289 UPREGULATION OF CAVEOLIN PROTEINS BY ANGIOTENSIN II ENHANCES DETRUSOR CONTRACTION

Abstract

INTRODUCTION AND OBJECTIVES: Bladder smooth muscle (BSM) cells release Angiotensin II (AngII) in response to mechanical stress and the angiotensin system is upregulated with bladder obstruction. AngII-mediated bladder contractions require intact caveolae, membrane microdomains involved in signal transduction regulation. These microdomains were found to be altered in hypertrophic detrusor muscle from obstructed bladders. This study investigated the effect of AngII on caveolin expression and whether changes in caveolins induced by AngII are associated with functional alterations. METHODS: Rat bladder tissue without mucosa was challenged for 8 hours with exogenous AngII, or with continuous electrical field stimulation (EFS) in the presence or absence of the AngII receptor-1 (AT1R) antagonist losartan (los). Changes in amplitude of contractions in response to treatments were recorded during the 8 hours. The effect of EFS on the release of endogenous AngII was determined by enzyme immunoassay. After stimulation, caveolin gene and protein expression was determined by real time rt-PCR and western blot. The interaction between AT1R and caveolin proteins was investigated by immunoprecipitation. RESULTS: Exposure of BSM tissue to AngII upregulated Cav-1, Cav-2 and Cav-3 gene and protein expression compared to non-stimulated tissue. Caveolin gene expression was similarly upregulated after 8 hours of EFS, but remained unchanged if EFS was delivered in the presence of los. Release of endogenous AngII was increased in EFS stimulated tissue compared to control. Continuous AngII exposure increased the amplitude of spontaneous activity compared to non-stimulated tissue. In addition, the amplitude of EFSinduced contractions was augmented after 8 hours. However, the increase in EFS-induced contractions was prevented by los. AT1R co-precipitated with Cav-1, Cav-2 and Cav-3. CONCLUSIONS: The caveolin protein-AT1R interaction is consistent with the localization of AT1R in caveolae and its regulation by caveolin proteins. The effect of AT1R inhibition on caveolin expression during mechanical perturbations indicates that autocrine/paracrine AngII positively regulates caveolin expression. These findings suggest that endogenous AngII released by BSM initiates a positive feedback response in which AT1R-activated detrusor contractions that are facilitated by caveolae become augmented by an enhanced caveolin-AT1R interaction. These findings may have important implications for pathophysiologic processes associated with altered AngII secretion such as the hypertrophic response to bladder obstruction.