Endothelium-derived factors and hyperpolarization of the carotid artery of the guinea-pig

Abstract

1 Transmembrane potentials were recorded from isolated carotid arteries of the guinea-pig superfused with modified Krebs-Ringer bicarbonate solution. Smooth muscle cells were impaled from the adventitial side with intracellular glass microelectrodes filled with KCl (30-80 MΩ). 2 Acetylcholine (1 μM) in the presence of inhibitors of nitric oxide synthase, (N(ω)-nitro-L-arginine (L-NOARG) 100 μM) and cyclo-oxygenase, (indomethacin 5 μM) induced an endothelium-dependent hyperpolarization (-18.9 ± 1.6 mV, n = 15). 3 In the presence of these two inhibitors, S-nitroso-L-glutathione (10 μM), sodium nitroprusside (10 μM), 3-morpholinosydnonimine (SIN-1, 10 μM) and iloprost (0.1 μM) induced endotheliumin-dependent hyperpolarizations of the smooth muscle cells (respectively: -16.0 ± 2.3, -16.3 ± 3.4, -12.8 ± 2.0 and -14.5 ± 1.5 mV, n = 4-6). 4 The addition of glibenclamide (1 μM) did not influence the acetylcholine-induced L-NOARG/ indomethacin-resistant hyperpolarization(-18.0 ± 1.8 mV, n = 10). In contrast, the responses induced by S-nitroso-L-glutathione, sodium nitroprusside, SIN-1 and iloprost were abolished (changes in membrane potential: -0.8 ± 1.1, 1.3 ± 3.9, 4.5 ± 4.6 and 0.3 ± 0.8 mV respectively, n = 4-5). 5 In the presence of NO synthase and cyclo-oxygenase inhibitors, charybdotoxin (0.1 μM) or apamin (0.5 μM) did not influence the hyperpolarization produced by acetylcholine. However, in the presence of the combination of charybdotoxin and apamin, the acetylcholine-induced L-NOARG/indomethacin-resistant hyperpolarization was converted to a depolarization (4.4 ± 1.2 mV, n = 20) while the endothelium-independent hyperpolarizations induced by S-nitroso-L-glutathione, sodium nitroprusside, SIN-1 and iloprost were not affected significantly (respectively: -20.4 ± 3.4, -22.5 ± 4.9, -14.5 ± 4.7 and -14.5 ± 0.5 mV, n = 4-5). 6 In the presence of the combination of charybdotoxin and apamin and in the absence of L-NOARG and indomethacin, acetylcholine induced a hyperpolarization (-19.5 ± 3.7 mV, n = 4). This hyperpolarization induced by acetylcholine was not affected by the addition of indomethacin (-18.3 ± 4.6 mV, n = 3). In the presence of the combination of charybdotoxin, apamin and L-NOARG (in the absence of indomethacin), acetylcholine, in 5 out of 7 vessels, still produced hyperpolarization which was not significantly smaller (-9.1 ± 5.6 mV, n = 7) than the one observed in the absence of L-NOARG. 7 These findings suggest that, in the guinea-pig isolated carotid artery, the endothelium-independent hyperpolarizations induced by NO donors and iloprost involve the opening of K(ATP) channels while the acetylcholine-induced endothelium-dependent hyperpolarization (resistant to the inhibition of NO-synthase and cyclo-oxygenase) involves the opening of Ca2+-activated potassium channel(s). Furthermore, in this tissue, acetylcholine induces the simultaneous release of various factors from endothelial origin: hyperpolarizing factors (NO, endothelium derived hyperpolarizing factor (EDHF) and prostaglandins) and possibly a depolarizing factor.