Investigating tonic and synaptic excitatory signaling in the bed nucleus of the stria terminalis across models of alcohol exposure

摘要/项目摘要 虽然酒精使用障碍(AUD)对谷氨酸能信号转导的影响已被广泛报道，但其作用 AUD中的β-谷氨酸受体(GluD1和GluD2)的分布尚不清楚。尽管他们并没有参与 传统的兴奋性突触对谷氨酸的反应，Glud家族被认为介导了许多更微妙的方面 兴奋性信号可能导致在AUD模型中观察到的神经元活动功能障碍。 几项全基因组关联研究也表明GluD1与AUD有关。然而，没有研究表明 直接评价了酒精暴露对GluD1功能的影响。 终纹床核(BNST)是一种前脑核，被认为是一种危重的 酒精暴露后的神经调节中枢。先前的研究表明，酒精暴露可以调节 BNST中的突触谷氨酸，但其确切发生机制尚不清楚。突触 谷氨酸能传递部分是由AMPA受体介导的，约30%的AMPA受体可以传递钙(钙- 渗透性AMPAR、CP-AMPAR)。我们的初步数据显示，慢性戒断急性期 间歇性乙醇蒸气暴露(CIE)降低CP-AMPAR突触电流和GluD1 BNST中介导的紧张性兴奋电流。尽管在此之前已经观察到GluD1在 BNST，这是它们可能参与调节BNST信号的第一个数据。这些数据还 暗示GluD1调制可能是酒精暴露后神经调节的新靶点。这 该提案旨在进一步研究CP-AMPAR和GluD1在BNST中的作用，它们可能如何相互作用，以及 用荧光原位杂交、膜片钳结合的方法研究酒精暴露对它们的影响 电生理学和体外钙成像。AIM 1将使用GluD1基因敲除(GluD1KO)小鼠和野生型 (WT)产仔建立BNST中GluD1的功能图谱及其与CP-AMPAR的重叠 表达BNST细胞。AIM 2将评估CIE对BNST CP-AMPAR和GluD1表达的影响 以及这些小鼠模型中的信号、神经元兴奋性和自发钙动力学。最后，目标3 我将使用这些方法来检查BNST CP-AMPAR和GluD1信号的翻译相关性 不同型号的澳元。

Abstract/Project Summary While the effects of alcohol use disorder (AUD) on glutamatergic signaling have been widely reported, the role of delta glutamate receptors (GluD1 and GluD2) in AUD remains unknown. Although they are not involved in traditional excitatory synaptic responses to glutamate, the GluD family is known to mediate many subtler aspects of excitatory signaling which may contribute to the dysfunctions of neuronal activity observed in models of AUD. GluD1 has also been implicated in AUD by several genome-wide association studies. However, no research has been conducted directly evaluating the effect of alcohol exposure on GluD1 function. The bed nucleus of the stria terminalis (BNST) is a forebrain nucleus that has been heavily implicated as a critical hub for neuromodulation following alcohol exposure. Previous work has shown that alcohol exposure regulates synaptic glutamate in the BNST, but the exact mechanisms by which this happen remain unclear. Synaptic glutamatergic transmission is in part mediated by AMPA receptors, ~30% of which can pass calcium (calcium- permeable AMPARs, CP-AMPARs). Our preliminary data suggests that acute withdrawal from chronic intermittent ethanol vapor exposure (CIE) decreases both CP-AMPAR synaptic current and reduces a GluD1 mediated tonic excitatory current in the BNST. Although it has been observed before that GluD1 is expressed in the BNST, these are the first data to that they may be involved in regulating BNST signaling. These data also implicate GluD1 modulation as a possible novel target for neuromodulation following alcohol exposure. This proposal aims to further examine the role of CP-AMPARs and GluD1 in the BNST, how they may interact, and how they are affected by alcohol exposure using a combination of fluorescent in situ hybridization, patch-clamp electrophysiology, and ex vivo calcium imaging. Aim 1 will use GluD1 knockout (GluD1 KO) mice and wildtype (WT) littermates to establish a functional profile of GluD1 in the BNST, as well as its overlap with CP-AMPAR expressing BNST cells. Aim 2 will then evaluate the impact of CIE on BNST CP-AMPAR and GluD1 expression and signaling, neuronal excitability, and spontaneous calcium dynamics in these mouse models. Finally, Aim 3 will use these approaches to examine the translational relevance of BNST CP-AMPAR and GluD1 signaling across different models of AUD.