Takimoto Eiki, Belardi Diego, Tocchetti Carlo G, Vahebi Susan, Cormaci Gianfrancesco, Ketner Elizabeth A, Moens An L, Champion Hunter C, Kass David A
Division of Cardiology, Department of Medicine, Johns Hopkins Medical Institutions, Baltimore, MD 21205, USA.
Circulation. 2007 Apr 24;115(16):2159-67. doi: 10.1161/CIRCULATIONAHA.106.643536. Epub 2007 Apr 9.
Recent cell-based studies have found that cGMP synthesis and hydrolysis by phosphodiesterase (PDE) appear compartmentalized, with nitric oxide synthase-derived and/or PDE type 5 (PDE-5)-hydrolyzable cGMP undetected at the sarcolemmal membrane in contrast to cGMP stimulated by natriuretic peptide. In the present study, we determine the functional significance of such compartments with a comparison of beta-adrenergic modulation by PDE-5 inhibition to that of natriuretic peptide stimulation in both cardiomyocytes and intact hearts. The potential role of differential cGMP and protein kinase G stimulation by these 2 modulators was also studied.
Intact C57/BL6 mouse hearts were studied with pressure-volume analysis, and adult isolated myocytes were studied with fluorescence microscopy. PDE-5 inhibition with 0.1 to 1 micromol/L sildenafil (SIL) suppressed isoproterenol (ISO)-stimulated contractility, whereas 10 micromol/L atrial natriuretic peptide (ANP) had no effect. ISO suppression by SIL was prevented in cells pretreated with a protein kinase G inhibitor. Surprisingly, myocardial cGMP changed little with SIL+ISO yet rose nearly 5-fold with ANP, whereas protein kinase G activation (vasodilator-stimulated protein phosphorylation; ELISA assay) displayed the opposite: increased with SIL+ISO but unaltered by ANP+ISO. PDE-5 and ANP compartments were functionally separated, as inhibition of nitric oxide synthase by N(w)-nitro-L-arginine methyl ester eliminated antiadrenergic effects of SIL, yet this was not restorable by co-stimulation with ANP.
Regulation of cardiac beta-adrenergic response by cGMP is specifically linked to a nitric oxide-synthesis/PDE-5-hydrolyzed pool signaling via protein kinase G. Natriuretic peptide stimulation achieves greater detectable increases in cGMP but not protein kinase G activity and does not modulate beta-adrenergic response. Such disparities likely contribute to differential cardiac regulation by drugs that modulate cGMP synthesis and hydrolysis.
最近基于细胞的研究发现，磷酸二酯酶（PDE）介导的环磷酸鸟苷（cGMP）合成与水解似乎存在区室化现象，与利钠肽刺激的cGMP相反，一氧化氮合酶衍生的和/或5型磷酸二酯酶（PDE-5）可水解的cGMP在肌膜上未被检测到。在本研究中，我们通过比较PDE-5抑制与利钠肽刺激对心肌细胞和完整心脏中β-肾上腺素能调节的作用，来确定这种区室化的功能意义。还研究了这两种调节剂对cGMP和蛋白激酶G刺激差异的潜在作用。
用压力-容积分析研究完整的C57/BL6小鼠心脏，用荧光显微镜研究成年分离的心肌细胞。用0.1至1微摩尔/升西地那非（SIL）抑制PDE-5可抑制异丙肾上腺素（ISO）刺激的收缩力，而10微摩尔/升心房利钠肽（ANP）则无作用。用蛋白激酶G抑制剂预处理的细胞可防止SIL对ISO的抑制作用。令人惊讶的是，心肌cGMP在SIL + ISO处理时变化不大，但在ANP处理时升高近5倍，而蛋白激酶G激活（血管舒张剂刺激的蛋白磷酸化；ELISA测定）则相反：在SIL + ISO处理时增加，但在ANP + ISO处理时未改变。PDE-5和ANP区室在功能上是分开的，因为N（ω）-硝基-L-精氨酸甲酯抑制一氧化氮合酶消除了SIL的抗肾上腺素能作用，但与ANP共同刺激不能恢复这种作用。
cGMP对心脏β-肾上腺素能反应的调节通过蛋白激酶G与一氧化氮合成/PDE-5水解池信号传导特异性相关。利钠肽刺激可使cGMP有更大的可检测增加，但不会增加蛋白激酶G活性，也不会调节β-肾上腺素能反应。这种差异可能导致调节cGMP合成和水解的药物对心脏有不同的调节作用。
