Astrocytic mechanism for regulating dopaminergic burst firing

 描述（由申请人提供）：位于黑质丘脑部和腹侧被盖区的多巴胺（DA）神经元产生涉及动机行为的信号。DA神经元放电模式编码不同方面的奖励。滥用药物（如可卡因）会侵占多巴胺能系统，导致多巴胺能爆发放电增加，从而使目标区域的多巴胺释放增加到自然奖励（如进食、梳理、社交互动）无法达到的水平。爆发放电的产生涉及NMDA受体（NMDAR）的激活; NMDAR的激活需要谷氨酸和甘氨酸。已知AMPA受体（AMPAR）的激活和星形胶质细胞的去极化在黑质中释放甘氨酸。因此，可以调节NMDAR活化的一种机制是经由AMPAR活化从星形胶质细胞释放甘氨酸。 本研究的目的是通过激活AMPAR来研究中脑星形胶质细胞释放甘氨酸调节多巴胺能神经元爆发性放电的新机制。首先，将使用体外生理学来确定操纵甘氨酸的细胞外浓度是否引起爆发放电的变化。将诱发爆发性放电 通过电刺激多巴胺能传入神经。在增加和降低细胞外甘氨酸浓度之前和之后测量爆发放电和NMDAR介导的电流。其次，星形胶质细胞是否可以调节DA神经元爆发放电将使用光遗传学进行测试。这将通过星形胶质细胞膜电位的光遗传学操纵来实现。将在电刺激和与星形胶质细胞的光活化配对的电刺激之间比较突发放电和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AMPAR阻断剂之前和之后首先测量来自DA神经元周围的星形胶质细胞的电诱发电流来完成。接下来，将通过电刺激诱发爆发放电，然后添加AMPAR阻断剂;将在两种条件下比较爆发和NMDAR介导的电流。接下来，在AMPAR阻断剂的存在下，电刺激将与星形胶质细胞的光活化配对;将比较爆发和NMDAR介导的电流。该提议将使我们更好地理解DA神经元爆发性放电的细胞机制。

 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.