Mechanisms of nitric oxide-independent relaxations induced by carbachol and acetylcholine in rat isolated renal arteries

1. In rat isolated renal artery segments contracted with 0.1 μM phenylephrine and in the presence of the NO synthase inhibitor N(ω)-nitro-L-arginine methyl ester (L-NAME), carbachol and acetylcholine produced endothelium-dependent relaxations. The mechanisms underlying these relaxations were studied. 2. These relaxations were not affected by ODQ (1H-[1,2,4]oxadiazolo[4,3,-a]quinoxalin-1-one) or indomethacin. In arteries contracted with 20-30 mM K⁺, L-NAME-resistant relaxations induced by carbachol and acetylcholine were virtually absent. 3. The Na⁺-K⁺ ATPase inhibitor ouabain reduced these relaxations in a concentration-dependent manner. 4. In K⁺-free media, addition of K⁺ (5 mM) produced 90.5 ± 3.9% (n = 3) relaxation of phenylephrine-induced tone. This relaxation was endothelium-independent and ouabain-sensitive. 5. Tetraethylammonium (TEA), charybdotoxin (ChTX) and iberiotoxin (IbTX) reduced the sensitivity of carbachol-induced relaxations, but did not change the maximal response. These relaxations were not altered by 4-aminopyridine (4-AP), glibenclamide or apamin. Acetylcholine (1 μM)-induced relaxation was reduced by ChTX, but not by TEA or IbTX. 6. The cytochrome P450 inhibitor miconazole, but not 17-octadecynoic acid, reduced the sensitivity of carbachol-induced relaxations, without changing the maximal response. 7. In conclusion, in rat isolated renal arteries, acetylcholine and carbachol produced a non-NO/non-PGI₂ relaxation which is mediated by an endothelium-derived hyperpolarizing factor (EDHF). This factor does not appear to be a cytochrome P450 metabolite. The inhibition by ouabain of these relaxations suggests the possible involvement of Na⁺-K⁺ ATPase activation in EDHF responses, although other mechanisms cannot be totally ruled out.