Osmolar regulation of endothelin-1 production by the inner medullary collecting duct

Endothelin-1 (ET-1) is an autocrine inhibitor of collecting duct (CD) Na+ and water reabsorption. CD ET-1 production is increased by a high salt diet and is important in promoting a natriuretic response. The mechanisms by which a high salt diet enhances CD ET-1 are being uncovered. In particular, elevated tubule fluid flow, as occurs in salt loading, enhances CD ET-1 synthesis. Tubule fluid solute content and interstitial osmolality can also be altered by a high salt diet, however their effect on CD ET-1 alone, or in combination with flow, is poorly understood. ET-1 mRNA production by a mouse inner medullary CD cell line (mIMCD3) in response to changing flow and/or osmolality was assessed. Flow or hyperosmolality (using NaCl, mannitol or urea) individually caused an ~2-fold increase in ET-1 mRNA, while flow and hyperosmolality together increased ET-1 mRNA by ~14 fold. The hyperosmolality effect alone and the synergistic effect of flow + hyperosmolality was inhibited by chelation of intracellular Ca2+, however were not altered by blockade of downstream Ca2+-signaling pathways (calcineurin or NFATc), inhibition of cellular Ca2+ entry channels (purinergic receptors or polycystin-2), or blockade of the epithelial Na+ channel. Inhibition of NFAT5 with rottlerin or NFAT5 siRNA greatly reduced the stimulatory effect of osmolality alone and osmolality + flow on mIMCD3 ET-1 mRNA levels. Both flow and osmolality individually and synergistically stimulate mIMCD3 ET-1 mRNA content. These findings may be relevant to explaining high salt diet induction of CD ET-1 production.