INTERACTIONS BETWEEN H+.1 AND CA2+1 IN MYXICOLA AXON

长期目标是更好地了解 调节细胞内游离钙（Ca 2+）i和游离钙（Ca 2+）i， 质子，（H+），以及这两种离子之间的相互依赖性 浓度的 现在已知或怀疑这些离子 调节或影响许多细胞功能，包括肌肉 收缩，神经分泌，离子渗透性，细胞间 电偶联、细胞分裂和分化，以及 新陈代谢. 钙调节机制将在 四个位点：（a）内质网，（B）质膜，（c） 细胞内Ca结合蛋白，和（d）线粒体。 钙 神经细胞内质网摄取和释放 将使用新的Ca电极表征细胞质， 检测nM浓度范围内的游离Ca。 atp依赖 将在存在线粒体的情况下研究Ca摄取。 抑制剂作为ATP、pH和（cA 2+）的函数。 的影响 咖啡因和肌醇磷酸盐对细胞器钙释放的影响 靠近质膜的细胞将被比作细胞器 在轴突的中心附近。 细胞内钙结合蛋白将 特别强调对神经丝的研究。 十五万- 160，000分子量的神经丝蛋白将被分离 Ca与这些蛋白质的结合将被定量地 在不同pH值、离子强度和 温度 使用有限的、特异性的蛋白水解切割， 放射自显影分析，神经丝的部分 将检查含有钙结合位点的分子。 的 将研究pH对线粒体的影响。 使用 四酚鳞（TPP+），TPP+-微电极，和 含有线粒体的轴浆样品，pH对 将检查线粒体膜电位以查看是否 这解释了先前关于pH和Ca吸收的观察结果。 使用pH微电极和分离的细胞质样品， 神经细胞质中“有效”氢离子扩散系数 将被确定，固定和移动的pH值的重要性 将评估缓冲液在消散细胞内pH梯度中的作用。

Long-term objectives are to gain a better understanding of the regulation of both intracellular free calcium (Ca2+) i, and free protons, (H+), and the interdependence between these two ion concentrations. These ions are now known or suspected to regulate or influence numberous cell functions, including muscle contraction, neuro-secretion, ion permeability, intercellular electrical coupling, cell division and differentiation, and metabolism. Calcuium regulatory mechanisms will be examined at four loci: (a) endoplasmic reticulum, (b) plasma membrane, (c) intracellular Ca-binding proteins, and (d) mitochondria. Calcium uptake and release from the endoplasmic reticulum of neural cytoplasm iwll be characterized using a new Ca electrode that detects free Ca in the nM concentration range. ATP-dependent Ca uptake will be studied in the presence of mitochondrial inhibitors as a function of ATP, pH, and (cA2+). The effects of caffeine and inositol phosphates on Ca release from organelles close to the plasma membrane will be compared to organelles near the center of the axon. Intracellular Ca-binding proteins will be studied with special emphasis on neurofilaments. The 150,000- 160,000 molecular-weight neurofilament proteins will be isolated and the binding of Ca to these proteins will be quantitiatively examined in solutions of different pH, ionic strength, and temperature. Using limited, specific proteolytic cleavage and autoradiographic analysis, the portions of the neurofilament molecule containg the calcium-binding site will be examined. The influence of pH on mitochondria will be studied. Using tetraphenol phosphonium (TPP+), a TPP+-microelectrode, and samples of axoplasm containing mitochondria, the effect of pH on the mitochondrial membrane potential will be examined to see if this explains a previus observation concerning pH and Ca uptake. Using pH microelectrodes and isolated samples of cytoplasm, the "effective" hydrogen ion diffusion coefficient in neural cytoplasm will be determined, and the importance of fixed and mobile pH buffers in dissipating intracellular pH gradients will be evaluated.