In-vivo Modulation of Synapses by Endocannabinoids

描述(申请人提供)：拟议研究的长期目标是研究内源性大麻素对中枢神经系统突触传递的调制作用。实验模型包括第八神经听觉初级传入与金鱼Mauthner(M-)细胞和邻近的(GABA/Glyine)抑制终末之间已识别的混合电和化学(谷氨酸)突触。虽然大多数描述内源性大麻素在突触传递中的作用的研究都是利用体外系统，但这种制备方法独特地允许连续监测和量化体内电和化学传递的变化。到目前为止，已报道内源性大麻素通过突触前激活大麻素1型受体(CB1Rs)来抑制化学突触传递。与这一概念相反，我们的初步结果表明，CB1Rs的激活增强了M细胞上这些输入的突触传递。树突内记录、分子生物学技术和免疫细胞化学将被用来测试这种激动剂诱导的突触传递改变的特定假说和机制。目的1、探讨不同的大麻素激动剂和内源性大麻素对混合突触和抑制性终末突触效应的影响。它基于的数据表明，CB1R的激活导致混合突触的电和化学传递的长期增强。这些变化还包括附近的抑制终末。我将探索局部应用的大麻素激动剂和内源性大麻素对与这种听觉输入功能相关的单一和群体突触反应和膜电导的作用。目的2是研究这些突触传递的长期变化的机制。这是基于多巴胺受体拮抗剂阻断CB1R激活所触发的增强的发现。我们以前已经报道过多巴胺能神经支配的存在和多巴胺的应用引起突触反应的持久增强。我们将检验这一假说，即大麻素诱发的增强是通过邻近曲张的多巴胺释放来调节的。我们还将询问在哪种生理条件下以及从哪种特定细胞类型释放内源性大麻素。这项拟议的研究提出了这样的概念，即内源性大麻素对细胞间通讯的调制不仅限于化学突触，还包括缝隙连接介导的电突触。此外，基于与多巴胺能系统的功能相互作用，它可以导致突触反应的长期增强。这种调节作用可能是一种广泛的性质，不仅与基底节、视网膜和新皮质等两种传播形式共存的结构中的正常大脑功能有关，而且与许多与健康有关的问题，如药物滥用。

DESCRIPTION (provided by applicant): The long-term objective of the proposed research is to study the modulatory actions of endocannabinoids on synaptic transmission in the CNS. The experimental model involves identified mixed electrical and chemical (glutamatergic) synapses between eighth nerve auditory primary afferents and the goldfish Mauthner (M-) cell and neighboring (GABA/Glycine) inhibitory terminals. While most studies describing the role of endocannabinoids on synaptic transmission have utilized in-vitro systems, this preparation uniquely allows continuous monitoring and quantification of changes in electrical and chemical transmission in-vivo. So far, endocannabinoids have been reported to depress chemical synaptic transmission via presynaptic activation of cannabinoid type 1 receptors (CB1Rs). Contrasting this notion, our preliminary results show that activation of CB1Rs enhances synaptic transmission at these inputs on the M-cell. Intradendritic recordings, molecular biology techniques, and immunocytochemistry, will be used to test specific hypotheses and mechanisms underlying modifications of synaptic transmission induced by this agonist. Aim 1, explores the action of different cannabinoid agonists and endocannabinoids on the synaptic efficacy of mixed synapses and inhibitory terminals. It is based on data suggesting that activation of CB1R leads to long-lasting enhancement of both electrical and chemical transmission at mixed synapses. These changes also included nearby inhibitory terminals. I will explore the actions of locally applied cannabinoid agonists and endocannabinoids on unitary and population synaptic responses and membrane conductances that are relevant for the function of this auditory input. Aim 2 is to investigate the mechanisms underlying these long-term changes in synaptic transmission. It is based on the finding that dopamine receptor antagonists block the potentiation triggered by CB1R activation. We have previously reported the presence of a dopaminergic innervation and application of dopamine evoked lasting enhancements of the synaptic response. We will test the hypothesis that cannabinoid-evoked potentiation is mediated via dopamine release from neighboring varicosities. We will also ask under which physiological conditions and from which particular cell type endocannabinoids are released. The proposed research addresses the concept that modulation of intercellular communication by endocannabinoids is not restricted to chemical synapses but also include gap-junction mediated electrical synapses. Moreover, based on a functional interaction with the dopaminergic system, it can lead to long-term potentiation of synaptic responses. This modulatory action could constitute a widespread property, relevant not only to normal brain function in structures such as the basal ganglia, retina, and neocortex where both forms of transmission co-exist, but also to numerous health-related issues such as drug abuse.