Role of NO in endothelin-regulated drug transport in the renal proximal tubule. Am J Physiol Renal Physiol 282: F458-F464, 2002; 10.1152/ajprenal.00173.2001.-We previously demonstrated in intact killifish renal proximal tubules that endothelin (ET), acting through an ETB receptor and protein kinase C (PKC), reduced transport mediated by multidrug resistance-associated protein 2 (Mrp2), i.e., luminal accumulation of fluorescein methotrexate (FL-MTX) (Masereeuw R, Terlouw SA, Van Aubel RAMH,

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... l FGM, and Miller DS. Mol Pharmacol 57: 59-67, 2000). In the present study, we used confocal microscopy and quantitative image analysis to measure Mrp2-mediated transport of FL-MTX in killifish tubules as an indicator of the status of this ET-fired, intracellular signaling pathway. Exposing tubules to sodium nitroprusside (SNP), a nitric oxide (NO) donor, signaled a reduction in luminal accumulation of FL-MTX, which suggested pathway activation. N G -monomethyl-L-arginine (L-NMMA), an NO synthase inhibitor, blocked the action of ET-1 on transport. Because SNP effects on transport were blocked by bisindoylmaleide, a PKC-selective inhibitor, but not by RES-701-1, an ET B-receptor antagonist, generation of NO occurred after ET B receptor signaling but before PKC activation. NO generation was implicated in the actions of several nephrotoxicants, i.e., diatrizoate, gentamicin, amikacin, HgCl 2, and CdCl 2, each of which decreased Mrp2-mediated transport by activating ET signaling. For each nephrotoxicant, decreased FL-MTX transport was prevented when tubules were exposed to L-NMMA. ET-1 and each nephrotoxicant stimulated NO production by the tubules, as determined by a fluorescencebased assay. Together, the data show that NO generation follows ET binding to the basolateral ET B receptor and that, in activating the ET-signaling pathway, nephrotoxicants produce NO, a molecule that could contribute to subsequent toxic effects. multidrug resistance-associated protein 2; calcium; endothelin signaling; protein kinase C; xenobiotic transport; nitric oxide METABOLISM AND EXCRETION provide a first line of defense against the wide variety of potentially toxic chemicals to which we are continually exposed. In the kidney, the Address for reprint requests and other correspondence: R. Masereeuw,