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 sensory activity. To begin to understand the cellular mechanisms associated with the actions of NO in the carotid body, we monitored the effects of NO donors on the macroscopic Ca(2+) current in glomus cells isolated from rabbit carotid bodies. Experiments were performed on freshly dissociated glomus cells 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 of 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-carboxyphenyl)-4, 4,5,5-tetramethyl-imidazoline-1-oxyl-3-oxide (carboxy-PTIO; 500 mu M). an NO scavenger, prevented inhibition of the Ca(2+) current by SNP (n = 7), whereas neither superoxide dismutase (SOD; 2,000 U/ml, n = 4), a superoxide scavenger, nor sodium hydrosulfite (SHS; 1 mM, n = 7), a reducing agent, prevented inhibition of the Ca(2+) current by SNP. However, SNP inhibition of the Ca(2+) current was reversible in the presence of either SOD or SHS. These results suggest that NO itself inhibits Ca(2+) current in a reversible manner and that subsequent formation of peroxynitrites results in irreversible inhibition. SNP inhibition of the Ca(2+) 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 Ca(2+) current are mediated, in part, via a cGMP-independent mechanism. N-ethylmaleimide (NEM; 2.5 mM, n = 6) prevented the inhibition of the Ca(2+) current by SNP, indicating that SNP is acting via a modification of sulfhydryl groups on Ca(2+) channel proteins. Norepinephrine (NE; 10 mu M) further inhibited the Ca(2+) current in the presence of NEM (n = 7), implying that NEM did not nonspecifically eliminate Ca(2+) current modulation. Nisoldipine, an L-type Ca(2+) channel blocker (2 mu M, n = 6), prevented the inhibition of Ca(2+) current by SNP, whereas omega-conotoxin GVIA, an N-type Ca(2+) channel blocker (1 mu M, n = 9), did not prevent the inhibition of Ca(2+) current by SNP. These results demonstrate that NO inhibits L-type Ca(2+) channels in adult rabbit glomus cells, in part, due to a modification of calcium channel proteins. The inhibition might provide one plausible mechanism for efferent inhibition of carotid body activity by NO.