The goal of this project is to identify differences in the mechanisms that regulate Na+ reabsorption in cortical collecting ducts (CCD) and medullary thick ascending limbs (MTAL) from Sprague-Dawley (SD), Dahl salt-sensitive (Rapp strain, S/Jr), and Dahl salt-resistant (R/Jr) rats. Such differences may underlie the varying natriuretic response to salt loading among these strains. CCDs and MTALs will be dissected from the kidneys of 5-6 week-old rats of all three strains, before hypertension has developed in the S/Jr rats. Osmotic water permeability, unidirectional fluxes of 22Na+, and transepithelial voltage will be measured. Intracellular electrophysiologic techniques will also be used to monitor changes in the amiloride-sensitive Na+ conductance of the luminal membrane of the CCD. Recognizing that mineralocorticoids and vasopressin act synergistically to increase Na+ reabsorption in the rat CCD, we will determine the role of such synergism at basal and physiologically elevated aldosterone concentrations, and over a 1-200 pM range of AVP concentrations. It is possible that differences in salt reabsorption among the strains may be manifested as a shifted dose response to AVP or aldosterone. We will also look for differences among the strains by comparing the increase in principal cell basolateral membrane surface density in CCDs from rats with elevated aldosterone concentrations. In other experiments, we will determine whether the expression of an electrically-silent, thiazide-sensitive NaC1 cotransporter in the CCD may be altered by increased or decreased NaC1 delivery to the distal nephron. We will conduct similar experiments on salt-loaded rats treated with L-arginine, an agent which has been shown to prevent the development of hypertension in S/Jr rats. We will also examine whether differences in epinephrine sensitivity among the three strains may account for differences in their response to an increased salt load. Na+ reabsorption in isolated perfused MTALs will also be examined to determine if it is higher in the S/Jr strain than in the R/Jr strain as suggested by in vivo loop perfusion, and to establish the reason for this difference. In other experiments, we will use reverse transcription of mRNA extracted from individual nephron segments followed by the polymerase chain reaction (RT-PCR) to determine whether Na,K- ATPase subunits other than alpha1 and beta1 may be expressed in specific nephron segments. We hypothesize that more ouabain-sensitive subunits in the CCD may be the target for the ouabain-like natriuretic factor. If such expression is found at the mRNA level, it will also be confirmed at the protein level by Western blot analysis, and techniques of competitive PCR will be used to quantify relative message levels of the subunits. We will also use this technology to examine which alpha2 adrenergic receptors are expressed in individual nephron segments. In the case of both the Na,K-ATPase subunit isoforms and alpha adrenergic receptors, we will be particularly interested to see if there are differences in expression among the strains of rats, or after Na+ depletion or Na+-loading, or with varying aldosterone replacement in adrenalectomized rats.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK045768-03
Application #
2145007
Study Section
General Medicine B Study Section (GMB)
Project Start
1993-08-01
Project End
1997-07-31
Budget Start
1995-08-01
Budget End
1996-07-31
Support Year
3
Fiscal Year
1995
Total Cost
Indirect Cost
Name
University of Alabama Birmingham
Department
Physiology
Type
Schools of Dentistry
DUNS #
004514360
City
Birmingham
State
AL
Country
United States
Zip Code
35294
Schafer, J A; Li, L; Sun, D (2000) The collecting duct, dopamine and vasopressin-dependent hypertension. Acta Physiol Scand 168:239-44
Schafer, J A (2000) Interaction of modeling and experimental approaches to understanding renal salt and water balance. Ann Biomed Eng 28:1002-9
Li, L; Schafer, J A (1998) Dopamine inhibits vasopressin-dependent cAMP production in the rat cortical collecting duct. Am J Physiol 275:F62-7
Sun, D; Wilborn, T W; Schafer, J A (1998) Dopamine D4 receptor isoform mRNA and protein are expressed in the rat cortical collecting duct. Am J Physiol 275:F742-51
Schafer, J A; Chen, L (1998) Low Na+ diet inhibits Na+ and water transport response to vasopressin in rat cortical collecting duct. Kidney Int 54:180-7
Hirsch, J R; Cermak, R; Forssmann, W G et al. (1997) Effects of sodium nitroprusside in the rat cortical collecting duct are independent of the NO pathway. Kidney Int 51:473-6
Schafer, J A; Watkins, M L; Li, L et al. (1997) A simplified method for isolation of large numbers of defined nephron segments. Am J Physiol 273:F650-7
Sun, D; Schafer, J A (1996) Dopamine inhibits AVP-dependent Na+ transport and water permeability in rat CCD via a D4-like receptor. Am J Physiol 271:F391-400
Hawk, C T; Li, L; Schafer, J A (1996) AVP and aldosterone at physiological concentrations have synergistic effects on Na+ transport in rat CCD. Kidney Int Suppl 57:S35-41
Schlatter, E; Cermak, R; Forssmann, W G et al. (1996) cGMP-activating peptides do not regulate electrogenic electrolyte transport in principal cells of rat CCD. Am J Physiol 271:F1158-65