Molecular Mechanisms of Ethanol Sensitivity in ILS and ISS Mice

描述（由申请人提供）：我们之前已经证明，ILS小鼠的GABAARs表现出乙醇增强，而乙醇对ISS小鼠海马中的GABAARs没有影响。同样，本研究提出的初步数据表明，乙醇增强了ILS小鼠皮质的GABAAR，而ISS小鼠没有；抑制了ILS小鼠海马和皮质的NMDAR，而ISS小鼠没有。在乙醇影响海马和皮质GABAAR和NMDAR的能力方面，ILS和ISS小鼠的这些显着差异将是我们提议的主要焦点。我们建议研究在ILS和ISS小鼠中GABAAR和NMDAR的乙醇敏感性差异的分子机制。我们预计，这两种小鼠将因此为我们提供一个独特的机会来测试关于这些机制的假设。这将是拟议研究的主要目标。为了实现这一目标，我们的建议将有三个具体目标。目标# 1。验证ILS和ISS小鼠中NMDAR和/或GABAAR磷酸化的差异可能是两种小鼠中乙醇对这些受体作用差异的基础。我们和其他人最近表明，磷酸化可能在NMDAR和GABAAR的乙醇敏感性中起重要作用。并且，我们已经证明ILS小鼠的GABAAR和NMDAR对乙醇敏感，而ISS小鼠的GABAAR和NMDAR对乙醇不敏感。因此，我们将比较ILS和ISS小鼠中NMDAR和GABAAR特定亚基和/或磷酸基的基础磷酸化和乙醇刺激磷酸化。目标# 2。验证ILS和ISS小鼠中NMDAR和/或GABAAR转运机制的乙醇敏感性差异可能是两种小鼠中乙醇对这些受体影响差异的基础。我们和其他人已经表明，NMDAR和GABAAR膜运输可能在这些受体的功能中发挥重要作用。此外，我们最近获得的初步数据表明，乙醇降低了ILS中NMDAR的膜表面表达，而在ISS小鼠中则没有。因此，在本研究中，我们将通过比较ILS和ISS小鼠NMDAR和GABAAR在基础和乙醇刺激下的表面表达来证实和扩展这一观察结果。目的3：验证调节NMDAR和GABAAR的磷酸化和/或表面表达的干预措施将减少乙醇对ILS小鼠中GABAAR和NMDAR反应的影响，并“挽救”乙醇对ISS小鼠中这些受体的影响。基于上述目标1和2的结果，我们将进行干预，以药理学方式调节磷酸化和/或表面表达。然后，我们将测试这些干预措施，以确定目的1和目的2中看到的生化差异是否必要或足以解释乙醇对ILS和ISS小鼠中这些受体诱发反应的影响差异。

DESCRIPTION (provided by applicant): We have previously shown that GABAARs in ILS mice exhibit ethanol potentiation while ethanol had no effect on GABAARs in hippocampi of ISS mice. Similarly, preliminary data presented in this proposal show that ethanol potentiates the GABAAR in cortex of ILS but not ISS mice and inhibits the NMDAR in hippocampus and cortex of ILS but not ISS mice. These dramatic differences in ILS and ISS mice in terms of the ability of ethanol to affect the GABAAR and NMDAR in hippocampus and cortex will be the principal focus of our proposal. We are proposing to study the molecular mechanisms that underlie the differences in ethanol sensitivity of the GABAAR and NMDAR in ILS and ISS mice. We anticipate that these two strains of mice will thus provide us with a unique opportunity to test hypotheses about these mechanisms. This will be the principal goal of the proposed studies. In pursuit of this goal our proposal will have three Specific Aims. Aim #1. Test the hypothesis that differences in the phosphorylation of the NMDAR and/or GABAAR in ILS and ISS mice may underlie the differences in ethanol effects on these receptors in the two strains of mice. We and others have recently shown that phosphorylation may play an important role in the ethanol sensitivity of the NMDAR and the GABAAR. And, we have shown that GABAAR and NMDAR of ILS mice are sensitive to ethanol and those of ISS mice are not. Therefore, in this aim we will compare the basal and ethanol-stimulated phosphorylation of specific subunits and/or phosphosites of the NMDAR and GABAAR in ILS and ISS mice. Aim #2. Test the hypothesis that differences in the ethanol sensitivity of the trafficking mechanisms for the NMDAR and/or GABAAR in ILS and ISS mice may underlie the differences in ethanol effects on these receptors in the two strains of mice. We and others have shown that NMDAR and GABAAR membrane trafficking may play important roles in the function of these receptors. Moreover, we have recently obtained preliminary data that ethanol reduces membrane surface expression of the NMDAR in ILS but not in ISS mice. Therefore, in this aim we will confirm and extend this observation by comparing the basal and ethanol- stimulated surface expression of the NMDAR and GABAAR in ILS and ISS mice. Aim #3 Test the hypothesis that interventions that modulate phosphorylation and/or surface expression of the NMDAR and GABAAR will reduce the effect of ethanol on GABAA R and NMDAR responses in ILS mice and and "rescue" ethanol effects on these receptors in ISS mice. Based on results from Aims 1 and 2 above, we will intervene to pharmacologically modulate phosphorylation and/or surface expression. We will then test these interventions to determine whether the biochemical differences seen in Aims 1 and 2 or either necessary or sufficient to explain the differences in ethanol effects on the evoked responses of these receptors in ILS and ISS mice.