Habuchi Y, Nishio M, Tanaka H, Yamamoto T, Lu L L, Yoshimura M
Department of Laboratory Medicine, Kyoto Prefectural University of Medicine, Japan.
Am J Physiol. 1996 Dec;271(6 Pt 2):H2274-82. doi: 10.1152/ajpheart.1996.271.6.H2274.
Effects of acetylcholine (ACh) on L-type Ca2+ current (ICa) were examined in isolated atrioventricular (AV) node cells exhibiting spontaneous contractions and pacemaker current (If). ACh at a saturating concentration of 10 microM reduced basal ICa by 48 +/- 6%. The ACh effect was abolished by dialysis with 8-bromoadenosine 3',5'-cyclic monophosphate (8-BrcAMP), an adenosine 3',5'-cyclic monophosphate (cAMP)-dependent protein kinase inhibitor, or guanosine-5'-O-(2-thiodiphosphate). Dialysis with guanosine 3',5'-cyclic monophosphate (cGMP) or NG-monomethyl-L-arginine (L-NMMA) and application of the cGMP-dependent protein kinase inhibitor KT-5823 (1 microM) did not affect ACh inhibition of ICa. Nitric oxide donor 3-morpholinosydnonimine (100 microM) and type III phosphodiesterase (PDE) inhibitor trequinsin (10 nM) enhanced basal ICa by 10-20%, whereas type IV PDE inhibitor Ro-20-1724 (30 microM) together with trequinsin caused a large ICa stimulation comparable to that by 3-isobutyl-1-methylxanthine (IBMX). These findings indicate that ACh inhibits basal ICa primarily by suppressing cAMP synthesis and that these cells have a potent type III and IV PDE activity to determine the basal cAMP concentration. When ICa was stimulated by IBMX (100 microM), the inhibitory effect of ACh was slightly reduced by L-NMMA, cGMP, and methylene blue but not by KT-5823 or Ro-20-1724. ACh hardly inhibited, or even enhanced, IBMX-stimulated Ica when forskolin (3 microM) was coapplied or the IBMX concentration was increased to 500 microM. These findings suggest that cAMP is degraded in the presence of 100 microM IBMX to some extent. Type II PDE, for which IBMX has a relatively high inhibitor constant, seems to contribute partially to the cAMP degradation.
在表现出自发性收缩和起搏电流（If）的离体房室（AV）结细胞中，研究了乙酰胆碱（ACh）对L型钙电流（ICa）的影响。10微摩尔饱和浓度的ACh使基础ICa降低了48±6%。用8-溴腺苷3',5'-环磷酸（8-BrcAMP）（一种腺苷3',5'-环磷酸（cAMP）依赖性蛋白激酶抑制剂）或鸟苷-5'-O-（2-硫代二磷酸）进行透析可消除ACh的作用。用鸟苷3',5'-环磷酸（cGMP）或NG-单甲基-L-精氨酸（L-NMMA）进行透析以及应用cGMP依赖性蛋白激酶抑制剂KT-5823（1微摩尔）均不影响ACh对ICa的抑制作用。一氧化氮供体3-吗啉代辛二胺（100微摩尔）和III型磷酸二酯酶（PDE）抑制剂曲喹辛（10纳摩尔）使基础ICa增强了10 - 20%，而IV型PDE抑制剂罗-20-1724（30微摩尔）与曲喹辛共同作用引起的ICa大幅刺激与3-异丁基-1-甲基黄嘌呤（IBMX）相当。这些发现表明，ACh主要通过抑制cAMP合成来抑制基础ICa，并且这些细胞具有强大的III型和IV型PDE活性以决定基础cAMP浓度。当用IBMX（100微摩尔）刺激ICa时，L-NMMA、cGMP和亚甲蓝可使ACh的抑制作用略有降低，但KT-5823或罗-20-1724则无此作用。当共同应用福斯可林（3微摩尔）或将IBMX浓度增加至500微摩尔时，ACh几乎不抑制甚至增强了由IBMX刺激的Ica。这些发现提示，在100微摩尔IBMX存在的情况下，cAMP会在一定程度上降解。对于IBMX具有相对较高抑制常数的II型PDE似乎部分参与了cAMP的降解。
