Anandamide Receptors: Role in Motor Neuron Protection

描述（由申请人提供）：在理解ALS的遗传基础以及介导散发性疾病进展的机制和分子途径方面正在取得重大进展;然而，有效的药物治疗仍然难以捉摸。ALS发展和进展的两个主要理论表明，运动神经元变性和死亡是由于对兴奋性毒性和氧化损伤的易感性。内描述的实验系统扩展了当前的ALS范例，提出兴奋性毒性和氧化损伤可能是由假定的调节控制的丧失引起的，并且刺激内源性大麻素/CB受体可能提供该调节的模拟物或替代物。内源性大麻素/CB受体系统有效地调节多巴胺能神经传递和毒性。我们已经在与运动控制相关的大脑区域中发现了大麻素/CB受体，包括它们在运动神经元中的存在。我们证明了大麻素/CB受体激动剂在脊髓培养中减弱兴奋性毒性。此外，CB受体激动剂是抗氧化剂，可以对两种假设的致病机制协同作用。这些研究已经成为可能，我们最近的克隆和表征的anandamide受体，内源性化合物的发现，通过这些受体（anandamides）的行为，并开发新的药理学工具，使我们能够监测的影响所产生的anandamide受体的激活。在第一个具体的目标，我们将确定，本地化和表征内源性大麻素/CB受体存在于小鼠脊髓。我们假设谷氨酸受体和花生四烯酸受体共同定位于小鼠脊髓运动神经元，提供了一个解剖学原理的神经保护作用，在体外和体内。在第二个具体的目标，我们将扩展我们的初步研究结果在脊髓文化中的兴奋性毒性衰减，并确定anandamide受体激动剂在调节神经毒性的作用，从小鼠脊髓运动神经元。我们将研究花生四烯酸/CB受体调制的影响，在两个范例产生神经毒性：兴奋性毒素的直接应用产生的兴奋性毒性;和氧化损伤产生的毒性。这些范例将在正常小鼠以及ALS模型小鼠（携带突变的人SOD 1基因的转基因小鼠）中进行。CBr受体在神经保护中的相对作用将通过检查CBr敲除小鼠中的激动剂作用来评估。提出的系统将使我们能够实验性地操纵复杂运动神经元环境的主要组成部分，从而测试ALS疾病发展的特定假设和可能的改善疗法。大麻素/CB受体系统提供了一种潜在的细胞机制，通过该机制，抗氧化剂和抗兴奋性毒性作用可以同时实现，具有更实质性治疗效果的可能性。

DESCRIPTION (provided by applicant): Significant advances are being made towards understanding the genetic basis for ALS as well as the mechanistic and molecular pathways mediating progression of the sporadic forms of the disease; however, effective pharmacotherapy remains elusive. Two of the primary theories for the development and progression of ALS suggests that motor neuron degeneration and death is due to susceptibility to excitotoxicity and oxidative damage. The experimental system described within extends current ALS paradigms, proposing that excitotoxicity and oxidative damage may result from presumed loss of modulatory control and that stimulating the endogenous anandamide/CB receptors may provide a mimic or substitution for that modulation. The endogenous anandamide/CB receptor system effectively modulates glutamatergic neurotransmission and toxicity. We have found anandamide/CB receptors in brain areas associated with motor control, including their presence in motor neurons. We demonstrate that anandamide/CB receptor agonists attenuate excitotoxicity in spinal cord cultures. In addition, CB receptor agonists are antioxidants that may work synergistically on both hypothesized pathogenic mechanisms. These studies have been made possible by our recent cloning and characterization of anandamide receptors, the discovery of endogenous compounds that act via these receptors (anandamides), and the development of novel pharmacological tools that allow us to monitor the effects produced by activation of anandamide receptors. In the first specific aim, we will identify, localize and characterize endogenous anandamide/CB receptors present in the mouse spinal cord. We hypothesize that both glutamate receptors and anandamide receptors co-localize in mouse spinal cord motor neurons, providing an anatomical rationale for neuroprotective effects in vitro and in vivo. In the second specific aim, we will extend our preliminary findings on attenuation of excitotoxicity in spinal cord cultures and define the role of anandamide receptor agonists in modulating neurotoxicity in motor neurons from mouse spinal cord. We will study the effects of anandamide/CB receptor modulation in two paradigms producing neurotoxicity: excitotoxicity produced by direct application of excitotoxins; and toxicity produced by oxidative damage. These paradigms will be conducted in normal mice as well as ALS model mice (transgenic mice carrying a mutant human SOD 1 gene). The relative role of the CB, receptor in neuroprotection will be evaluated by examining agonist effects in CBr knockout mice. The system proposed will allow us to experimentally manipulate major components of the complex motor neuron environment and thereby test specific hypotheses of ALS disease development and possible ameliorating therapies. The anandamide/CB receptor system offers a potential cellular mechanism by which both antioxidant and anti-excitotoxic effects might be achieved simultaneously, with the possibility of a more substantial therapeutic effect.