培养的血管平滑肌细胞中钠和钾通量的容量依赖性调节：对培养基渗透压的依赖性及信号系统的调节

Volume-dependent regulation of sodium and potassium fluxes in cultured vascular smooth muscle cells: dependence on medium osmolality and regulation by signalling systems.

作者信息

Orlov S N, Resink T J, Bernhardt J, Buhler F R

机构信息

Department of Research, University Hospitals, Basel, Switzerland.

出版信息

J Membr Biol. 1992 Aug;129(2):199-210. doi: 10.1007/BF00219515.

DOI:10.1007/BF00219515

PMID:1331467

Abstract

To identify ion transport systems involved in the maintenance of vascular smooth muscle cell volume the effects of incubation medium osmolality and ion transport inhibitors on the volume and 86Rb and 22Na transport in cultured smooth muscle cells from rat aorta (VSMC) have been studied. A decrease of medium osmolality from 605 to 180 mosm increased intracellular water volume from 0.6 to 1.3 microliters per 10(6) cells. Under isosmotic conditions, cell volume was decreased by ouabain (by 10%, P less than 0.005) but was not influenced by bumetanide, furosemide, EIPA and quinidine. These latter compounds were also ineffective in cell volume regulation under hypotonic buffer conditions. Under hyperosmotic conditions, cell volume was decreased by bumetanide (by approximately 7%, P less than 0.05) and by ethylisopropyl amiloride (by approximately 13%, P less than 0.005). Ouabain-sensitive 86Rb influx was decreased by 30-40% under hypoosmotic conditions. An increase in medium osmolality from 275 to 410 mosm resulted in an approximately eightfold increase in bumetanide-inhibited 86Rb influx and 86Rb efflux. The (ouabain and bumetanide)-insensitive component of 86Rb influx was not dependent on the osmolality of the incubation medium. However (ouabain and bumetanide)-insensitive 86Rb efflux was increased by approximately 1.5-2 fold in VSMC incubated in hypotonic medium. Ethylisopropyl amiloride-inhibited 22Na influx was increased by approximately sixfold following osmotic-shrinkage of VSMC. The data show that both Na+/H+ exchange and Na+/K+/2Cl- cotransport may play a major role in the regulatory volume increase in VSMC. Basal and shrinkage-induced activities of Na+/K+/2Cl- cotransport in VSMC were similarly sensitive to inhibition by either staurosporin, forskolin, R24571 or 2-nitro-4-carboxyphenyl N,N-diphenylcarbomate (NCDC). In contrast basal and shrinkage-induced Na+/K+/2Cl- cotransport were differentially inhibited by NaF (by 30 and 65%, respectively), suggesting an involvement of guanine nucleotide binding proteins in the volume-sensitive activity of this carrier. Neither staurosporin, forskolin, R24571 nor NCDC influenced shrinkage-induced Na+/H+ exchange activity. NaF increased Na+/H+ exchanger activity under both isosmotic and hyperosmotic conditions. These data demonstrate that different intracellular signalling mechanisms are involved in the volume-dependent activation of the Na+/K+/2Cl- cotransporter and the Na+/H+ exchanger.

摘要

为了确定参与维持血管平滑肌细胞体积的离子转运系统，研究了孵育培养基渗透压和离子转运抑制剂对大鼠主动脉培养平滑肌细胞（VSMC）体积以及⁸⁶Rb和²²Na转运的影响。培养基渗透压从605降至180 mosm时，每10⁶个细胞的细胞内水体积从0.6微升增加到1.3微升。在等渗条件下，哇巴因使细胞体积减小（减小10%，P<0.005），但布美他尼、呋塞米、EIPA和奎尼丁对其无影响。在低渗缓冲液条件下，后几种化合物对细胞体积调节也无效。在高渗条件下，布美他尼（约7%，P<0.05）和乙基异丙基氨氯吡咪（约13%，P<0.005）使细胞体积减小。在低渗条件下，哇巴因敏感的⁸⁶Rb内流减少30 - 40%。培养基渗透压从275 mosm增加到410 mosm导致布美他尼抑制的⁸⁶Rb内流和⁸⁶Rb外流增加约8倍。⁸⁶Rb内流的（哇巴因和布美他尼）不敏感成分不依赖于孵育培养基的渗透压。然而，在低渗培养基中孵育的VSMC中，（哇巴因和布美他尼）不敏感的⁸⁶Rb外流增加约1.5 - 2倍。VSMC渗透收缩后，乙基异丙基氨氯吡咪抑制的²²Na内流增加约6倍。数据表明，Na⁺/H⁺交换和Na⁺/K⁺/2Cl⁻共转运在VSMC调节性体积增加中可能起主要作用。VSMC中Na⁺/K⁺/2Cl⁻共转运的基础活性和收缩诱导活性对星形孢菌素、福斯可林、R24571或2 - 硝基 - 4 - 羧基苯基N,N - 二苯基氨基甲酸酯（NCDC）的抑制同样敏感。相比之下，基础和收缩诱导的Na⁺/K⁺/2Cl⁻共转运受到NaF的不同抑制（分别为30%和65%），表明鸟嘌呤核苷酸结合蛋白参与了该载体的体积敏感活性。星形孢菌素、福斯可林、R24571或NCDC均不影响收缩诱导的Na⁺/H⁺交换活性。在等渗和高渗条件下，NaF均增加Na⁺/H⁺交换活性。这些数据表明，不同的细胞内信号机制参与了Na⁺/K⁺/2Cl⁻共转运体和Na⁺/H⁺交换体的体积依赖性激活。