CALCIUM WAVES IN PYRAMIDAL NEURONS

描述（由申请人提供）：海马锥体神经元突触后[Ca2+]i的变化在诱导各种形式的突触可塑性、基因表达和膜电导调节中起重要作用。所有这些机制都可以影响这些神经元在学习和记忆回路中的行为。因此，详细了解这些过程对于理解大脑功能非常重要。Ca2+通过NMDA受体进入和通过电压依赖性Ca2+通道进入已经被深入研究，但对Ca2+从内部储存释放知之甚少。我们将从CA1区研究代谢性谷氨酸受体的突触激活介导的IP3介导的Ca2+释放的特性和功能。先前的实验证实Ca2+释放在锥体细胞树突的受限区域以波的形式扩散。我们将研究控制波传播到不同区域的机制，包括斜树突、体细胞和细胞核。我们将研究非常小的Ca2+释放事件的性质和空间分布，并将它们与再生波的性质进行比较。我们将尝试确定它们是否与最小突触刺激引起的基本信号事件相对应。我们将研究Ca2+从商店释放在几个重要的生理过程中的功能，特别强调Ca2+在不同树突区域释放的不同后果。一组实验将检查树突Ca2+波是否阻断了突触可塑性中重要的反向传播动作电位。第二组实验将研究Ca2+波在抑制内源性大麻素介导的锥体神经元突触抑制中的作用。第三组实验将检查Ca2+释放在诱导长期增强中的作用。我们将在年轻的斯普拉格·道利大鼠的海马体上做这些实验。我们将采用来自体细胞和树突的全细胞斑块记录，并使用冷却CCD相机和2光子共聚焦显微镜对[Ca2+]i的变化进行高速测量，以检测注射Ca2+指示剂的荧光变化。我们将使用细胞外谷氨酸和细胞内IP3和Ca2+的局部解锁来实现在厚或薄树突区域的信号事件的精确定位。

DESCRIPTION (provided by applicant): Postsynaptic [Ca2+]i changes in pyramidal neurons in the hippocampus play an important role in the induction of various forms of synaptic plasticity, gene expression, and modulation of membrane conductances. All of these mechanisms can affect the behavior of these neurons in circuits involved in learning and memory. Therefore, a detailed understanding of these processes is important for understanding brain function. Ca2+ entry through NMDA receptors and entry through voltage-dependent Ca2+ channels have been studied intensively but less is known about Ca2+ release from internal stores. We will investigate the properties and functions of Ca2+ release mediated by IP3 mobilized by synaptic activation of metabotropic glutamate receptors in pyramidal cells from the CA1 region. Previous experiments established that Ca2+ release spreads as a wave in restricted regions of pyramidal cell dendrites. We will examine the mechanisms that control the propagation of the waves into different regions including oblique dendrites, the soma and nucleus. We will examine the properties and spatial distribution of very small Ca2+ release events and compare them to the properties of regenerative waves. We will try to determine if they correspond to fundamental signaling events evoked by minimal synaptic stimulation. We will investigate the function of Ca2+ released from stores in several important physiological processes, with particular emphasis on the different consequences of Ca2+ released in different dendritic regions. One set of experiments will examine whether dendritic Ca2+ waves block back propagating action potentials important in synaptic plasticity. A second set of experiments will examine the role of the Ca2+ waves in suppressing synaptic inhibition onto pyramidal neurons mediated by endogenous cannabinoids. A third set of experiments will examine the role of Ca2+ release in the induction of long term potentiation. We will make these experiments in slices from the hippocampus of young Sprague Dawley rats. We will employ whole cell patch recordings from the soma and dendrites, and make high speed measurements of [Ca2+]i changes using cooled CCD cameras and 2- photon confocal microscopy to detect the fluorescence changes of injected Ca2+ indicators. We will use focal uncaging of extracellular glutamate and intracellular IP3 and Ca2+ to achieve precise localization of signaling events in thick or thin dendritic regions.