Central amygdala input circuits control stress-induced anxiety after chronic ETOH

杏仁核中央输入电路控制慢性 ETOH 后压力引起的焦虑

• 批准号: 8868866
• 负责人: GEORGE R BREESE
• 金额: $ 29.49万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-07-15 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8868866
• 关键词: Abstinence; Address; Affect; Agonist; Alcoholism; Alcohols; Amygdaloid structure; Anxiety; Brain; Chronic; Clinical; Complement; Distant; Ethanol; Figs - dietary; Functional Magnetic Resonance Imaging; Glutamates; Halorhodopsins; Health; Hypothalamic structure; Investigation; Kindling (Neurology); Knowledge; Lateral; Link; Medial; Modeling; Neurons; Output; Oxytocin; Oxytocin Receptor; Pharmacology; Play; Rhodopsin; Role; Series; Site; Stress; Symptoms; Synapses; Testing; Thalamic structure; Therapeutic; Vasopressin Receptor; Vasopressins; alcohol abstinence; alcohol exposure; anxiety-like behavior; base; craving; expression vector; improved; innovation; midbrain central gray substance; neural circuit; neuropathology; optogenetics; prevent; problem drinker; promoter; relating to nervous system; response; social; vector

DESCRIPTION (provided by applicant): During abstinence from alcohol, stress in alcoholics can result in negative affect and craving (1), a response which is accompanied by a change in brain functional magnetic resonance imaging (fMRI)-effects of stress not seen in social drinkers (2). To model the negative affect induced by stress during abstinence in alcoholics, an extended period of chronic intermittent alcohol (CIA) exposure was found to cause an enduring adaptation that sensitizes stress-induced negative affect during abstinence. These series of clinical and basic findings are consistent with the kindling/stress hypothesis of alcoholism. Even though the central amygdala (CeA) is known to play an important role in stress-induced negative affect after CIA exposure, unknown is the role input circuits to the CeA have in facilitating this stress-induced anxiety. In addition to CRF, other inputs that synapse on CeA neurons are glutamate (GLU) terminals from the basolateral amygdala (BLA), oxytocin (OXY) terminals from the hypothalamus, and vasopressin (VP ) terminals. Nonetheless, the influence these other inputs have on stress-induced anxiety after CIA exposure has not been identified. Likewise unidentified is whether CeA medial division (CEM) neurons that output terminals to the periaqueductal gray (PAG) and other sites are a critical component of the neural circuit that supports stress-induced anxiety associated with CIA. To guide addressing these unknowns, the hypothesis tested is that stress facilitates anxiety during abstinence from CIA exposure by influencing a neural circuit composed of terminal inputs to the lateral (CEL) and medial (CEM) divisions of the CeA that modulate CEM output. First, to permit optogenetic investigations of the potential role BLA terminals have in facilitating stress-induced anxiety after CIA exposure, an AAV5-eYFP vector with a CAMKII promoter containing the rhodopsin derivatives halorhodopsin (NpHR3.0) or channel- rhodopsin (ChR2) will be placed into the BLA to confirm localization of GLU terminals from the BLA to the lateral (CEL) and the medial (CEM) divisions of the CeA. Subsequently tested with optogenetics will be whether ChR2 activation or NpHR3.0 inhibition of BLA terminal release of GLU on CEL neurons will influence facilitation of stress-induced anxiety after CIA exposure. This latter optogenetic strategy will be complemented by determining if activation of CEL neurons with an OXY receptor agonist will block facilitation of stress-induced anxiety after CIA exposure. Upon completing investigations of CEL neural inputs, optogenetic inhibition or excitation of GLU-containing BLA terminals synapsing on CEM neurons will assess if the stress-induced anxiogenic action after CIA exposure is affected. Because VP-containing terminals synapse on CEM neurons to induce anxiety- like behavior, it is reasoned that activation of CEM neurons with VP may contribute to the anxiety induced by stress following CIA exposure. To confirm a direct involvement of CEM neurons in VP action, an AAV-eYFP vector with the NpHR3 expressed in CEM neurons will test if optogenetic inhibition of CEM neurons will alter both stress- as well as VP-induced anxiety after CIA exposure. To confirm that VP action contributes to stress-induced anxiety after CIA exposure, VP receptor subtypes will be pharmacologically antagonized in the CEM prior to stress to determine if the stress- induced anxiety after CIA exposure is prevented. Upon confirmation of CEM terminal presence in the PAG, involvement of these CEM terminals in the PAG in stress- and VP-induced anxiety observed after CIA exposure will be assessed by optogenetic inhibition or excitation of these CEM terminals- an approach to implicate both the CEM and the PAG in facilitation of the stress-induced anxiety after CIA exposure. Collectively, the proposed pharmacological and optogenetic strategies utilized can be expected to define whether involvement of specific neural inputs to the CEL and CEM, which are accompanied by CEM output to the PAG and other sites, form a circuit that influences facilitation of stress-induced negative affect after CIA exposure. This proposed circuit involving the CeA is felt to be associated with the neuropathology responsible for the facilitated negative affect observed to stress that facilitates craving in alcoholics durin abstinence. New knowledge concerning adaptations in this circuit after CIA exposure could possibly provide clues by which to minimize the barrier to developing new and improved therapeutics to treat the negative symptoms to stress observed in abstinent alcoholics.

描述（由申请人提供）：在戒酒期间，酗酒者的压力会导致负面影响和渴望(1)，这种反应伴随着脑功能磁共振成像（fMRI）的变化——在社交饮酒者中没有看到压力的影响(2)。为了模拟戒酒期间压力引起的负面影响，我们发现长时间的慢性间歇性酒精（CIA）暴露会导致戒酒期间压力引起的负面影响的持久适应。这一系列的临床和基础发现与酒精中毒的点燃/压力假说是一致的。尽管中央杏仁核（CeA）已知在CIA暴露后应激诱导的负面影响中起重要作用，但CeA的输入回路在促进这种应激诱导的焦虑中的作用尚不清楚。除了CRF外，CeA神经元上突触的其他输入还有来自基底外侧杏仁核（BLA）的谷氨酸（GLU）终端，来自下丘脑的催产素（OXY）终端和加压素（VP）终端。尽管如此，这些其他输入对CIA暴露后压力引起的焦虑的影响尚未确定。同样不确定的是，向导水管周围灰质（PAG）和其他部位输出终端的CeA内侧分裂（CEM）神经元是否是支持与CIA相关的应激性焦虑的神经回路的关键组成部分。为了指导解决这些未知的问题，我们测试的假设是，压力通过影响一个神经回路，该神经回路由调节CEM输出的CeA的外侧（CEL）和内侧（CEM）分支的终端输入组成，从而促进了CIA暴露戒断期间的焦虑。首先，为了对CIA暴露后BLA末端在促进应激诱导焦虑中的潜在作用进行光遗传学研究，将一个带有CAMKII启动子的AAV5-eYFP载体放入BLA中，该载体含有视紫红质衍生物halorhodopsin （NpHR3.0）或通道视紫红质（ChR2），以确认GLU末端从BLA定位到CeA的外侧（CEL）和内侧（CEM）分裂。随后，我们将利用光遗传学的方法测试在CIA暴露后，CEL神经元上GLU的BLA末端释放的ChR2激活或NpHR3.0抑制是否会影响应激性焦虑的促进。后一种光遗传学策略将通过确定使用OXY受体激动剂激活CEL神经元是否会阻断CIA暴露后应激性焦虑的促进来补充。在完成对CEL神经输入的研究后，光遗传学抑制或激发在CEM神经元上突触的含有glu的BLA终端将评估CIA暴露后应激诱导的焦虑作用是否受到影响。由于含有VP的神经末梢在CEM神经元上突触诱导焦虑样行为，因此可以推断，VP激活CEM神经元可能有助于CIA暴露后应激引起的焦虑。为了证实CEM神经元直接参与VP的作用，在CEM神经元中表达NpHR3的AAV-eYFP载体将测试CEM神经元的光遗传抑制是否会改变CIA暴露后应激和VP诱导的焦虑。为了证实VP作用有助于CIA暴露后应激诱导的焦虑，我们将在应激前在CEM中对VP受体亚型进行药理学拮抗，以确定CIA暴露后应激诱导的焦虑是否被预防。在确认CEM终端存在于PAG后，将通过光遗传抑制或激发CEM终端来评估这些CEM终端在CIA暴露后所观察到的应激和vp诱导的焦虑中所参与的PAG，这一方法暗示CEM和PAG在CIA暴露后应激诱导焦虑中的促进作用。总的来说，所提出的药理学和光遗传学策略可以用来确定特定的神经输入是否参与CEL和CEM，并伴随CEM输出到PAG和其他部位，形成一个回路，影响CIA暴露后应激诱导的负面影响的促进。这个涉及CeA的电路被认为与神经病理学有关，负责观察到的压力促进负面影响，在戒酒期间促进酗酒者的渴望。有关CIA暴露后该回路适应的新知识可能提供线索，从而最大限度地减少开发新的和改进的治疗方法的障碍，以治疗在戒断酗酒者中观察到的负面压力症状。