Spectrophotometric assay of renal ouabain-resistant Na(+)-ATPase and its regulationby leptin and dietary-induced obesity.
Journal Title: Acta Biochimica Polonica - Year 2004, Vol 51, Issue 4
Abstract
Apart from Na(+),K(+)-ATPase, a second sodium pump, Na(+)-stimulated, K(+)-independentATPase (Na(+)-ATPase) is expressed in proximal convoluted tubule of the mammalian kidney. The aim ofthis study was to develop a method of Na(+)-ATPase assay based on the method previously used by us tomeasure Na(+),K(+)-ATPase activity. The ATPase activity was assayed as the amount of inorganic phosphateliberated from ATP by isolated microsomal fraction. Na(+)-ATPase activity was calculated as the differencebetween the activities measured in the presence and in the absence of 50 mM NaCl. Na(+)-ATPase activitywas detected in the renal cortex (3.5 +/- 0.2 mumol phosphate/h per mg protein), but not in the renalmedulla. Na(+)-ATPase was not inhibited by ouabain or an H(+),K(+)-ATPase inhibitor, Sch 28080, but wasalmost completely blocked by 2 mM furosemide. Leptin administered intraperitoneally (1 mg/kg) decreasedthe Na(+),K(+)-ATPase activity in the renal medulla at 0.5 and 1 h by 22.1% and 27.1%, respectively,but had no effect on Na(+)-ATPase in the renal cortex. Chronic hyperleptinemia induced by repeated subcutaneousleptin injections (0.25 mg/kg twice daily for 7 days) increased cortical Na(+),K(+)-ATPase, medullaryNa(+),K(+)-ATPase and cortical Na(+)-ATPase by 32.4%, 84.2% and 62.9%, respectively. In rats with dietary-inducedobesity, the Na(+),K(+)- ATPase activity was higher in the renal cortex and medulla by 19.7% and 34.3%,respectively, but Na(+)-ATPase was not different from control. These data indicate that both renal Na(+)-dependentATPases are separately regulated and that up-regulation of Na(+)-ATPase may contribute to Na(+) retentionand arterial hypertension induced by chronic hyperleptinemia.
Authors and Affiliations
Grazyna Wójcicka, Jarosław Nazar, Anna Jamroz-Wiśniewska, Jerzy Bełtowski
Keywords
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