(Verbatim from the application): Dopamine and D1-like receptors in the proximal tubules and other regions of the nephron play an important role in regulating renal sodium excretion. A defective DIA receptor (a D1-like receptor) function is reported in the proximal tubules of hepertensive animals and humans. In our preliminary studies we found that D1-Iike receptors are reduced by 50 percent and dopamine was unable to stimulate second messengers (cAMP and IP3) and thereby failed to inhibit Na,K-ATPase and Na,H-exchanger in the proximal tubules of obese Zucker rats (12 week old). Therefore, it is hypothesized that the diminished inhibitory effect of dopamine on sodium transporting proteins in the proximal tubules is due to a dysfunction in DIA receptor which results from altered phosphorylation of membranal DIA receptor as well as impaired recruitment of newer D1A receptors from cytosol. Such a defect in dopamine receptor function contributes to the diminished natriuretic response to exogenously administered as well as endogenously produced dopamine. This hypothesis will be tested by conducting a series of experiments to identify the biochemical/molecular mechanism(s) contributing towards the defect in DIA receptors. These will include: measurement of D1A receptor agonist affinity, determination of the effect of GTPyS on ligand binding, determination of agonist-dependent and -independent phosphorylation of D1A receptor, determination of agonist recmitment of D1A receptors on the membrane and measurement of D1A receptor mRNA and DIA receptor protein content in proximal tubules of lean and obese rats. The functional consequence of the defective DIA receptor function will be examined by studying the natriuretic and diuretic effect of dopamine and SKF38393 as well as by determining the effect of acute volume expansion on renal production of dopamine and sodium and water excretion in anesthetized lean and obese Zucker rats. In order to delineate the role of hyperinsulinemia in causing the defective D1A receptor function and the natriuretic response, experiments described above will also be performed in lean and obese rats (7-8 weeks) treated for four weeks with darglitazone, a drug which decreases insulin levels and improve insulin sensitivity as well as in 3-4 week old non-treated hyperinsulinemic and control rats. Identification of the mechanism(s) leading to dopamine receptor dysfunction and the diminished natriuretic response to dopamine would allow us to better understand the relationship between kidney D1A receptor function and hyperinsulinemia in obese rats.
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