Two types of non‐selective cation channel opened by muscarinic stimulation with carbachol in bovine ciliary muscle cells
Two types of non‐selective cation channel opened by muscarinic stimulation with carbachol in bovine ciliary muscle cells
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DOI：
10.1113/jphysiol.2004.065607
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发表时间：
2004-09
期刊：
影响因子：
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通讯作者：
Y. Takai;R. Sugawara;H. Ohinata;A. Takai
中科院分区：
文献类型：
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作者：
Y. Takai;R. Sugawara;H. Ohinata;A. Takai
在睫状肌中，紧张性收缩需要持续的Ca 2+通过细胞膜流入。然而，迄今为止，人们对这种缺乏电压门控Ca 2+通道的组织中Ca 2+内流的途径知之甚少。为了鉴定作为Ca 2+进入途径的离子通道，我们通过全细胞电压钳研究了卡巴胆碱（CCh）对新鲜分离的牛睫状肌细胞的影响。使用装有含100 mm天冬氨酸铯、5 mm BAPTA和180 μm GTP的无K+溶液（pH 7.0;细胞内游离Ca 2+浓度，[Ca 2 +]i= 70 nm）的移液器进行实验。CCh诱发的内向电流在保持电位接近0 mV时显示极性反转。对电流噪声的分析区分了两种类型的非选择性阳离子通道（NSCCL和NSCCS），它们具有非常不同的单位电导（35 pS和100 fS）。用阳离子置换法测得非小细胞肺癌细胞对Li+、Na+、Cs+、Mg ~（2+）、Ca ~（2+）、Sr ~（2+）和Ba ~（2+）的渗透率分别为0.9：1.0：1.5：0.2：0.3：0.4：0.5和1.0：1.0：1.8：2.5：2.6：3.2：5.0。[Ca ~（2+）]i升高可明显抑制NSCCS，但对NSCCL无明显抑制作用。1-100 μm SKF 96365、La 3+和Gd 3+均呈剂量依赖性抑制NSCCL和NSCCS，这也抑制了CCh在肌束中产生的收缩的紧张性成分，而不显著影响初始相位成分。NSCCL和/或NSCCS可能是牛睫状肌持续收缩所需的主要Ca 2+进入途径。牛睫状肌（整个组织）中的RT-PCR实验检测了几种瞬时受体电位（TRP）通道同源物（TRPC 1，TRPC 3，TRPC 4和TRPC 6）的mRNA，这些通道现在被认为是受体操作阳离子通道的可能分子候选者。
In the ciliary muscle, the tonic contraction requires a sustained influx of Ca2+ through the cell membrane. However, little has hitherto been known about the route(s) of Ca2+ influx in this tissue that lacks voltage‐gated Ca2+ channels. To identify ion channels as the Ca2+ entry pathway we studied the effects of carbachol (CCh) on freshly isolated bovine ciliary muscle cells by whole‐cell voltage clamp. Experiments were carried out using pipettes filled with K+‐free solution containing 100 mm caesium aspartate, 5 mm BAPTA and 180 μm GTP (pH 7.0; the intracellular free Ca2+ concentration, [Ca2+]i= 70 nm). CCh evoked an inward current showing polarity reversal at a holding potential near 0 mV. Analysis of the current noise distinguished two types of non‐selective cation channel (NSCCL and NSCCS) with widely different unitary conductances (35 pS and 100 fS). The ratios of the permeabilities to Li+, Na+, Cs+, Mg2+, Ca2+, Sr2+ and Ba2+, estimated by cation replacement procedures, were 0.9: 1.0: 1.5: 0.2: 0.3: 0.4: 0.5 for NSCCL, and 1.0: 1.0: 1.8: 2.5: 2.6: 3.2: 5.0 for NSCCS. NSCCS, but not NSCCL, was strongly inhibited by elevation of [Ca2+]i. Both NSCCL and NSCCS were dose‐dependently inhibited by 1–100 μm SKF96365, La3+ and Gd3+, which also inhibited the tonic component of the contraction produced in muscle bundles by CCh without markedly affecting the initial phasic component. NSCCL and/or NSCCS may serve as a major Ca2+ entry pathway required for sustained contraction of the bovine ciliary muscle. RT‐PCR experiments in the bovine ciliary muscle (whole tissue) detected mRNAs of several transient receptor potential (TRP) channel homologues (TRPC1, TRPC3, TRPC4 and TRPC6), which are now regarded as possible molecular candidates for receptor‐operated cation channels.