Synaptic Mechanisms of General Anesthetic Action

描述（由申请人提供）：对于这样一种广泛使用和医学上重要的药物类别来说，全身麻醉剂的药理学和毒理学是非常不完整的，这种药物被越来越多的老年人和病情越来越重的患者使用。对麻醉作用机制的了解还不足以解释麻醉剂如何产生健忘症、无意识或（随着剂量的增加）不动，这是全身麻醉的主要特征。麻醉剂对突触传递具有强大的特异性作用，包括突触前对神经递质释放的作用和突触后对受体的作用。本研究计划的主要目的是通过实验将麻醉对神经递质释放的影响从突触后的作用中分离出来，从而了解麻醉对神经递质释放的突触前机制。突触前作用可能涉及麻醉药的治疗作用（无意识、健忘症、不动）和/或其毒性作用（神经毒性、呼吸抑制、心血管抑制）。了解麻醉药的突触机制对于开发具有更好副作用的麻醉药和优化当前高危患者的麻醉技术至关重要。我们已经证明，全身麻醉剂通过突触前机制抑制谷氨酸释放，这些作用是递质特异性的，涉及特定Na+通道亚型的区域特异性抑制。我们现在建议将重点放在挥发性麻醉剂的区域和递质特异性作用以及Na+通道阻断机制上，以便更全面地了解它们的突触前作用。我们的中心假设是全身麻醉剂通过突触前离子通道影响突触特异性机制来影响神经递质释放。我们将使用综合和协作的多学科方法来验证这一假设，具体目的如下：目的1：确定挥发性麻醉剂对孤立神经末梢神经递质释放的不同影响机制，以验证由于突触前机制的差异，它们对递质释放具有突触特异性影响的假设；目的2：确定挥发性麻醉药对完整神经元胞吐的神经递质特异性作用和机制，以验证它们通过神经递质特异性和离子通道依赖性机制不同地抑制突触囊泡胞吐的假设；目的3：确定挥发性麻醉剂对电压门控Na+通道的作用机制和调控，以验证其通过状态依赖机制抑制Na+通道亚型的假设。补充方法包括分析麻醉对完整神经末梢递质释放的影响，单个培养海马神经元的突触囊泡胞吐，以及特定Na+通道亚型的生物物理特性。这些研究对于突触前麻醉机制的分子理解以及对兴奋性和抑制性突触传递的理想和潜在毒性麻醉作用之间的平衡至关重要。

DESCRIPTION (provided by applicant): The pharmacology and toxicology of general anesthetics are remarkably incomplete for such a widely used and medically important class of drugs that are administered to increasingly older and sicker patients. Knowledge of the mechanisms of anesthetic action is insufficient to explain how any anesthetic produces amnesia, unconsciousness or immobilization (with increasing doses), the cardinal features of general anesthesia. Anesthetics have potent and specific effects on synaptic transmission, including both presynaptic actions on the release of neurotransmitters and postsynaptic actions on receptors. The principal objective of this research proposal is to understand the presynaptic mechanisms of anesthetic effects on neurotransmitter release by experimentally isolating these effects from their better understood postsynaptic actions. Presynaptic actions could be involved in therapeutic effects (unconsciousness, amnesia, immobility) and/or their toxic effects (neurotoxicity, respiratory depression, cardiovascular depression) of anesthetics. Understanding synaptic mechanisms of anesthetics is essential for development of anesthetics with improved side-effect profiles and for optimization of current anesthetic techniques in high-risk patients. We have shown that general anesthetics inhibit glutamate release by presynaptic mechanisms and that these effects are transmitter-specific and involve region-specific inhibition of specific Na+ channel subtypes. We now propose to focus on the region- and transmitter-specific actions and Na+ channel blocking mechanisms of volatile anesthetics in order to more fully understand their presynaptic actions. Our central hypothesis is that general anesthetics affect neurotransmitter release by synapse-specific mechanisms due to effects on presynaptic ion channels. We will test this hypothesis using an integrative and collaborative multidisciplinary approach by the following Specific Aims: Aim 1-Determine the mechanisms by which volatile anesthetics differentially affect neurotransmitter release from isolated nerve terminals to test the hypothesis that they have synapse-specific effects on transmitter release due to differences in presynaptic mechanisms; Aim 2-Determine the neurotransmitter-specific effects and mechanisms of volatile anesthetics on exocytosis in intact neurons to test the hypothesis that they differentially inhibit synaptic vesicle exocytosis by neurotransmitter-specific and ion channel-dependent mechanisms; and Aim 3-Determine the mechanisms and regulation of volatile anesthetic effects on voltage- gated Na+ channels to test the hypothesis that they inhibit Na+ channel subtypes by state-dependent mechanisms. Complementary approaches include analysis of anesthetic effects on transmitter release from intact nerve terminals, synaptic vesicle exocytosis from single cultured hippocampal neurons, and biophysical properties of specific Na+ channel subtypes. Such studies are essential to a molecular understanding of presynaptic anesthetic mechanisms and the balance between desirable and potentially toxic anesthetic effects on excitatory and inhibitory synaptic transmission.