Endothelin inhibits thick ascending limb chloride flux via ETB receptor-mediated NO release

Endothelin-1 (ET-1) inhibits transport in various nephron segments, and the thick ascending limb of the loop of Henle (TALH) expresses ET-1 receptors. In many tissues, activation of ETB receptors stimulates release of NO, and we recently reported that endogenous NO inhibits TALH chloride flux (J(Cl) ). However, the relationship between ET-1 and NO in the control of nephron transport has not been extensively studied. We hypothesized that ET-1 decre ases NaCl transport by cortical TALHs through activation of ETB receptors a nd release of NO. Exogenous ET-1 (1 nM) decreased J(Cl) from 118.3 +/- 15.0 to 62.7 +/- 13.6 pmol . mm(-1) . min(-1) (48.3 +/- 8.2% reduction), wherea s removal of ET-1 increased J(Cl) in a separate group of tubules from 87.6 +/- 10.7 to 115.2 +/- 10.3 pmol . mm(-1) . min(-1) (34.5 +/- 6.2% increase) . To determine whether NO mediates the inhibitory effects of ET-1 on J(Cl), we examined the effect of inhibiting of NO synthase (NOS) with N-G-nitro-L -arginine methyl ester (L-NAME) on ET-1-induced changes in J(Cl). L-NAME (5 mM) completely prevented the ET-1-induced reduction in J(Cl), whereas D-NA ME did not. L-NAME alone had no effect on J(Cl). These data suggest that th e effects of ET-1 are mediated by NO. Blockade of ETB receptors with BQ-788 prevented the inhibitory effects of 1 nM ET-1. Activation of ETB receptors with sarafotoxin S6c mimicked the inhibitory effect of ET-1 on J(Cl) (from 120.7 +/- 12.6 to 75.4 +/- 13.3 pmol . mm(-1) . min(-1)). In contrast, ETA receptor antagonism with BQ-610 did not prevent ET-1-mediated inhibition o f TALH J(Cl) (from 96.5 +/- 10.4 to 69.5 +/- 8.6 pmol . mm(-1) . min(-1)). Endothelin increased intracellular calcium from 96.9 +/- 14.0 to 191.4 +/- 11.9 nM, an increase of 110.8 +/- 26.1%. We conclude that exogenous endothe lin indirectly decreases TALH J(Cl) by activating ETB receptors, increasing intracellular calcium concentration, and stimulating NO release. These dat a suggest that endothelin acts as a physiological regulator of TALH NO synt hesis, thus inhibiting chloride transport and contributing to the natriuret ic effects of ET-1 observed in vivo.