Clinical states in which angiotensin II (AII) is increased are associated with renal Na and water retention. AII has direct physiologic effects on renal epithelial cells. In microperfused renal proximal tubules, picomolar concentrations of AII added to either luminal or vascular perfusing solutions enhance Na reabsorption. AII receptors have been identified on both apical and basolateral membranes prepared from proximal tubules of intact rats. However, signalling and function of AII receptors on proximal tubule cells are not known. Part of the explanation for this lack of information is that until recently, cell culture methods for growing epithelial cells with differentiated apical and basolateral membranes have not been available. Our hypothesis is that apical and basolateral AII receptor expression, itinerary, signalling and function in renal epithelial cells differ and are determined by the polarized nature of these cells. As a consequence of these differences, AII binding to basolateral sites results in transduction of basolateral cell signals which enhance apical Na transport. Apical binding does not result in Na transport unless AII or AII receptors are endocytosed to basolateral signalling sites.
The specific aims are to determine differences between apical and basolateral AII receptors with regard to 1) the number and binding affinity, 2) receptor and ligand itinerary and the role of endosomes and cytoskeleton components in the process; 3) signal transduction (adenylate cyclase and phospholipase c); 4) Na 22 transport; 5) factors which control binding, signalling and 22 Na transport; 6) the relationship between the number of receptors on each membrane and a) signal transduction and b) 22 Na transport. The methods to be utilized include: 1) apical and basolateral membrane preparations from intact rats; 2) monolayers of epithelial cells from rat proximal tubules grown on porous filters. With this method, basolateral membranes become differentiated and selective access to either apical or basolateral surfaces can be gained. A variety of morphologic, biochemical and physiological techniques will be applied. Significance: These studies will provide insight on the mechanisms by which receptor expression and function are controlled in polarized cells. This information may be relevant to understanding the role of AII in hypertension and the major edematous disorders.
Showing the most recent 10 out of 11 publications
