García-Chacón Luis E, Nguyen Khanh T, David Gavriel, Barrett Ellen F
Department of Physiology and Biophysics, University of Miami Miller School of Medicine, Miami, FL 33101, USA.
J Physiol. 2006 Aug 1;574(Pt 3):663-75. doi: 10.1113/jphysiol.2006.110841. Epub 2006 Apr 13.
Mitochondria sequester much of the Ca2+ that enters motor nerve terminals during repetitive stimulation at frequencies exceeding 10-20 Hz. We studied the post-stimulation extrusion of Ca2+ from mitochondria by measuring changes in matrix [Ca2+] with fluorescent indicators loaded into motor terminal mitochondria in the mouse levator auris longus muscle. Trains of action potentials at 50 Hz produced a rapid increase in mitochondrial [Ca2+] followed by a plateau, which was usually maintained after the end of the stimulus train and then slowly decayed back to baseline. Increasing the Ca2+ load delivered to the terminal by increasing the number of stimuli (from 500 to 2000) or the stimulation frequency (from 50 to 100 Hz), by increasing bath [Ca2+], or by prolonging the action potential with 3,4-diaminopyridine (100 microM) prolonged the post-stimulation decay of mitochondrial [Ca2+] without increasing the amplitude of the plateau during stimulation. Inhibiting the opening of the mitochondrial permeability transition pore with cyclosporin A (5 microM) had no significant effect on the decay of mitochondrial [Ca2+]. Inhibition of the mitochondrial Na+-Ca2+ exchanger with CGP-37157 (50 microM) dramatically prolonged the post-stimulation decay of mitochondrial [Ca2+], reduced post-stimulation residual cytosolic [Ca2+], and reduced the amplitude of endplate potentials evoked after the end of a stimulus train in the presence of both low and normal bath [Ca2+]. These findings suggest that Ca2+ extrusion from motor terminal mitochondria occurs primarily via the mitochondrial Na+-Ca2+ exchanger and helps to sustain post-tetanic transmitter release at mouse neuromuscular junctions.
在频率超过10 - 20Hz的重复刺激过程中，线粒体可隔离进入运动神经末梢的大部分Ca2+。我们通过使用装载到小鼠耳长肌运动终板线粒体中的荧光指示剂测量基质[Ca2+]的变化，研究了刺激后Ca2+从线粒体中的排出情况。50Hz的动作电位串使线粒体[Ca2+]迅速升高，随后达到平稳状态，该平稳状态通常在刺激串结束后仍持续，然后缓慢衰减回基线水平。通过增加刺激次数（从500次增加到2000次）、刺激频率（从50Hz增加到100Hz）、增加浴液[Ca2+]或用3,4 - 二氨基吡啶（100μM）延长动作电位来增加传递到终板的Ca2+负荷，可延长刺激后线粒体[Ca2+]的衰减时间，而不会增加刺激期间平稳状态的幅度。用环孢素A（5μM）抑制线粒体通透性转换孔的开放对线粒体[Ca2+]的衰减没有显著影响。用CGP - 37157（50μM）抑制线粒体Na+-Ca2+交换体可显著延长刺激后线粒体[Ca2+]的衰减时间，降低刺激后残余胞质[Ca2+]，并在低浴液[Ca2+]和正常浴液[Ca2+]条件下，降低刺激串结束后诱发的终板电位幅度。这些发现表明，运动终板线粒体中的Ca2+排出主要通过线粒体Na+-Ca2+交换体进行，并有助于维持小鼠神经肌肉接头处强直后递质的释放。
