VOLATILE ANESTHETICS AND NMDA RECEPTORS

挥发性麻醉剂和 NMDA 受体

• 批准号: 3305837
• 负责人: Robert S. Aronstam
• 金额: $ 16.15万
• 依托单位: GUTHRIE FOUNDATION FOR EDUCATION AND RES
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-09-30 至 1995-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3305837
• 关键词: NMDA receptors; anesthetics; biological signal transduction; brain metabolism; calcium metabolism; chemical binding; chloroform; cyclopropanes; enflurane; glutamates; glycine; halothane; inhibitor /antagonist; laboratory rat; membrane channels; neural inhibition; neurochemistry; nitrous oxide; polyamines; radiotracer; stimulant /agonist

The goal of the proposed research is to understand the effects of volatile anesthetics on the N-methyl-D-aspartate (NMDA) receptor-channel complex and to appreciate the role of these actions in the development and maintenance of the anesthetic state. We have shown that 2 volatile anesthetics (enflurane and halothane) selectively inhibit glutamate-stimulated [3H]MK-801 binding to NMDA receptor ion channels and block NMDA receptor-mediated 45Ca flux into rat brain microvesicles. Moreover, this inhibition is attenuated by the allosteric NMDA modulator, glycine. These findings raise the intriguing possibility that volatile anesthetics depress synaptic transmission at NMDA receptors by disrupting glutamate activation (opening) of the intrinsic ion channel. The hypotheses to be tested are that volatile anesthetics depress excitatory neurotransmission at NMDA receptors by disrupting 1) agonist binding to the glutamate recognition site, 2) ligand binding to the glycine modulatory site, and/or 3) glutamate, glycine, cation and polyamine activation of NMDA receptor ion channels. These hypotheses will be tested using radiolabelled probes for agonist, regulatory and ion channel binding sites on the NMDA receptor complex, as well as direct measurement of receptor-generated intra-cellular calcium signals. Accordingly, the specific aims are to determine the effects of several volatile anesthetics (halothane, enflurane, isoflurane, diethyl ether, cyclopropane, nitrous oxide and chloroform) on the following processes in rat brain (hippocampus): 1) Ligand binding to glutamate binding sites on the NMDA receptor complex, including receptor density, affinity and kinetic constants using [3H]CGS 19755 as a specific probe. 2) Ligand binding to the glycine modulatory site, using [3H]CGS 19755 as a specific probe. 3) Glutamate, glycine, divalent cation and spermidine activation of NMDA receptor ion channels, using [3H]MK-801 as the probe. [3H]MK-801 binding is allosterically stimulated by glutamate, glycine and polyamines (such as spermidine) and inhibited by divalent cations (such as Zn2+). 4) NMDA-receptor mediated changes in calcium content of rat brain microvesicles due to flux through transmembrane channels and release from intracellular stores, using a fluorescent dye and 45Ca. The potency of the different anesthetics in affecting NMDA receptor binding functions will be compared to their 1) anaesthetic potency, 2) physiochemical properties, and 3) membrane-perturbing actions, in order to assess the relationship of these actions to the elaboration of the anesthetic state. This research will increase our understanding of volatile anesthetic action on signal transduction processes in the brain, and explore a mechanism that may contribute to the development of the anesthetic state.

拟议研究的目标是了解挥发性物质的影响， N-甲基-D-天冬氨酸（NMDA）受体通道复合物上的麻醉剂， 了解这些行动在发展和维护 麻醉状态。 我们已经证明，2种挥发性麻醉剂（安氟醚和氟烷） 选择性抑制谷氨酸刺激的[3 H]MK-801与NMDA的结合 阻断NMDA受体介导的~（45）Ca流入大鼠体内 脑微泡 此外，这种抑制作用被 别构NMDA调节剂，甘氨酸。 这些发现提出了有趣的 挥发性麻醉剂抑制NMDA突触传递的可能性 受体通过破坏谷氨酸激活（开放）的内在离子 频道 待检验的假设是挥发性麻醉剂抑制 兴奋性神经传递在NMDA受体通过破坏1）激动剂 结合谷氨酸识别位点，2）配体结合甘氨酸 调节位点，和/或3）谷氨酸、甘氨酸、阳离子和多胺 NMDA受体离子通道的激活。 这些假设将得到检验 使用放射性标记探针进行激动剂、调节剂和离子通道结合 NMDA受体复合物上的位点，以及直接测量 受体产生的细胞内钙信号。 因此，具体目标是确定几个因素的影响， 挥发性麻醉剂（氟烷，恩氟烷，异氟烷，乙醚， 环丙烷、一氧化二氮和氯仿）对下列过程的影响， 大鼠脑（海马）： 1)配体与NMDA受体复合物上的谷氨酸结合位点结合， 包括受体密度、亲和力和动力学常数，使用[3 H]CGS 19755作为特异性探针。 2)使用[3 H]CGS 19755作为配体与甘氨酸调节位点的结合， 特异性探针 3)谷氨酸、甘氨酸、二价阳离子和亚精胺激活NMDA 受体离子通道，以[3 H]MK-801为探针。 [3 H]MK-801结合 被谷氨酸、甘氨酸和多胺（如 亚精胺）和抑制二价阳离子（如Zn 2+）。 4)NMDA受体介导的大鼠脑内钙含量变化 微囊泡由于通过跨膜通道的通量和从 细胞内储存，使用荧光染料和45 Ca。 不同麻醉药对NMDA受体结合的影响 功能将与其1）麻醉效力，2） 生理化学性质，和3）膜扰动作用，以 评估这些行动与制定 麻醉状态 本研究将增加我们对挥发性麻醉剂作用的了解 对大脑中信号转导过程的研究，并探索一种机制， 可能会导致麻醉状态的发展。