IMMUNOHISTOCHEMICAL LOCALIZATION OF ANG-II AT(1) RECEPTOR IN ADULT-RAT KIDNEY USING A MONOCLONAL-ANTIBODY

Molecular and functional studies have suggested that AT(1) receptors a re present in most nephron segments, yet direct demonstration of AT(1) at these sites is lacking. The present study was performed to determi ne the intrarenal localization of the AT(1) receptor utilizing a monoc lonal anti-peptide (amino acid residues 8-17) antibody (6313/G2) in ad ult male Sprague-Dawley rats. Western blot analysis of kidney protein extracts showed a predominant 41-kDa immunoreactive band corresponding to the molecular weight of the deduced cDNA sequence. To determine op timal fixation conditions, kidney tissues were immersion fixed in Boui n's solution, 10% buffered Formalin, or 4% paraformaldehyde. Specifici ty of immunostaining was documented by preadsorption of the antibody w ith the immunogenic peptide sequence. Prominent AT(1) immunostaining w as visualized in the proximal tubule brush-border and basolateral memb ranes. In addition, distal tubules, cortical and medullary collecting ducts, and the renal arterial vasculature exhibited specific immunorea ctivity. Glomerular staining for AT(1) was observed in mesangial cells and podocytes. Macula densa cells stained positively. Similar localiz ation of the AT(1) receptor was obtained using the three tissue fixati on methods, although the intensity of vascular and glomerular staining was highest in Bouin-fixed tissues. The present study demonstrates th at the AT(1) receptor is more widely distributed along the nephron tha n previously described and includes renal vascular smooth muscle and p roximal and distal epithelial sites.