SODIUM-CHANNEL INACTIVATION IS IMPAIRED IN EQUINE HYPERKALEMIC PERIODIC PARALYSIS
SODIUM-CHANNEL INACTIVATION IS IMPAIRED IN EQUINE HYPERKALEMIC PERIODIC PARALYSIS
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DOI：
10.1152/jn.1995.73.5.1892
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发表时间：
1995-05-01
影响因子：
2.5
通讯作者：
BROWN, RH
中科院分区：
文献类型：
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作者：
CANNON, SC;HAYWARD, LJ;BROWN, RH
1. 马高钾血症性周期性麻痹（E-HPP）是一种主要的遗传性肌肉疾病，可引起与血清K+升高相关的反复发作的僵硬（肌强直）和虚弱。受影响的马携带Na通道α亚基骨骼肌同种异构体的突变等位基因。为了了解这种突变如何导致疾病表型，通过记录正常和患病马肌管上细胞附着斑块的单一电流，从生理学上定义了Na通道行为的功能缺陷。通过聚合酶链反应（PCR）扩增的基因组DNA的限制性消化，证实了该突变的存在。来自受影响马的肌管为杂合的点突变，该突变编码IV结构域S-3中的苯丙氨酸到亮氨酸的替换。该试验提供了一种快速筛选高危马突变的技术。突变体钠通道的主要生理缺陷是失活损伤。这种缺陷表现为持续活动的爆发，在此期间通道关闭和重新开放，整个过程保持去极化。中断的失活减缓了总体平均电流的衰减，并使40毫秒脉冲结束时测量的稳态打开概率增加了8倍。这个点突变确定了α亚基的一个新区域，该区域对通道的快速失活很重要。持续的钠电流是由一种不同的门控模式产生的。突变通道失活的失败并不常见，并且发生在连续的组试验中。此外，突变Na通道的开放时间分布包含一个第二慢分量（tau(s) = 1-2 ms）和一个快速分量（tau(f) = 0.4 ms），其本身足以代表正常通道的分布。这些观察结果与通道在两种失活模式之间缓慢切换的概念是一致的：快速失活与非失活。虽然变异很大，但当细胞外K+增加时，突变通道有更频繁的非失活行为爆发的趋势。对于E-HPP肌管中的Na通道，10 mM [K+]与0 mM [K+]相比，稳态与峰值打开概率之比增加了三倍（从0.008增加到0.024）。相反，在正常的Na通道中，峰值P- P-开度的稳态为0.003，且随[K+](o)不变。
1. Equine hyperkalemic periodic paralysis (E-HPP) is a dominantly inherited disorder of muscle that causes recurrent episodes of stiffness (myotonia) and weakness in association with elevated serum K+. Affected horses carry a mutant allele of the skeletal muscle isoform of the Na channel alpha-subunit. To understand how this mutation may cause the disease phenotype, the functional defect in Na channel behavior was defined physiologically by recording unitary currents from cell-attached patches on normal and affected equine myotubes.2. The presence of the mutation was confirmed in our cell line by restriction digest of polymerase chain reaction (PCR)-amplified genomic DNA. Myotubes from the affected horse were heterozygous for the point mutation that codes for a Phe to Leu substitution in S-3 of domain IV. This assay provides a rapid technique to screen for the mutation in horses at risk.3. The primary physiological defect in mutant Na channels was an impairment of inactivation. This defect was manifest as bursts of persistent activity during which the channel closed and reopened throughout a maintained depolarization. Disrupted inactivation slowed the decay of the ensemble-averaged current and produced an eightfold increase in the steady-state open probability measured at the end of a 40-ms pulse. This point mutation identifies a new region of the alpha subunit that is important for rapid inactivation of the channel.4. The persistent Na current was produced by a distinct mode of gating. Failure of a mutant channel to inactivate was infrequent and occurred in groups of consecutive trials. Furthermore, the open time distributions for mutant Na channels contained a second, slow component (tau(s) = 1-2 ms) in addition to a fast component (tau(f) = 0.4 ms), which by itself was sufficient to represent the distribution in normal channels. These observations are consistent with the notion that channels slowly switch between two modes of inactivation: rapid versus noninactivating. Although the variance was high, for mutant channels there was a trend toward more frequent bursts of noninactivating behavior when extracellular K+ was increased. For Na channels in E-HPP myotubes, the ratio of steady-state to peak open probability increased threefold (0.024 from 0.008) in 10 versus 0 mM [K+](o). Conversely, in normal Na channels the steady-state to peak P P-open was 0.003 and invariant with [K+](o).