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We raised a polyclonal antibody against a decapeptide corresponding to the carboxyl terminus of the rat angiotensin II AT1 receptor. This antibody was demonstrated to be specific for the rat receptor according to a number of approaches. These included (a) the ultrastructural localization of immunogold-labeled receptor on the surfaces of zona glomerulosa cells in the adrenal cortex, (b) the specific labeling of Chinese hamster ovarian (CHO) cells transfected with AT1 receptors, (c) the identification of a specific band on Western blots, (d) the immunocytochemical co-localization of angiotensin receptors on neurons in the lamina terminalis of the brain shown to be responsive to circulating angiotensin II, as shown by the expression of c-fos, and (e) the correlation between the expression of the mRNA of the AT1 receptor and AT1 receptor immunoreactivity.(J Histochem Cytochem 47:507-515, 1999)

Renomedullary interstitial cells (RMICs) are prominent in the inner medullary interstitium and have binding sites for several vasoactive agents, including angiotensin II (ANG II). Although the functional role of RMICs remains largely unknown, it is likely that the interaction between RMICs and vasoactive peptides is important in the regulation of renal function. The current investigation characterizes the cellular responses following treatment of RMICs with ANG II. Studies were performed on RMICs isolated from Sprague-Dawley rat kidneys. 125I-labeled [Sar1,Ile8]ANG II specifically bound to RMICs at sites determined by reverse transcription-polymerase chain reaction to be of the AT1A subtype. ANG II (10(-6) and 10(-10) M) had no effect on either basal or forskolin-stimulated adenosine 3',5'-cyclic monophosphate accumulation in RMICs but increased intracellular inositol 1,4,5-trisphosphate concentration after 10 s and intracellular calcium concentration after 18 s. For RMICs plated at low densities, ANG II (10(-6) M) induced an increase in [3H]thymidine incorporation, mediated through the AT1-receptor subtype. For RMICs plated at high densities, ANG II (10(-6) M) induced an increase in extracellular matrix synthesis as detected by trans-35S incorporation, an effect also mediated by AT1 receptors. We conclude that ANG II AT1A receptors on cultured RMICs are coupled to intracellular second messenger pathways leading to hyperplasia and synthesis of extracellular matrix.

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