Acetylcholine-stimulated changes of membrane potential and intracellular Ca2+ concentration recorded in endothelial cells in situ in the isolated rat aorta.
The intracellular free Ca2+ concentration and membrane potential changes evoked by acetylcholine were recorded from whole-cell patch-clamped endothelial cells in situ in the isolated rat aorta. The endothelium had a resting membrane potential of -52 +/- 3 mV (SEM, range -35 mV to -76 mV n = 34) and a low input resistance (32 - 54 M omega). The membrane potential hyperpolarised by 3-30 mV on continuous application of acetylcholine at concentrations that produced endothelium-dependent relaxations in isolated rat aortic rings (range 1-500 nM). The response often comprised complex fluctuations of hyperpolarised membrane potential. Calcium concentration was measured with the fluorescent indicator furaptra, which has a wide range and minimises Ca2+ buffering. Acetylcholine evoked an initial rapid elevation of intracellular Ca2+ concentration, peaking in the range 6-35 microM, which declined with a half time of approximately 6 s, followed by repetitive [Ca2+] spikes of amplitude 2-18 microM in 23 of 34 cells. The initial [Ca2+] transient and hyperpolarisation were unaffected by removal of external Ca2+, whilst subsequent [Ca2+] spikes and maintained hyperpolarisations required the presence of external Ca2+. Both the hyperpolarisation and Ca2+ responses elicited by acetylcholine were abolished by atropine (1 microM). These results show that endothelial cells in situ exhibit large, fast repetitive [Ca2+] spikes in response to extracellular acetylcholine.
Division of Neurophysiology and Neuropharmacology, National Institute for Medical Research, Mill Hill, London, UK.
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Carter
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Ogden
D000109:Acetylcholine / Q000494:pharmacology*; D000818:Animals; D001013:Aorta, Thoracic / Q000378:metabolism* / Q000502:physiology; D002118:Calcium / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000378:metabolism / Q000502:physiology; D007425:Intracellular Membranes / Q000378:metabolism / Q000502:physiology; D008564:Membrane Potentials / Q000187:drug effects*; D051381:Rats; D017208:Rats, Wistar
D000109:Acetylcholine; D002118:Calcium