Presynaptic Mechanisms of Neurotransmission in NTS

描述(申请人提供)：心血管系统的节拍调节依赖于完整的压力感受器反射。这种反射弧首先在释放谷氨酸的孤束核(NTS)内突触。压力感受器反射功能在常见的威胁生命的疾病状态中受到损害：高血压、休克和心力衰竭。在大型、可接触的中枢神经系统终末的研究表明，突触前控制谷氨酸释放涉及离子通道和第二信使系统，这些系统调节囊泡的胞吐。不同神经元的谷氨酸调节不同，但对其机制知之甚少。调节心血管和呼吸系统的反射通路依赖于脑干神经元，这些反射是由作用于孤束核(NTS)内的脑神经初级传入启动的。我们对初级传入神经的中枢行为知之甚少。这些航站楼的面积很小，难以直接调查。我们已经开发了允许从NTS的单个神经末梢进行膜片钳记录的方法。我们的研究计划将使用这些方法来解决我们的驱动假说，即NTS中重要的调节机制取决于传入神经元的身份。我们的初步工作表明，NTS神经元提供了一个独特的机会，因为：1.我们可以直接从单个神经末梢可视化、识别、刺激和电生理记录；2.NTS接受药理上可区分的传入终末，这些传入终末形成亚类，已知起源于分子上不同的外周神经元；3.这些终末的亚群可以被标记，以提供与功能不同的传入神经元的联系。我们的计划包括两个实验室的努力，这些实验室具有适合这一问题的互补专业知识。我们将使用直接斑片记录和刺激单末梢以及成像的方法，研究频率依赖性突触抑制和多肽调节谷氨酸释放的机制。这项工作利用初级传入终末的标记来区分不同的亚类(分别为TRPV1和P2X3；C型和A型传入终末)。具体目标涉及A型和C型颅脑传入突触终末钠、钾和钙通道的不同表达以及突触前调节突触间隙钙的新机制的存在。不同亚类的传入神经末梢也将在它们对谷氨酸释放的控制和多肽的调节方面进行比较。

DESCRIPTION (provided by applicant): Beat to beat regulation of the cardiovascular system depends on an intact baroreflex. This reflex arc first synapses in the nucleus tractus solitarius (NTS) where glutamate is released. Baroreflex function is compromised in common life threatening disease states: hypertension, shock, and heart failure. Work in large, accessible CNS terminals suggests that the presynaptic control of glutamate release involves ion channels and 2nd messenger systems that regulate vesicle exocytosis. Glutamate regulation differs across neurons, but the mechanisms are poorly understood. Reflex pathways regulating the cardiovascular and respiratory systems depend on brainstem neurons and these reflexes are initiated by cranial nerve primary afferents acting within the nucleus tractus solitarius (NTS). Little is known about how primary afferents behave centrally. The small size of these terminals has made direct investigation difficult. We have developed methods to permit patch clamp recording from single nerve terminals in NTS. Our Research Plan will use these methods to address our driving hypothesis that important mechanisms of regulation in NTS depend on the identity of the afferent neuron. Our Preliminary Work demonstrates that NTS neurons offer a unique opportunity because: 1. we can directly visualize, identify, stimulate and electrophysiologically record from single nerve terminals, 2. NTS receives pharmacologically distinguishable afferent terminals that form subclasses known to arise from molecularly distinct peripheral neurons, 3. Subsets of these terminals can be labeled to provide links to functionally distinct afferents. Our Plan encompasses the efforts of two labs with complementary expertise suited to this problem. We will use direct patch recording and stimulation of single terminals, as well as imaging, to study the mechanism of frequency dependent synaptic depression and peptide modulation of glutamate release. The work capitalizes on using markers of primary afferent terminals to distinguish the various sub classes (TRPV1 and P2X3; C- and A-type afferent terminals, respectively). Specific Aims concern differential sodium, potassium and calcium channel expression across A- and C-type cranial afferent synaptic terminals plus the presence of new presynaptic mechanisms regulating synaptic cleft calcium. The different sub classes of afferent nerve terminal will be also be compared in terms of their control of glutamate release and modulation by peptides.