The effects of thiol modulators on nitrergic nerve- and S-nitrosothiols-induced relaxation in duodenum

Abstract

The aim of this study was to investigate whether thiols are involved in the nitrergic neurotransmission in mouse duodenum. The effects of thiol-modulating agents, ethacrynic acid (100 μM), a non-specific sulfhydryl alkylator, and diamide (100 μM), an alkylating agent that oxidizes protein sulfhydryl groups and depletes intracellular glutathione, on relaxations to nitrergic stimulation (electrical field stimulation, EFS;10 Hz, 25 V, 1 ms, 15 s-train), S-nitrosoglutathione (GSNO; 5 μM), S-nitroso-acetylpenicillamine (SNAP; 5 μM), and S-nitrosocysteine (CysNO; 10 μM) were investigated. Moreover, the effects of buthionine sulfoximine (100 μM), an inhibitor of γ-glutamylcysteine synthetase, and sulfobromophthalein (100 μM), an inhibitor of glutathione-S-transferase, were studied on relaxant responses to EFS and S-nitrosothiols in mouse duodenum. Exogenous free thiol, glutathione (GSH, 100 μM) did not influence relaxation to EFS, GSNO, SNAP, and CysNO. Ethacrynic acid and diamide significantly decreased relaxation of duodenum to EFS, GSNO, SNAP, and CysNO. This inhibition was prevented by addition of GSH. Buthionine sulfoximine and sulfobromophthalein significantly decreased relaxation to EFS and GSNO but did not influence relaxation to SNAP and CysNO. The inhibitory effect of buthionine sulfoximine and sulfobromophthalein on the relaxant response to EFS and GSNO was prevented by addition of GSH. These results suggest that relaxation to nitrergic stimulation is thiol-dependent, and nitrosothiols, possibly S-nitrosoglutathione may play a role, as an intermediate compound in nitrergic neurotransmission in mouse duodenum.