IONIC CONTROL OF NERVE FIBER GROWTH AND SURVIVAL

局部应用神经生长因子调控神经纤维生长和存活的研究 提出了细胞外钾的变化。 区室化 将使用培养制备物，其中来自大鼠细胞体的轴突 位于一个隔室中的神经元穿过硅脂屏障生长， 进入不同的隔间。 以这种方式， 远端轴突可以独立于流体环境进行控制， 细胞体和近端轴突。 远端轴突可以在任何位置被移除。 时间和迅速再生。 这使得轴突再生的相同 在不同条件下依次观察神经元群体 实验条件和随着神经元在体外发育和老化。 神经突延伸的显微测量和显微摄影 将使用神经突密度的比较来评价生长。 还有， 隔室化培养物中的神经元可以长期电 刺激，允许有关的实验性能 电活动在生长和发育中的作用。 除了工作 对于区室化培养物，建议测量[K+]e的变化 通过K+选择性微电极在体内发生。 本提案中所述问题的产生是因为 在这个实验室里使用的是分区培养。 神经突 交感神经元、背根神经节神经元和脊髓神经元 可以被排除进入局部增加的细胞外 钾（[K+]e）。 此外，[K+]e沿着 神经突对神经突生长具有多种退化效应， 再生和生存：伸长可以减慢，这是一个极端。 神经突的回缩可随之发生，并可发生完全变性。 这些观察结果对神经纤维的机制有启示 延长，他们也提出了深远的可能性，自然 [K+]e在体内正常发育过程中的变化可能强烈 影响连接的建立和/或维护 神经元 例如，神经纤维在活动过程中释放的K+可能 介导突触后位点的竞争性相互作用。 也是 [K+]e变化引起的回归变化可能具有 再生、衰老和癫痫发作问题的意义 在神经系统中。 实验提出：（1）表征 局部升高的K+对外周和中枢神经元的影响 特别强调随着发育和衰老而发生的变化， 体内和体外，（2）研究这些作用的机制，（3） 开始收集关于[K+]e变化幅度的信息， 在体内发育过程中发生，以及（4）建立体外系统， 研究活动对突触形成的长期影响， 上维护

Investigations into the control of nerve fiber growth and survival by local variations in extracellular potassium are proposed. A compartmentalized culture preparation will be employed in which axons from cell bodies of rat neurons located in one compartment grow across silicone grease barriers to enter separate compartments. In this way the fluid environment of the distal axons can be controlled independently of the fluid environment of the cell bodies and proximal axons. The distal axons can be removed at any time and promptly regenerate. This allows axonal regeneration of the same population of neurons to be observed sequentially under different experimental conditions and as the neurons develop and age in vitro. Micrometric measurements of neurite extension and photomicrographic comparisons of neurite density will be used to evaluate growth. Also, neurons in compartmentalized cultures can be chronically electrically stimulated, permitting the performance of experiments concerned with the role of electrical activity in growth and development. In addition to work with compartmentalized cultures, it is proposed to measure changes in [K+]e that occur in vivo by means of K+-selective microelectrodes. The questions addressed in this proposal arise because of observations made in this laboratory employing compartmentalized cultures. Neurites of sympathetic neurons, dorsal root ganglion neurons and spinal cord neurons can be excluded from entering a region of locally increased extracellular potassium ([K+]e). Also, a proximo-distal increase in [K+]e along the neurites has a variety of regressive effects on neurite growth, regeneration, and survival: elongation can be slowed, an extreme retraction of neurites can ensue, and complete degeneration can occur. These observations have implications for the mechanism of nerve fiber elongation, and they also raise the far-reaching possibility that natural variations in [K+]e in vivo during normal development may strongly influence the establishment and/or maintenance of connections between neurons. For example, K+ released by nerve fibers during activity may mediate competitive interactions for postsynaptic sites. It is also possible that regressive changes induced by variations in [K+]e may have implications for the problems of regeneration, aging, and seizure activity in the nervous system. Experiments are proposed to: (1) characterize the effects of locally elevated K+ on peripheral and central neurons with particular emphasis on changes that occur with development and aging in vivo and in vitro, (2) investigate the mechanisms of these effects, (3) begin to collect information about what magnitude of changes in [K+]e can occur during development in vivo, and (4) establish an in vitro system to study the long-term effects of activity on synapse formation and maintenance.