Evidence for expression and function of phosphodiesterase type 5 (PDE-V) in rat resistance arteries

Evidence is provided for expression and a functional role for phosphodiesterase type V (PDE-V) in the rat isolated small mesenteric artery. The reverse transcription polymerase chain reaction (RT- PCR) demonstrated mRNA for PDE-V, while Western blotting and immunocytochemical studies showed corresponding protein expression. Smooth muscle relaxation to the nitric oxide donor, diethylamine NONOate (DEA NONOate; 1 nM-10mM; pEC50=6.7+0.3) was potentiated significantly by the specific inhibitor of PDE-V, 4-((3,4-(methylenedioxy)benzyl)amino)-6-chloroqui- nazoline (MBCQ; 1 mM; pEC50=10.5+0.04). These data show that PDE-V is expressed in both the smooth muscle and endothelial cells of a resistance artery, and the enzyme can significantly influence nitric oxide-evoked vasorelaxation. British Journal of Pharmacology (2001) 132, 13-17 Introduction Sildenafil (Viagra TM ) a selective inhibitor of phosphodiesterase type 5 (PDE-V), has been shown to be a clinically eAective treatment for erectile dysfunction (Boolell et al., 1996). Its action results from increased levels of cyclic guanosine monophosphate (cyclic GMP) which is normally degraded by PDE-V. This cyclic nucleotide is a second messenger for nitric oxide, which is involved in the regulation of numerous functions, including vascular smooth muscle tone (Corbin & Francis, 1999). Reported cardiovascular side eAects of sildenafil are typically minor and associated with vasodilatation (headache, flushing and small decreases in systolic and diastolic blood pressure). However, although the incidence is small, serious cardiovascular events, including significant hypotension can occur. Most at risk are individuals taking organic nitrates, commonly prescribed to manage the symptoms of angina pectoris. The coadministra- tion of nitrates and sildenafil significantly increases the risk of potentially life threatening hypotension. The mechanism of this interaction has not been defined. The present study sought to identify the possible expression of PDE type V in a resistance artery and to correlate expression with the eAect of a specific inhibitor of this enzyme on the relaxant responses to an exogenously applied NO donor. Methods Animals Male Sprague-Dawley rats (250-300 g) were stunned and killed by cervical dislocation. RNA isolation and RT-PCR Small mesenteric arterial segments were dissected, cleaned of fat and connective tissue and snap-frozen prior to isolation of total RNA. Sequence specific sense (5'-ttgaccgacctcttagacc-3') and antisense (5'-ctacaccaacgacctcttcc-3') oligonucleotide primers were designed against the published genetic sequences for the rat PDE-V isoform (Kotera et al., 1997). A one tube RT- PCR reaction was carried out on RNA isolated from