Hill R R, Robbins N, Fang Z P
Center for Neurosciences, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106.
J Neurocytol. 1991 Mar;20(3):165-82. doi: 10.1007/BF01186990.
In order to assay the extent of ongoing synaptic remodelling in adult mouse neuromuscular junctions, dynamic structural changes of identified neuromuscular junctions were monitored in vivo over periods up to three months. Nerve terminal outgrowths as small as 1 micron were detectable with a new fluorescent tetanus toxin C-fragment stain combined with fluoresceinated alpha-bungarotoxin to stain postsynaptic acetylcholine receptors. With limited illumination, the new stain did not affect miniature endplate potential frequency, nor morphometric parameters of repeatedly observed neuromuscular junctions. At each observation, areas of presynaptic nerve terminal extending beyond underlying acetylcholine receptor ('preprojections'), and areas of acetylcholine receptor without overlying nerve terminal ('postprojections') were measured. Regions of the neuromuscular junction in which nerve terminal-postsynaptic acetylcholine receptor complexes either 'lengthened' or 'shortened' between observations were also measured. The total area of pre- and postprojections (relative to total junctional area) remained the same over three months but most had been replaced; only 20% of preprojections gave rise to lengthenings, the rest retracted or were unchanged. Lengthening and shortening of branches were about 1-2% of junctional area per month. These more permanent changes occurred against a background of ongoing transient nerve terminal outgrowth and retraction (which constituted 80% of all neuromuscular junction shape changes from one observation to the next, compared with 20% for the postsynaptic component). Breaks in the continuity of the underlying acetylcholine receptor were also observed between observations as were instances where acetylcholine receptor continuity was re-established. A newly observed form of plasticity was a shift in position and angle of pre-existing branches. Establishment of new acetylcholine receptor-positive synaptic regions was mostly preceded by nerve terminal outgrowth on the previous observation. In animals in which spontaneous wheel-running increased locomotor activity approximately tenfold over a period of 35 days, the findings were identical to those in unexercised mice. In summary, in the adult neuromuscular junction, the nerve terminal, not the postsynaptic component, is the dynamic entity, continually changing shape on the scale of micrometers, with relatively small permanent changes. These ongoing exploratory excursions may supply the substrate for synaptic plasticity, which would involve regulation of the dynamics or stability of nerve outgrowth.
为了测定成年小鼠神经肌肉接头处正在进行的突触重塑程度，在长达三个月的时间里，对体内已确定的神经肌肉接头的动态结构变化进行了监测。结合荧光α-银环蛇毒素对突触后乙酰胆碱受体进行染色，使用一种新的荧光破伤风毒素C片段染色法，可检测到小至1微米的神经末梢生长。在有限光照下，这种新染色法不影响微小终板电位频率，也不影响反复观察的神经肌肉接头的形态学参数。每次观察时，测量突触前神经末梢延伸至其下方乙酰胆碱受体之外的区域（“前突出”），以及没有上方神经末梢覆盖的乙酰胆碱受体区域（“后突出”）。还测量了在观察期间神经末梢-突触后乙酰胆碱受体复合物“延长”或“缩短”的神经肌肉接头区域。前突出和后突出的总面积（相对于总接头面积）在三个月内保持不变，但大部分已经被替换；只有20%的前突出导致延长，其余的缩回或保持不变。分支的延长和缩短每月约为接头面积的1-2%。这些更持久的变化发生在持续的短暂神经末梢生长和缩回的背景下（从一次观察到下一次观察，这构成了所有神经肌肉接头形状变化的80%，而突触后成分的变化为20%）。在观察期间还观察到了其下方乙酰胆碱受体连续性的中断，以及乙酰胆碱受体连续性重新建立的情况。一种新观察到的可塑性形式是先前存在的分支的位置和角度发生了变化。新的乙酰胆碱受体阳性突触区域的建立大多在先前观察到神经末梢生长之后。在自发轮转运动使运动活动在35天内增加约十倍的动物中，研究结果与未运动小鼠的结果相同。总之，在成年神经肌肉接头中，神经末梢而非突触后成分是动态实体，其形状在微米尺度上不断变化，永久性变化相对较小。这些持续的探索性活动可能为突触可塑性提供基础，而突触可塑性可能涉及对神经生长动力学或稳定性的调节。
