Synaptic mechanisms underlying reward seeking and compulsive drug use

项目1：中脑多巴胺神经元的谷氨酸和多巴胺传递具有相似的释放特性，但受可卡因的影响不同。(Adrover，Shin和Alvarez，J.NeuroScience，2014)腹侧被盖区和伏隔核(NAC)之间的突触传递在奖赏动机行为中至关重要，并被认为在成瘾中发生改变。除了多巴胺(DA)，谷氨酸还被中脑边缘DA神经元的子集包装和释放，诱导EPSCs进入NAC的中等棘神经元。关于DA中脑终末谷氨酸释放的性质和调节以及可卡因的作用，人们知之甚少。使用光遗传学方法选择性激活中脑DA纤维，我们比较了用快扫描循环伏安法和全细胞记录在小鼠脑片上测量的DA瞬变和EPSCs的性质和调制。DA瞬时和EPSCs被DA受体D2R激动剂抑制，并呈现明显的成对脉冲抑制，需要2分钟才能完全恢复。可卡因使EPSCs波幅降低50%，但增强了中脑DA神经元的DA传递。AMPA和NMDA受体介导的EPSCs同样受到可卡因的抑制，提示为突触前机制。药物阻断和D2R基因缺失可阻断可卡因对EPSCs的抑制作用，并使DA瞬变峰显著升高，证实突触前D2R参与其中。这些发现表明，急性可卡因通过突触前D2R抑制中脑DA神经元释放DA和谷氨酸，但对它们在NAC中的传递有不同的整体影响。我们推测，可卡因通过阻断DA的再摄取，延长了DA的瞬变，并促进了对DA和谷氨酸从这些终末释放的反馈抑制。 项目2：反复酗酒改变酒精饮用模式并抑制纹状体GABA能传递(Wilcox等人，神经精神药理学，2014) 对大量和狂饮酒精的神经生物学的研究一直受到大多数小鼠品系表现出的自愿酒精消耗水平低的限制(Crabbe等人，2011年)。最近，一种间歇性接触乙醇的模型已被证明在小鼠中引起类似狂饮的饮酒和与药物相关的血液乙醇浓度(BECs)(Rhodes等人，2005年)。这个被称为在黑暗中饮酒的模型利用了老鼠的昼夜节律模式，在两个小时的饮酒过程中实现了可靠的高水平饮酒。C57BL/6J小鼠的BECs达到80 mg/dl以上，并表现出中毒迹象，如运动障碍(Rhodes等人，2007年)。DID是一种稳健的范例，已成功用于研究调节酗酒样酒精消费的神经元电路和信号(Sprow，2012)。尽管这种间歇性获取模式取得了成功，但人们并不完全了解自愿饮用乙醇的潜在机制。在这项研究中，我们在我们的实验室建立了DID，并表明它可以可靠地增加自愿乙醇摄入量和血液中的乙醇浓度。我们用挠度计描述了间歇性饮酒小鼠的饮酒模式，以高时间分辨率记录了自愿饮酒数周的每一次饮酒，并分析了最后一次饮酒后2天和30天纹状体神经元的突触形态。这些结果代表了对酒精领域的一项新的和重要的贡献，因为他们发现了瓶子交换，这是DID的一个组成部分，作为一种可能的机制，间歇接触通过诱导小鼠以更高的速度饮酒，促进了自愿饮酒行为的获得。随着时间的推移，小鼠改变了他们的酒精饮用模式，并在每次DID会议开始时提高了饮用速度。更快的饮酒与更高的BEC相关，并且在治疗6周后观察到对酒精的偏好增强。有趣的是，纹状体和伏隔脊椎的密度没有发现变化，棘突长度的缩短只是暂时的，这表明这些行为变化与脑内突触形态的长期变化无关。

Project 1: Glutamate and dopamine transmission from midbrain dopamine neurons share similar release properties but are differentially affected by cocaine.(Adrover, Shin and Alvarez, J. Neuroscience 2014)Synaptic transmission between ventral tegmental area and nucleus accumbens (NAc) is critically involved in reward-motivated behaviors and thought to be altered in addiction. In addition to dopamine (DA), glutamate is packaged and released by a subset of mesolimbic DA neurons, eliciting EPSCs onto medium spiny neurons in NAc. Little is known about the properties and modulation of glutamate release from DA midbrain terminals and the effect of cocaine. Using an optogenetic approach to selectively activate midbrain DA fibers, we compared the properties and modulation of DA transients and EPSCs measured using fast-scan cyclic voltammetry and whole-cell recordings in mouse brain slices. DA transients and EPSCs were inhibited by DA receptor D2R agonist and showed a marked paired-pulse depression that required 2 min for full recovery. Cocaine depressed EPSCs amplitude by 50% but enhanced the overall DA transmission from midbrain DA neurons. AMPA and NMDA receptor-mediated EPSCs were equally inhibited by cocaine, suggesting a presynaptic mechanism of action. Pharmacological blockage and genetic deletion of D2R in DA neurons prevented the cocaine-induced inhibition of EPSCs and caused a larger increase in DA transient peak, confirming the involvement of presynaptic D2R. These findings demonstrate that acute cocaine inhibits DA and glutamate release from midbrain DA neurons via presynaptic D2R but has differential overall effects on their transmissions in the NAc. We postulate that cocaine, by blocking DA reuptake, prolongs DA transients and facilitates the feedback inhibition of DA and glutamate release from these terminals. Project 2: Repeated binge-like ethanol drinking alters ethanol drinking patterns and depresses striatal GABAergic transmission.(Wilcox et al., Neuropsychopharmacology 2014) Research into the neurobiology of heavy and binge-like ethanol drinking has been limited by the low-levels of voluntary ethanol consumption shown by most mouse strains (Crabbe et al., 2011). Recently, a model of intermittent access to ethanol has been shown to elicit binge-like drinking and pharmacologically relevant blood ethanol concentrations (BECs) in mice (Rhodes et al., 2005). Termed Drinking in the Dark, this model takes advantage of the circadian patterns of mice to achieve reliably high levels of consumption in a two hour drinking session. C57BL/6J mice reach BECs higher than 80 mg/dl, and show signs of intoxication such as motor impairment (Rhodes et al., 2007). DID is a robust paradigm that has been successfully used to investigate neuronal circuits and signals that modulate binge-like ethanol consumption (Sprow, 2012). Despite the success of this intermittent access model, the mechanisms underlying the acquisition of voluntary ethanol drinking are not completely understood. In this study, we established DID in our laboratory and showed that it produces reliable escalation of voluntary ethanol intake and blood ethanol concentration. We characterized the drinking pattern of mice with intermittent access to ethanol using lickometers to record each bout with high temporal resolution over many weeks of voluntary ethanol consumption, and analyzed the synaptic morphology of striatal neurons 2 days and 30 days after the last ethanol binge. The results represent a novel and important contribution to the alcohol field because they identify the bottle exchange, an integral part of DID, as a likely mechanism by which intermittent access facilitates the acquisition of voluntary ethanol drinking behavior by inducing mice to drink at a higher rate. Over time, mice change their ethanol drinking pattern and increase the speed of drinking at the beginning of each DID session. Faster ethanol drinking is associated with higher BEC, and an enhanced preference for ethanol was observed after 6 weeks of treatment. Interestingly, no changes were detected in either striatal or accumbal spine density, and a shortening of spine length was seen only transiently, suggesting that these behavioral changes occur independent of long-term changes in synaptic morphology in brain.