Developing focal RNA interference for the identification of neural entry points for sleep-promoting adenosine

腺苷通过激活A_2a和gt；受体来促进睡眠。A&lt；2A&gt；受体密集表达于基底节纹状体神经元，多巴胺D_2受体与A&lt；2A&gt；受体共表达，参与运动功能、习惯形成和奖赏/成瘾行为，所有这些活动都需要清醒。在帕金森氏症等多巴胺含量较低的情况下，保持觉醒的能力会受到影响，但基底节中的A&lt；2a和gt；受体在多大程度上对睡眠和觉醒的调节作用尚不清楚。我们使用强大的定点基因操作工具，包括基于Cre/lox技术的条件性A&lt；2A&gt；受体敲除小鼠；通过局部感染携带A&lt；2A&gt；受体短发夹状RNA的腺相关病毒局部感染大鼠；以及通过基因工程受体-通道系统的体内刺激和抑制来调节神经元活动，例如，光学开关，包括通道视紫红质，以及由设计药物(DREADD)专门激活的设计受体。我们的研究表明，咖啡因的唤醒作用主要依赖于伏隔核壳上神经元上的A&lt；2a和gt；受体，这些神经元的瞬时激活促进睡眠。这些观察表明，伏核中的A&lt；2a和gt；受体是控制睡眠和觉醒的关键结构元件。腹侧纹状体具有整合行为功能的独特能力，因此，它是需要意识的过程调节睡眠和觉醒的理想场所。我们认为，动机可以被认为是调节睡眠和清醒的一个重要的基本原则。

Adenosine promotes sleep through the activation of A_<2A> receptors. A_<2A> receptors are densely expressed on striatopallidal neurons of the basal ganglia, where dopamine D_2 receptors are co-expressed with A_<2A> receptors and involved in motor function, habit formation, and reward/addictive behaviors, all activities that require wakefulness. Abilities to maintain arousal are compromised under low dopamine conditions such as Parkinson's disease, but the extent to which A_<2A> receptors in the basal ganglia contribute to the regulation of sleep and wakefulness is not known. We investigated the role of A_<2A> receptors in the basal ganglia for wakeful consciousness by using powerful tools for site-specific gene manipulations, including conditional A_<2A> receptor knockout mice based on the Cre/lox technology ; focal A_<2A> receptor knockdown in rats through the local infection with adeno-associated virus carrying short-hairpin RNA of A_<2A> receptors ; and modulation of neuronal activity through in-vivo stimulation and inhibition with genetically engineeredreceptor-channel systems, e. g., optical switches, including channelrhodopsin, and designer receptors exclusively activated by a designer drug(DREADD). Our studies have revealed that the arousal effect of caffeine critically depends on A_<2A> receptors on neurons in the shell of the nucleus accumben and that transient activation of these neurons promotes sleep.These observations imply that A_<2A> receptors in the nucleus accumbens are key structural elements for the control of sleep and wakefulness. The ventral striatum has the unique capability to integrate behavioral functions and thus, it is an ideal site where sleep and wakefulness are regulated by processes that require consciousness. We believe that motivation may be considered as an important fundamental principle by which sleep and waking are regulated.