced subtly by raising extracellular [Ca(2+)] from 2 mm to 10 mm and was reduced by half in endothelium of TRPV4(-/-) mice. Thus, during submaximal activation of muscarinic receptors, V(m) can modulate Ca(2+) entry through the plasma membrane in accord with the electrochemical driving force.
authors.affiliation: Department of Medical Pharmacology and Physiology, University of Missouri, Columbia, MO, 65212, USA.
authors.forename: Erik J
authors.initials: EJ
authors.lastname: Behringer
authors.affiliation: Department of Medical Pharmacology and Physiology, University of Missouri, Columbia, MO, 65212, USA.|Dalton Cardiovascular Research Center, Columbia, MO, 65211, USA.
authors.forename: Steven S
authors.initials: SS
authors.lastname: Segal
mesh_terms: D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000502:physiology*; D042783:Endothelial Cells / Q000502:physiology; D004730:Endothelium, Vascular / Q000502:physiology*; D019074:Epigastric Arteries / Q000502:physiology; D007211:Indoles / Q000494:pharmacology; D008297:Male; D008564:Membrane Potentials / Q000502:physiology*; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D055806:Microvessels / Q000502:physiology*; D010091:Oximes / Q000494:pharmacology; D011189:Potassium Chloride / Q000494:pharmacology; D011976:Receptors, Muscarinic / Q000502:physiology*; D050916:TRPV Cation Channels / Q000819:agonists / Q000037:antagonists & inhibitors / Q000502:physiology
chemical_list: C493327:6,7-dichloro-1H-indole-2,3-dione 3-oxime; D007211:Indoles; D010091:Oximes; D011976:Receptors, Muscarinic; D050916:TRPV Cation Channels; C486576:Trpv4 protein, mouse; D011189:Potassium Chloride; D000109:Acetylcholine; D002118:Calcium