Electrophysiological studies suggest that activation of large-conductance Ca-activated K channels (KCa) with nitric oxide (NO) causes hyperpolarization and relaxation of smooth muscle. We determined whether KCa blockers decreased relaxation to the NO donors S-nitroso-N-acetylpenicillamine (SNAP) and 3-morpholinosydonimine-hydrochloride (SIN-1) in isolated segments from main pulmonary artery (MPA), its left branch (LPA), aorta (Ao), carotid artery (CA), and trachea (Tr). NO donors caused concentration-dependent relaxation of tissues precontracted with histamine whereas the inactive carrier molecule C88-3934 was without effect. The rank order profiles of SNAP and SIN-1 sensitivity were CA = Ao = MPA > LPA = Tr. Compared with histamine, 80 mM KCl precontraction caused variable reductions in tissue sensitivity and maximum relaxation to SNAP. The KCa antagonists charybdotoxin, iberiotoxin, and tetraethylammonium decreased sensitivity to SNAP and SIN-1 2- to 11-fold in MPA, LPA, and Tr, with variable shifts in Ao and CA. The effect of iberiotoxin was not altered by removing the endothelium or epithelium. Furthermore, charybdotoxin or iberiotoxin did not alter basal or SNAP-stimulated guanosine 3',5'-cyclic monophosphate content. Glibenclamide, noxiustoxin, and leiurotoxin I, antagonists of ATP-dependent, delayed rectifier, and small-conductance KCa channels, respectively, had no effect. In conclusion, antagonists of KCa decrease NO donor-mediated relaxation of pulmonary arterial and tracheal smooth muscle.

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We studied the effects of cholesterol enrichment on arterial function by evaluating its effects on 45Ca2+ uptake and tension development in the carotid artery of the rabbit. Arterial segments were enriched with cholesterol in vitro, using media containing liposomes composed of free (unesterified) cholesterol (FC) and phospholipid (PL) in a 2:1 molar ratio. Control segments were simultaneously perfused with 0.5:1 liposomal medium to compare the possible effects of PL. Rings from these arteries were then tested for basal and activated Ca2+ uptake and for contractile responses to norepinephrine (NE) and KCl. We found elevated 45Ca2+ uptake under basal and NE-activated conditions along with an increased contractile sensitivity (4-fold) to NE. These alterations correlated with a 78% increase in the FC/PL ratio reflecting cholesterol enrichment of cellular membranes. Cholesterol enrichment did not alter resting or maximal tensions, K+-activated Ca2+ uptake, or contractile sensitivity to K+. Pretreatment with 1 microM diltiazem abolished the cholesterol-induced increase in basal as well as NE-activated 45Ca2+ uptake but had no effect on either uptake in control vessels. These studies suggest that excess membrane cholesterol selectively increases NE contractile sensitivity by increasing basal or NE-activated Ca2+ influx (or both) as a result of fundamental alteration in the calcium channels in arterial smooth muscle cell membrane.

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