Neuron-astrocyte interactions mediating ethanol and noradrenergic modulation of wake-promoting vPAG dopamine neurons

项目总结 据估计，有5000万至7000万美国人患有睡眠障碍，这种疾病通常与许多 精神疾病，如酒精使用障碍(AUD)。患有AUD的人经常报告失眠，以及 睡眠困难会显著增加复发的可能性。尽管人们迫切需要控制睡眠 有助于防止复发的干扰，觉醒的基本生理学，特别是在 关于急性和慢性酒精使用的背景，人们还没有很好地了解。大脑的去甲肾上腺素能系统 许多行为状态，如觉醒、警觉和压力。急性和慢性酒精暴露 差异地改变去甲肾上腺素能神经元的活动以及去甲肾上腺素能神经元的释放、合成和周转 大脑中的去甲肾上腺素(NE)。最近，我们的实验室与合作者一起描绘了一种新的唤醒回路 从去甲肾上腺素能蓝斑(LC)到中脑导水管周围灰质腹侧的多巴胺能神经元 (VPAGDA)，刺激这个回路可以促进觉醒。急性酒精暴露也会增加 兴奋性驱动和促进觉醒的vPAGDA神经元的活动；然而，其机制尚不清楚。 α1肾上腺素能受体(α1AR)的特异性激活增加了对vPAGDA神经元的兴奋性驱动，并 随后会引起性唤醒。有趣的是，α1AR在vPAG星形胶质细胞上的表达尤其丰富，并且 激活星形细胞GQ信号足以促进觉醒。而星形胶质细胞在 通过嘌呤能信号调节其他脑区的突触传递，神经元- 酒精暴露期间VPAG中星形胶质细胞的相互作用需要进一步研究。这款F30旨在 检验中央假说，即来自邻近星形胶质细胞的嘌呤能传递介导 去甲肾上腺素能对vPAGDA神经元的调制，而乙醇增强了这一信号。我们的目标是 阐明促进尾流的vPAGDA上兴奋性驱动增加的机制细节 α受体激活神经元与去甲肾上腺素能系统的病理变化 长期酗酒。在具体目标1中，我将利用药理和病毒遗传方法 结合体外荧光成像和全细胞膜片钳电生理检测 VPAGDA神经元去甲肾上腺素能调制下的神经元-星形胶质细胞相互作用。在具体目标2中，我将 除了慢性间歇性酒精暴露的小鼠模型外，还使用类似的方法来确定 急性和慢性酒精暴露后vPAGDA神经元突触传递的变化。这个 这些研究的结果将使我们的实验室、我们的合作者和其他人能够进一步研究神经元- 星形胶质细胞生理学与酒精使用障碍的睡眠障碍有关。这将进一步推动以下工作 为开发缓解睡眠障碍的新疗法确定可能的治疗靶点 并帮助预防复发。

PROJECT SUMMARY An estimated 50-70 million Americans suffer from a sleep disorder, which is commonly comorbid with many psychiatric illnesses, such as alcohol use disorder (AUD). Individuals with AUD frequently report insomnia, and sleep difficulty significantly increases the likelihood of relapse. Despite the overwhelming need to control sleep disturbances to aid in the prevention of relapse, the underlying physiology of wakefulness, particularly in the context of acute and chronic alcohol use, is not well understood. The brain’s noradrenergic system mediates many behavioral states, such as arousal, alertness, and stress. Acute and chronic alcohol exposure differentially alter the activity of noradrenergic neurons as well as the release, synthesis, and turnover of norepinephrine (NE) in the brain. Recently, our lab alongside collaborators delineated a novel arousal circuit from the noradrenergic locus coeruleus (LC) to dopaminergic neurons in the ventral periaqueductal gray (vPAGDA), where stimulation of this circuit promotes wakefulness. Acute ethanol exposure also increases excitatory drive and the activity of wake-promoting vPAGDA neurons; however, the mechanism is unknown. Specific activation of alpha1-adrenergic receptors (α1ARs) increases excitatory drive onto vPAGDA neurons and subsequently causes arousal. Interestingly, α1AR expression is particularly enriched on vPAG astrocytes, and activation of astrocytic Gq signaling is sufficient to promote wakefulness. While astrocytes are essential in mediating synaptic transmission in other brain regions through purinergic signaling, the role of neuron- astrocyte interactions in the vPAG during ethanol exposure requires further investigation. This F30 aims to test the central hypothesis that purinergic transmission from neighboring astrocytes mediates the noradrenergic modulation of vPAGDA neurons, and that ethanol potentiates this signaling. The goal is to elucidate the mechanistic details mediating the increase in excitatory drive onto the wake-promoting vPAGDA neurons by activation of α1ARs and the pathologic changes to the noradrenergic system following acute and chronic alcohol exposure. In Specific Aim 1 I will utilize pharmacological and viral genetic approaches combined with ex vivo fluorescence imaging and whole cell patch-clamp electrophysiology to examine the neuron-astrocyte interactions underlying noradrenergic modulation of vPAGDA neurons. In Specific Aim 2 I will use similar approaches in addition to a mouse model of chronic intermittent ethanol exposure to determine the changes in synaptic transmission onto vPAGDA neurons following acute and chronic ethanol exposure. The results of these studies will allow our lab, our collaborators, and others to further investigate the role of neuron- astrocyte physiology related to sleep dysfunction in alcohol use disorder. This will further the work towards identifying possible therapeutic targets for the development of novel therapies to alleviate disordered sleeping and aid in relapse prevention.