Samuels P L, Riml H, Cohen M W
Department of Physiology, McGill University, Montreal, Quebec, Canada.
Dev Biol. 1990 Oct;141(2):399-411. doi: 10.1016/0012-1606(90)90394-x.
The formation and survival of nerve-induced clusters of acetylcholine receptors (AChRs) was monitored over a synaptogenic period of several days in cultures of myotomal muscle cells and spinal cord neurons derived from embryos of Xenopus laevis. AChRs were labeled with fluorescent alpha-bungarotoxin so that neurite-associated receptor patches (NARPs) could be viewed at daily intervals throughout the neuritic arbor of selected neurons. To avoid bleaching the NARPs and damaging the neurons, the intensity of the fluorescence excitation was reduced to 3%. Images were digitized and NARPs were measured with a computer-based image analysis system. Virtually all newly formed NARPs (greater than 90%) were detected at the same time as neurite-muscle contact and in the same proximal-distal sequence as neuritic growth. Those which formed in 6- to 13-day-old cocultures had similar distributions with respect to length, area, intensity, and area X intensity to those which formed in 1- to 2-day-old cocultures. NARPs exhibited variable daily changes in these parameters but on average they grew and reached close to their ultimate values within 1-2 days. Almost all (greater than 95%) survived as long as their contacts. In cases where NARP formation occurred on the same muscle on 2 or more different days, the ones which formed first were the most extensive. Spontaneous neurite withdrawal occurred mainly from young NARPs and resulted in their rapid disappearance. It is suggested that during the period when neurons grow and make new contacts with muscle cells there is no substantial change in their capacity to trigger the formation of new synaptic sites and maintain preexisting ones, and that the first-forming synapses on a muscle cell tend to be the largest because muscle cells have a limited capacity to generate postsynaptic membrane. Additional implications of the findings for synapse formation and elimination are discussed.
在源自非洲爪蟾胚胎的肌节肌细胞和脊髓神经元培养物中，在数天的突触形成期内监测了神经诱导的乙酰胆碱受体（AChR）簇的形成和存活情况。用荧光α-银环蛇毒素标记AChR，以便能够每天间隔观察选定神经元整个神经树突中与神经突相关的受体斑块（NARP）。为避免NARP褪色和损伤神经元，将荧光激发强度降低至3%。图像数字化后，使用基于计算机的图像分析系统测量NARP。几乎所有新形成的NARP（超过90%）在神经突与肌肉接触的同时被检测到，并且与神经突生长的近端-远端顺序相同。在6至13天大的共培养物中形成的NARP在长度、面积、强度以及面积×强度方面的分布与在1至2天大的共培养物中形成的NARP相似。NARP在这些参数上表现出每日变化，但平均而言，它们在1至2天内生长并接近其最终值。几乎所有（超过95%）只要保持接触就会存活。在同一肌肉上不同日期形成2个或更多NARP的情况下，最先形成的NARP最为广泛。自发的神经突退缩主要发生在年轻的NARP上，并导致它们迅速消失。研究表明，在神经元生长并与肌肉细胞建立新接触的期间，它们触发新突触位点形成和维持已有突触位点的能力没有实质性变化，并且肌肉细胞上最先形成的突触往往最大，因为肌肉细胞产生突触后膜的能力有限。还讨论了这些发现对突触形成和消除的其他影响。
