Chen B M, Grinnell A D
Department of Physiology, Jerry Lewis Neuromuscular Research Center, University of California, Los Angeles 90024.
Adv Second Messenger Phosphoprotein Res. 1994;29:383-98.
Changes in muscle length cause large changes in the probability of transmitter release from frog motor nerve terminals. A 5% to 10% stretch from rest length can increase EPP amplitude or mEPP frequency by more than 100%. The phenomenon is fully reversible and extremely rapid. Within 7-10 milliseconds of the stretch, the enhancement is complete, and it is maintained essentially constant at the new level for as long as the stretch is sustained. Given these properties, the length modulation of release is unquestionably of functional importance, strongly amplifying the spinal stretch reflex. The stretch-induced enhancement of transmitter release persists at a reduced level in a 0 Ca++, 2 mM Mg++ Ringer. This finding indicates a lack of dependence on Ca++ influx from outside the terminal. Release of Ca++ from intracellular stores close to release sites cannot be ruled out as a contributing factor. Our results, however, suggest a mechanism involving physical connections between the extracellular matrix and the nerve terminal that can alter release probability directly. Morphological evidence for connections that might be responsible can be demonstrated in micrographs of deep-etched freeze fractures through neuromuscular junctions. Hypothesizing that the ECM-nerve terminal connections responsible for the stretch effect involve proteins from the integrin family and knowing that many of the integrin-ECM binding interactions occur at sites on the ECM proteins containing the amino acid sequence RGD, we treated preparations with 0 Ca++, 2 mM Mg++ Ringer to reduce integrin binding and then returned the muscle to normal Ringer containing 0.1-0.2 mM of a six-amino-acid peptide containing the RGD sequence. This peptide strongly suppressed the stretch effect, while a control peptide (RGE) had no effect. A 50 microM Ca++/50 microM Mg++ Ringer had little effect on stretch enhancement but permitted a strong inhibition of enhancement when RGD was present. The identity of the ECM molecule(s), the integrin(s), and the mechanism of enhancement of release are unknown. However, our findings imply that much or all of the length-dependent modulation of release probability is mediated by an RGD-sensitive integrin-ECM interaction that depends more on external Ca++ than on Mg++.
肌肉长度的变化会导致青蛙运动神经末梢递质释放概率发生巨大变化。从静息长度拉伸5%至10%可使终板电位（EPP）幅度或微小终板电位（mEPP）频率增加超过100%。这种现象完全可逆且极其迅速。在拉伸后的7 - 10毫秒内，增强作用就已完成，并且只要拉伸持续，它就会在新的水平上基本保持恒定。鉴于这些特性，释放的长度调节无疑具有功能重要性，极大地增强了脊髓牵张反射。在无钙、2 mM镁的任氏液中，拉伸诱导的递质释放增强仍会在降低的水平持续存在。这一发现表明其不依赖于从神经末梢外部流入的钙离子。不能排除靠近释放位点的细胞内钙库释放钙离子作为一个促成因素。然而，我们的结果提示了一种机制，该机制涉及细胞外基质与神经末梢之间的物理连接，可直接改变释放概率。在通过神经肌肉接头的深度蚀刻冷冻断裂显微照片中可以显示出可能与此相关的连接的形态学证据。假设负责拉伸效应的细胞外基质 - 神经末梢连接涉及整合素家族的蛋白质，并且知道许多整合素 - 细胞外基质结合相互作用发生在细胞外基质蛋白质上含有氨基酸序列RGD的位点，我们用无钙、2 mM镁的任氏液处理标本以减少整合素结合，然后将肌肉放回含有0.1 - 0.2 mM含RGD序列的六氨基酸肽的正常任氏液中。这种肽强烈抑制了拉伸效应，而对照肽（RGE）则没有作用。50 microM钙/50 microM镁的任氏液对拉伸增强作用影响很小，但当存在RGD时允许对增强作用进行强烈抑制。细胞外基质分子、整合素以及释放增强机制的身份尚不清楚。然而，我们的发现意味着释放概率的大部分或全部长度依赖性调节是由一种对RGD敏感的整合素 - 细胞外基质相互作用介导的，这种相互作用更多地依赖于外部钙离子而非镁离子。
