Astrocytic mechanism for regulating dopaminergic burst firing

 描述(申请人提供)：位于黑质致密部和腹侧被盖区的多巴胺(DA)神经元产生与动机行为有关的信号。DA神经元的放电模式编码了奖赏的不同方面。滥用药物，如可卡因，会篡夺多巴胺能系统，导致多巴胺能爆发放电增加，从而增加目标区域的多巴胺释放，达到自然奖励(如进食、梳妆、社交)无法达到的水平。爆发式放电的产生涉及到NMDAR的激活；NMDAR的激活需要谷氨酸和甘氨酸。AMPA受体(AMPAR)的激活和星形胶质细胞的去极化可以在黑质中释放甘氨酸。因此，可能调节NMDAR激活的一个机制是通过AMPAR激活从星形胶质细胞释放甘氨酸。这项建议的目的是研究一种新的机制，通过激活AMPAR从中脑星形胶质细胞释放甘氨酸来调节多巴胺能神经元的突发性放电。首先，体外生理学将被用来确定控制细胞外甘氨酸浓度是否会导致突发放电的改变。爆炸射击将会被激发 通过电刺激谷氨酸能传入。在药物升高和降低胞外甘氨酸浓度前后，将测量爆发放电和NMDAR介导的电流。其次，星形胶质细胞是否可以调节DA神经元的突发性放电，将利用光遗传学进行测试。这将通过对星形细胞膜电位的光遗传操作来实现。我们将比较电刺激与星形胶质细胞光激活配对的电刺激和NMDAR介导的电流。最后，将测试星形胶质细胞上的AMPAR是否可以调节爆发式放电。这将通过在添加特定的AMPAR阻滞剂前后首先测量DA神经元周围的星形胶质细胞的电诱发电流来完成。下一步，电刺激将诱发爆发放电，然后加入AMPAR阻滞剂；比较两种情况下爆发和NMDAR介导的电流。接下来，在AMPAR阻滞剂存在的情况下，电刺激将与星形胶质细胞的光激活配对；猝发电流和NMDAR介导的电流将被比较。该提议将使人们更好地理解DA神经元爆发式放电背后的细胞机制。

 DESCRIPTION (provided by applicant): Dopamine (DA) neurons located in the substantia nigra pars compacta and the ventral tegmental area generate signals involved in motivated behavior. DA neuron firing patterns encode different aspects of reward. Drugs of abuse, such as cocaine, usurp the dopaminergic system causing an increase in dopaminergic burst firing which increases dopamine release at target regions to levels unattainable by natural reward (e.g. eating, grooming, social interactions). Generation of burst firing involves activation of NMDA receptors (NMDARs); activation of NMDARs requires glutamate and glycine. Activation of AMPA receptors (AMPARs) and depolarization of astrocytes are known to release glycine in the substantia nigra. Thus, one mechanism that may regulate NMDAR activation is glycine release from astrocytes via AMPAR activation. The goal of this proposal is to study a novel mechanism for regulating burst firing of dopaminergic neurons by glycine release from midbrain astrocytes via activation of AMPARs. First, in vitro physiology will be used to determine if manipulating the extracellular concentration of glycine causes a change in burst firing. Burst firing will be evoked by electrical stimulation of glutamatergic afferents. Burst firing and NMDAR- mediated currents will be measured before and after pharmacologically increasing and decreasing the extracellular glycine concentration. Second, whether astrocytes can regulate DA neuron burst firing will be tested using optogenetics. This will be accomplished by optogenetic manipulation of astrocytic membrane potentials. Burst firing and NMDAR-mediated currents will be compared between electrical stimulation, and electrical stimulation paired with photoactivation of astrocytes. Lastl, whether AMPARs on astrocytes can modulate burst firing will be tested. This will be done by first measuring electrically evoked currents from astrocytes surrounding DA neurons before and after addition of a specific AMPAR blockers. Next, burst firing will be evoked by electrical stimulation, then AMPAR blockers will be added; burst and NMDAR-mediated currents will be compared between the two conditions. Next, in the presence of AMPAR blockers, electrical stimulation will be paired with photoactivation of astrocytes; burst and NMDAR-mediated currents will be compared. The proposal will grant a better understanding of the cellular mechanism that underlies burst firing in DA neurons.