Ouabain modulation of endothelial calcium signaling in descending vasa recta

Abstract

Using fura 2-loaded vessels, we tested whether ouabain modulates endothelial cytoplasmic calcium concentration ([Ca 2+ ] CYT ) in rat descending vasa recta (DVR). Over a broad range between 10 −10 and 10 −4 M, ouabain elicited biphasic peak and plateau [Ca 2+ ] CYT elevations. Blockade of voltage-gated Ca 2+ entry with nifedipine did not affect the response to ouabain mitigating against a role for myo-endothelial gap junctions. Reduction of extracellular Na + concentration ([Na + ] o ) or Na + /Ca 2+ exchanger (NCX) inhibition with SEA-0400 (10 −6 M) elevated [Ca 2+ ] CYT , supporting a role for NCX in the setting of basal [Ca 2+ ] CYT . SEA-0400 abolished the [Ca 2+ ] CYT response to ouabain implicating NCX as a mediator. The transient peak phase of [Ca 2+ ] CYT elevation that followed either ouabain or reduction of [Na + ] o was abolished by 2-aminoethoxydiphenyl borate (5 × 10 −5 M). Cation channel blockade with La 3+ (10 μM) or SKF-96365 (10 μM) also attenuated the ouabain-induced [Ca 2+ ] CYT response. Ouabain pretreatment increased the [Ca 2+ ] CYT elevation elicited by bradykinin (10 −7 M). We conclude that inhibition of ouabain-sensitive Na + -K + -ATPase enhances DVR endothelial Ca 2+ store loading and modulates [Ca 2+ ] CYT signaling through mechanisms that involve NCX, Ca 2+ release, and cation channel activation.

Keywords

Affiliated Institutions

• University of Maryland, Baltimore US

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