The research proposed in this application deals with the control of urinary acidification in the collecting tubule (CT). We propose that this process is controlled by two pumps, an H-ATPase and an H-K-ATPase. We will characterize these pumps, both functionally and biochemically. Two hypotheses will be tested. Hypothesis I - Acidification in the CT is mediated by two proton pumps, an H-ATPase and an H-K-ATPase. The experiments testing this hypothesis will be performed in rat and rabbit nephron fragments. Considerable data concerning the H-ATPase is already available, much less is known about the H-K-ATPase. To show that these enzymes mediate distal acidification it is necessary to show that their activities change appropriately with acid-base perturbations and that their inhibition reduces acidification. We will study inhibition of the H-ATPase with bafilomycin A and N-ethylmaleimide; H-K-ATPase activity will be studied using vanadate, SCH28080, or K removal. The roles of aldosterone, K, and Cl in modulating enzyme activity will be examined. Hypothesis II - Inhibition of the H-K-ATPase results in hypokalemic distal renal tubular acidosis. These studies will be performed in rats and will employ clearance techniques, measurement of enzyme activities along the nephron, and functional studies in isolated tubules. We will study the effects of inhibiting the H-K-ATPase with vanadate and SCH28080. New data show, as predicted, that vanadate induces hypokalemic distal renal tubular acidosis. Other models of distal renal tubular acidosis will also be studied. By studying alterations in whole animal acid-base status and acid excretion, enzyme activities, net acidification in isolated tubules, and cell pH recovery when basolateral proton extension is blocked, we hope to elucidate the relationship between ATPase activity and tubular function.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
2R01DK036199-08A4
Application #
2139751
Study Section
Special Emphasis Panel (ZRG7-SSS-5 (02))
Project Start
1985-01-01
Project End
1998-06-30
Budget Start
1994-07-01
Budget End
1995-06-30
Support Year
8
Fiscal Year
1994
Total Cost
Indirect Cost
Name
Texas Tech University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
609980727
City
Lubbock
State
TX
Country
United States
Zip Code
79430
Hayward, A L; Hinojos, C A; Nurowska, B et al. (1999) Altered sodium pump alpha and gamma subunit gene expression in nephron segments from hypertensive rats. J Hypertens 17:1081-7
Eiam-Ong, S; Sabatini, S (1999) Age-related changes in renal function, membrane protein metabolism, and Na,K-ATPase activity and abundance in hypokalemic F344 x BNF(1) rats. Gerontology 45:254-64
Sabatini, S (1999) Experimental studies in distal urinary acidification: bringing the bedside to the bench. Semin Nephrol 19:188-94
Nguyen, A T; Hayward-Lester, A; Sabatini, S et al. (1998) Renal Na+, K+-ATPase in SHR: studies of activity and gene expression. Clin Exp Hypertens 20:641-56
Wesson, D E (1998) Na/H exchange and H-K ATPase increase distal tubule acidification in chronic alkalosis. Kidney Int 53:945-51
Wesson, D E (1998) Dietary acid increases blood and renal cortical acid content in rats. Am J Physiol 274:F97-103
Hayward, A L; Oefner, P J; Sabatini, S et al. (1998) Modeling and analysis of competitive RT-PCR. Nucleic Acids Res 26:2511-8
Wesson, D E; Simoni, J; Green, D F (1998) Reduced extracellular pH increases endothelin-1 secretion by human renal microvascular endothelial cells. J Clin Invest 101:578-83
Wesson, D E; Dolson, G M (1997) Endothelin-1 increases rat distal tubule acidification in vivo. Am J Physiol 273:F586-94
Wesson, D E (1997) Endogenous endothelins mediate increased distal tubule acidification induced by dietary acid in rats. J Clin Invest 99:2203-11

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