Nitric oxide inhibits L-type Ca2+ current in glomus cells of the rabbit carotid body via a cGMP-independent mechanism

Previous studies have shown that nitric oxide (NO) inhibits carotid body se nsory activity. To begin to understand the cellular mechanisms associated w ith the actions of NO in the carotid body, we monitored the effects of NO d onors on the macroscopic Ca2+ current in glomus cells isolated from rabbit carotid bodies. Experiments were performed on freshly dissociated glomus ce lls from adult rabbit carotid bodies using the whole cell configuration of the patch-clamp technique. The NO donors sodium nitroprusside (SNP; 600 mu M, n = 7) and spermine nitric oxide (SNO; 100 mu M, n = 7) inhibited the Ca 2+ current in glomus cells in a voltage independent manner. These effects o f NO donors were rapid in onset and peaked within 1 or 2 min. In contrast, the outward K+ current was unaffected by SNP (600 mu M, n = 6), indicating that the inhibition by SNP was not a nonspecific membrane effect. 2-(4-carb oxyphenyl)-4, 4,5,5-tetramethyl-imidazoline-1-oxyl-3-oxide (carboxy-PTIO; 5 00 mu M). an NO scavenger, prevented inhibition of the Ca2+ current by SNP (n = 7), whereas neither superoxide dismutase (SOD; 2,000 U/ml, n = 4), a s uperoxide scavenger, nor sodium hydrosulfite (SHS; 1 mM, n = 7), a reducing agent, prevented inhibition of the Ca2+ current by SNP. However, SNP inhib ition of the Ca2+ current was reversible in the presence of either SOD or S HS. These results suggest that NO itself inhibits Ca2+ current in a reversi ble manner and that subsequent formation of peroxynitrites results in irrev ersible inhibition. SNP inhibition of the Ca2+ current was not affected by 30 mu M LY 83,583 (n = 7) nor was it mimicked by 600 mu M 8-bromoguanosine 3':5'-cyclic monophosphate (8-Br-cGMP; n = 6), suggesting that the effects of NO on the Ca2+ current are mediated, in part, via a cGMP-independent mec hanism. N-ethylmaleimide (NEM; 2.5 mM, n = 6) prevented the inhibition of t he Ca2+ current by SNP, indicating that SNP is acting via a modification of sulfhydryl groups on Ca2+ channel proteins. Norepinephrine (NE; 10 mu M) f urther inhibited the Ca2+ current in the presence of NEM (n = 7), implying that NEM did not nonspecifically eliminate Ca2+ current modulation. Nisoldi pine, an L-type Ca2+ channel blocker (2 mu M, n = 6), prevented the inhibit ion of Ca2+ current by SNP, whereas omega-conotoxin GVIA, an N-type Ca2+ ch annel blocker (1 mu M, n = 9), did not prevent the inhibition of Ca2+ curre nt by SNP. These results demonstrate that NO inhibits L-type Ca2+ channels in adult rabbit glomus cells, in part, due to a modification of calcium cha nnel proteins. The inhibition might provide one plausible mechanism for eff erent inhibition of carotid body activity by NO.