The nitric oxide/cGMP signalling pathway is a major regulator of penile vascular smooth muscle tone and plays a critical role in erection. Phosphodiesterases, enzymes which hydrolyze cyclic nucleotides, are an integral part of this signalling pathway. Phosphodiesterase type 5 (PDE 5) is one of the main phosphodiesterases expressed in penile corpus cavernosum smooth muscle. Sildenafil, a reversible PDE 5 selective inhibitor, has been successfully utilized in the clinical treatment of erectile dysfunction. The efficacy of this inhibitor in ameliorating erectile function in men with impotence, resulting from a broad range of etiologies, emphasizes the crucial aspect which PDE5 plays in regulating penile smooth muscle tone. Yet the regulation of PDE 5 expression or activity is not well understood and the consequences of prolonged inhibition of this enzyme are unknown. Fundamental knowledge of the cellular and molecular mechanisms which regulate PDE 5 expression and/or activity is important to the understanding of erectile function. The novel perspective that intracellular cGMP levels are actively regulated by PDE 5 broadens our understanding of the cGMP signalling pathway and the integrated mechanisms which control penile trabecular smooth muscle tone. Thus, in this study we will investigate potential regulatory mechanisms which may alter the expression and/or activity of PDE 5 in human penile corpus cavernosum smooth muscle. We will utilize primary cultures of human penile trabecular smooth muscle cells to study and characterize mechanisms of PDE 5 regulation without disrupting the intracellular regulatory pathways which are under investigation. Using cyclic nucleotide radioimmunoassays, Northern and Western blot analyses, and enzyme activity assays, we will investigate: 1) the effects of nitric oxide and cGMP on PDE 5 expression and/or activity; 2) the effects of cAMP and agonists which stimulate adenylate cyclase on PDE 5 expression and/or activity; 3) the effects of cAMP and agonists which stimulate adenylate cyclase on PDE 5 expression and/or activity; 4) cross-talk regulation between cyclic nucleotides and their respective protein kinases, and estrogen and testosterone signalling pathways. This investigation of multiple signalling systems and their influence on PDE 5 will increase our current understanding of the extensive nature by which distinct signalling pathways can interact and provide integrated regulation of smooth muscle tone. This work will provide new and useful information on regulation of PDE5 which will improve treatment strategies for management o erectile dysfunction.
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