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Regulation of cell-volume and intracellular osmolarity by chloride channels in cardiac cells

心肌细胞中氯离子通道对细胞体积和细胞内渗透压的调节

基本信息

批准号：
14570041
负责人：
RHARA Tsuguhisa
金额：
$ 2.24万
依托单位：
Saga University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2003
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14570041/
关键词：
swelling-activated RVS current RVD CFTR PKA ventricular cell ATP PKC P2 purinoceptor 容積感受性Cl電流細胞容積調節性容量減少 DIDS

项目摘要

1.The cell-volume regulation by chloride currents was studied in guinea-pig cardiac myocytes, using a microscopic video-image analysis. Application ofhypotonic solution to the cells at normal [K]_0 induced a cell swelling, and the so-swollen cells showed a slight but clear spontaneous cell shrinkage, indicating operation of the mechanism of regulatory volume decrease (RVD)._This RVD could be pronounced at low [Cl]_o, at which E_<Cl> was far more positive than E_m. On the contrary, when the hypotonic solution was applied to the cells at high [K]_o, at which E_<Cl> was negative to E_m, the cells swelled monotonically without showing RVD, the swelling being much greater than that seen at normal [K]_0. Both the RVD at normal [K]_0 and the 'extra' cell inflation at high [K]_0 were suppressed by the inhibitors of I_<Cl, swell>. These findings suggest that the cell inflation activated. I_<Cl, swell>, and that the activation of I_<Cl, swell> induced the RVD or the extra cell inflation, dependi … More ng on the direction and magnitude of the driving force of Cl ions across the membrane.2.The effects of extracellular ATP on [β-adrenergic activation of CFTR Cl current (I_<Cl, PKA>) were examined with the whole-cell patch clamp method. The cells were initially exposed to isoproterenol (ISO) for 〜3 min to activate I_<Cl, PKA>, and then to 1-100 μM ATP in the presence of ISO. ATP was found to potentiate I_<Cl, PKA>, in most cells examined. With 50 μM ATP, the potentiation, on average, resulted in a 1.3 fold increase of the Cl^-conductance activated by ISO alone (0.02-1μM). The effects of ADP and ATPγS on I_<Cl, PKA> were similar to those of ATP, while AMP and adenosine never potentiated I_<Cl, PKA>. Thus the potentiation was attributed, to a stimulation of P2-purinoceptors. PDBu (0.5μM), an activator of PKC, facilitated I_<Cl, PKA>, and in the presence of PDBu ATP did not further potentiate I_<Cl, PKA>. When BIM (0.2μM), an inhibitor of PKC, was present, ATP did not facilitate I_<Cl, PKA>. These findings suggested involvement of PKC in the observed ATP action. When ATP was removed in the presence of ISO, the potentiated I_<Cl, PKA> decreased (recovered) only slowly, and, if ATP was reapplied during this slowly recovering phase, the subsequent current potentiation was weak. Thus the stimulation of P2 purinoceptors by ATP facilitates the β-adrenergic activation of I_<Cl, PKA> through PKC activation, and this potential appears to persist for several min after removal of ATP. Less

1.用显微视频图像分析技术研究了豚鼠心肌细胞氯电流对细胞体积的调节作用。在正常[K]_0下，低渗溶液可引起细胞肿胀，肿胀细胞出现轻微但明显的自发性细胞皱缩，表明调节性容积减少（RVD）机制的作用。这种RVD在低[Cl]_o时明显，此时E_<Cl>m远大于E_m。相反，当低渗溶液作用于高[K]_0（<Cl>E_0与E_m呈负相关）时，细胞呈单调性肿胀，但无RVD，肿胀程度远大于正常[K]_0时的肿胀程度。I_<Cl，swell>抑制剂可抑制正常[K]_0时的RVD和高[K]_0时的“额外”细胞膨胀。这些发现表明细胞膨胀被激活。I_<Cl，swell>的激活可引起RVD或细胞外膨胀，并依赖于I_<Cl，swell>的激活。 ...更多信息 2.用全细胞膜片钳技术观察了细胞外ATP对CFTR β-肾上腺素能激活Cl电流（I_<Cl，PKA>）的影响。细胞先暴露于异丙肾上腺素（ISO）0.3min以激活I_（Cl，PKA），然后在ISO存在下暴露于1-100 μM ATP。在大多数细胞中，ATP可增强I_（Cl，PKA）。在50 μM ATP的作用下，平均而言，ISO单独（0.02-1μM）激活的Cl^-电导增加了1.3倍。ADP和ATPγS对I <Cl，PKA>的作用与ATP相似，而AMP和腺苷对I <Cl，PKA>无增强作用。因此，增强归因于P2-嘌呤受体的刺激。PKC激活剂PDBu（0.5μM）使I_（Cl，PKA）增加，而在PDBu存在时，ATP不能进一步增强I_（Cl，PKA）。当有PKC抑制剂BIM（0.2μM）存在时，ATP对I_（Cl，PKA）无促进作用。这些发现表明PKC参与观察到的ATP作用。在ISO存在下去除ATP时，增强的I_（Cl，PKA）仅缓慢下降（恢复），在此缓慢恢复阶段再加ATP，则随后的电流增强作用很弱。因此，ATP刺激P2嘌呤受体可通过激活PKC而促进β-肾上腺素能I_<Cl，PKA>的激活，且这种电位在ATP去除后可持续数分钟。少