不同前负荷条件下去神经平滑肌的应力松弛与膜电流特征

Preload-induced stress relaxation and membrane current changes in the denervated intact smooth muscle

背景:肌丝牵拉实验是检验去神经后的肌丝标本在不同受力负荷状态下的肌肉自身力学变化的实验方法。目的:探讨不同前负荷平滑肌组织在肌动图上产生的自主性舒缩波形特点以及自主性舒缩时的膜电流特点。方法:平滑肌肌丝取自昆明小鼠主动脉平滑肌层及膀胱壁肌层。肌丝标本松弛状态下在任氏液中稳定5 min后将两端固定,微量步进定位器调整标本长度,使前负荷至1 g。以此作为标本初长度(L0)。微量器快速牵拉标本1次(L0+1)即低前负荷,每间隔5min快速牵拉标本1次,第10次为高负荷(L0+10),在快速牵拉前滴加3%Ca Cl2及0.05%Nitrendipine干预,观察L0+1及L0+10后标本的自主舒缩。制备2μm玻璃微电极,显微镜下贴附平滑肌组织并高阻封接,测量L0+1及L0+10两种前负荷后应力松弛时间相的膜电位变化。结果与结论:(1)平滑肌标本应力松弛期随前负荷上升而变短,且膀胱平滑肌张力松弛期短于主动脉平滑肌,显示两种组织在顺应性上存在差异;(2)应力松弛期自主舒缩幅度随前负荷的增加而加大;(3)主动脉平滑肌膜电流波动随前负荷的增加而显著;(4)高钙环境显著提高了主动脉平滑肌膜电流的幅度及频率,这可被L-型钙通道阻断剂(0.05%Nitrendipine)抑制而减弱;(5)结果提示,前负荷的增长会引起应力松弛期肌源性自主收缩加强,这种顺应性变化与机械性牵拉引起的跨膜离子流动有关。高负荷状态下的高钙环境引起平滑肌膜电流的加强,这种变化可被L-型钙通道阻断剂所抑制。从而说明快速牵拉不仅直接作用于平滑肌机械门控通道,同时也影响到L-型钙通道的激活。

BACKGROUND：Muscle stretch test is a method for testing mechanical properties of denervated muscle under diverse loads.OBJECTIVE：To observe changes of spontaneous tension wave in myography and membrane electric current in aorta smooth muscle samples under several preload conditions.METHODS：Denervated intact smooth muscle samples were taken from the Kunming mouse aorta and urinary bladder wall.Smooth muscle samples were fixed on a micro positioning device and the first stretch was induced for a passive tension up to 1 g,this position was determined as the initial length（L0）.Intermittently,with increasing sample length from L0 every 5 minutes,the 1st and 10 th stretch were recorded as low-（L0＋1） and high-preload（L0＋10）.3% CaC l2 and 0.05% nitrendipine were dropped on the samples before L0＋1 and L0＋10 stretch were recorded.Membrane current changes were evaluated by glass microelectrodes and MultiC lamp 700 B Amplifier and pC lamp 10 analyzer software.The membrane current changes after L0＋1 and L0＋10 were analyzed.RESULTS AND CONCLUSION：Prolonged stress relaxation phase was shortened with increasing preload in smooth muscle preparations.The stress relaxation phase in smooth muscle samples from urinary bladder wall was shorter than aorta samples.It was revealed that there were compliance differences between two kinds of smooth muscle samples.In aorta samples,myogenic spontaneous contraction amplitude during stress relaxation and membrane current were significantly increased with increasing preload.Increased amplitude and frequency of membrane current in aorta smooth muscle samples under high calcium concentration（3% Ca Cl2） were inhibited by L-type calcium channel blockade（0.05% nitrendipine）.Collectively,increased preload in smooth muscle samples induced a directly decrease of compliance with an enlargement of myogenic spontaneous contraction amplitude.It appeared vital in particular in aorta samples.Membrane current significantly enhanced in spontaneous contraction period suggested the mechanical stretching ionic inward flow involved in this period.Membrane current amplitude was increased by high-preload under elevated calcium conditions;however,this change was suppressed by L-type calcium channel blockade.This study indicates quick stretching influences not only mechanically-gated channels in smooth muscle,but also the activation of L-type calcium channel.