A redox-based mechanism for the contractile and relaxing effects of NO in the guinea-pig gall bladder

1. The purpose of this study was to determine the effects of sodium nitropr usside (SNP), 2,2'-(hydroxynitrosohydrazino)bis-ethanamine (DETA/NO) and 3- morpholinosydnonimine (BIN-1), NO donors which yield different NO reactive species (NO+, NO . and peroxynitrite, respectively), as well as exogenous p eroxynitrite, on gall bladder contractility. 2. Under resting tone conditions, NP induced a dose-dependent contraction w ith a maximal effect (10.3 +/- 0.7 mN, S.E.M.) at 1 mM. Consistent with the se findings, SNP caused a concentration-dependent depolarization of gall bl adder smooth muscle. The excitatory effects of SNP were dependent on extrac ellular calcium entry through L-type Ca2+ channels. Furthermore, the contra ction and depolarization were sensitive to tyrosine kinase blockade, and an associated increase in tyrosine phosphorylation was detected in Western bl ot studies. 3. DETA/NO induced dose-dependent relaxing effects. These relaxations were sensitive to the guanylyl cyclase inhibitor 1H-[1,2,4]oxidiazolo[4,3-a]quin oxaline-1-one (ODQ, 2 muM) but they were not altered by treatment with the potassium channel blockers tetraethylammoniun (TEA, 5 mM) and 4-aminopyridi ne (4-AP, 5 mM). 4. When tested in a reducing environment (created by 2.5 mar 1,4-dithiothre itol, DTT), SNP caused a relaxation of gall bladder muscle strips. Similarl y, the SNP-induced contraction was converted to a relaxation, and associate d hyperpolarization, when DTT was added during the steady state of an SNP-i nduced response. 5. SIN-1 (0.1 mM), which hlas been shown to release peroxynitrite, induced relaxing effects that were enhanced by superoxide dismutase (SOD, 50 U ml(- 1)). The relaxations induced by either SIN-1 alone or SIN-1 in the presence of SOD were strengthened by catalase (1000 U ml(-1)) and abolished by ODQ pretreatment. However, exogenous peroxynitrite induced a concentration depe ndent contraction, which was dependent on activation of leukotriene (LT) me tabolism and extracellular calcium. The peroxynitrite-induced contraction w as abolished in the presence of the peroxynitrite scavenger melatonin. Thes e results suggest that SIN-1 behaves as an NO . rather than a peroxynitrite source. 6. We conclude that, depending on the redox state, NO has opposing effects on the motility of the gall bladder, being a relaxing agent when in NO . fo rm and a contracting agent when in NO+ or peroxynitrite redox species form. Knowledge of the contrasting effects of the different redox forms of NO ca n clarify our understanding of the effects of NO donors on gall bladder and other smooth muscle cell types.