Abstract 16569: Melatonin Inhibits NO-Induced Activation of BKCa Channels and Relaxation of Coronary Arteries by Increasing PKG1-Dependent Phosphorylation of PDE5
Alterations in melatonin (MEL) blood levels are associated with several cardiovascular disorders, including hypertension, ischemic heart disease, and heart failure. We recently demonstrated that MEL, acting via MT2-receptors, impairs nitric oxide (NO)-induced relaxation in coronary arteries by inhibiting NO-induced increases in intracellular cGMP levels (JPET 336: 127, 2011). In vascular smooth muscle, PKG1 limits the physiological actions of NO/cGMP signaling by phosphorylating PDE5, thereby increasing PDE5 activity.
Hypothesis: We hypothesized that MEL impairs NO-induced relaxation of coronary arteries by stimulating PKG1-dependent phosphorylation of PDE5, resulting in attenuation of NO-activated BKCa function in coronary smooth muscle cells.
Methods and Results: MEL (10-9 - 10-7 M) inhibited relaxation to the NO-donors (10-10 - 10-4 M), diethylamine-NONOate and sodium nitroprusside (SNP) in isolated porcine coronary arteries (p<0.05; n=5-8), but had no effect on relaxation induced by 8-Br-cGMP, a cell permeable, non-hydrolyzable cGMP analog. The effect of MEL on NO-induced relaxation was abolished by the PDE5 inhibitors, sildenafil (10-7M) and zaprinast (10-5 M). MEL markedly reduced NO-induced increases in cGMP levels in coronary arteries, an effect that was abolished by inhibition of PDE5 (p<0.05). Immunoblot analysis demonstrated that incubation of the tissues with MEL (10-7 M) caused a significant increase in PDE5 phosphorylation, which was attenuated (p<0.05) in arteries that were treated with the PKG1-inhibitors Rp-8-Br-PET-cGMPS (3x10-5 M) or DT-2 (10-5 M). In whole cell patch clamp studies (10mv steps from -70mv to +60mv, 200ms duration), the SNP (10-5 M)-induced increase in BKCa currents was abolished by ODQ (10-5 M). MEL (10-7M) inhibited the SNP-induced increase in BKCa currents, but had no effect on BKCa currents elicited by 8-Br-cGMP (10-3M). The effect of MEL on the SNP-induced increase in BKCa currents was abolished by either the MT2-receptor antagonist, 4P-PDOT (10-7 M) or by inhibition of PDE5.
Conclusions: MEL increases PKG1-dependent phosphorylation of PDE5, resulting in increased activity of the enzyme, inhibition of NO/cGMP-dependent activation of BKCa channels, and impaired NO-induced vasorelaxation.
- © 2012 by American Heart Association, Inc.