Lugnier C, Gauthier C, Le Bec A, Soustre H
Laboratoire de Pharmacologie Cellulaire et Moléculaire, Centre National de la Recherche Scientifique Unité 600, Université Louis Pasteur de Strasbourg, Illkirch, France.
Am J Physiol. 1992 Mar;262(3 Pt 2):H654-60. doi: 10.1152/ajpheart.1992.262.3.H654.
The cyclic nucleotide phosphodiesterase (PDE) forms present in frog atrial fibers were isolated and characterized by their drug sensitivities. DEAE-sephacel chromatography of cytosolic PDE activity resolved three major PDE forms: peak A hydrolyzed both adenosine 3',5'-cyclic monophosphate (cAMP) and guanosine 3',5'-cyclic monophosphate (cGMP) and was activated by calcium-calmodulin (PDE I); peak B also hydrolyzed both cAMP and cGMP but was activated by 5 microM cGMP (PDE II); peak C specifically hydrolyzed cAMP (PDE IV). Rolipram specifically inhibited PDE IV (Ki = 1.1 microM), whereas dipyridamole potently inhibited both PDE II (Ki = 4.6 microM) and PDE IV (Ki = 0.8 microM). Atrial fiber PDE I was preferentially inhibited by zaprinast (Ki = 10 microM). 3-Isobutyl-1-methyl xanthine (IBMX) and theophylline inhibited nonspecifically all three different enzymes. The positive inotropic drug CI 930 only inhibited the different isolated atrial PDE forms at concentrations greater than 200 microM. However, under assay conditions for which PDE IV was specifically inhibited (presence of 100 microM rolipram), an IC50 of 17 microM for CI 930 was observed on the remaining 26% cAMP hydrolytic activity of peak C (which could represent a cGMP-inhibited PDE form: PDE III). The same PDE forms were also found in frog ventricle. The major difference between frog atrial fiber (and ventricular tissue) PDEs and mammalian cardiac PDEs is that the main cytosolic cAMP-specific hydrolytic activity in frog heart is due to PDE IV rather than PDE III. Rolipram, dipyridamole, and zaprinast might be useful tools to investigate the participation of cAMP in frog atrial contraction (unpublished observations).
通过药物敏感性对青蛙心房纤维中存在的环核苷酸磷酸二酯酶（PDE）形式进行了分离和表征。对胞质PDE活性进行DEAE-琼脂糖凝胶柱层析，分离出三种主要的PDE形式：峰A可水解腺苷3',5'-环磷酸（cAMP）和鸟苷3',5'-环磷酸（cGMP），并被钙调蛋白激活（PDE I）；峰B也可水解cAMP和cGMP，但被5μM cGMP激活（PDE II）；峰C特异性水解cAMP（PDE IV）。咯利普兰特异性抑制PDE IV（Ki = 1.1μM），而双嘧达莫则强烈抑制PDE II（Ki = 4.6μM）和PDE IV（Ki = 0.8μM）。扎普司特优先抑制心房纤维PDE I（Ki = 10μM）。3-异丁基-1-甲基黄嘌呤（IBMX）和茶碱非特异性地抑制所有三种不同的酶。正性肌力药物CI 930仅在浓度大于200μM时抑制不同的分离心房PDE形式。然而，在PDE IV被特异性抑制的测定条件下（存在100μM咯利普兰），观察到CI 930对峰C剩余26%的cAMP水解活性的IC50为17μM（这可能代表一种cGMP抑制的PDE形式：PDE III）。在青蛙心室中也发现了相同的PDE形式。青蛙心房纤维（和心室组织）PDE与哺乳动物心脏PDE的主要区别在于，青蛙心脏中主要的胞质cAMP特异性水解活性是由PDE IV而非PDE III引起的。咯利普兰、双嘧达莫和扎普司特可能是研究cAMP参与青蛙心房收缩的有用工具（未发表的观察结果）。
