Plasticity of bridge collaterals in Parkinonian state and treatment

 描述(申请人提供)：在帕金森病(PD)中，多巴胺能神经元的退化会导致运动缓慢、僵硬、震颤和步态异常等症状。多巴胺(DA)前体左旋多巴(L-多巴)可显著缓解运动症状，但在长期治疗后会出现L-多巴诱发的运动障碍(LID)等副作用，从而限制了其治疗应用。纹状体中棘神经元(MSN)是帕金森病患者运动障碍和LID过度运动的关键介质，它是纹状体的唯一输出，在正常情况下接受DA输入。帕金森病缺乏DA输入，慢性L-多巴中毒后DA输入过多、非生理性，导致MSN功能异常，分别导致运动迟缓和LID。经典的基底节(BG)模型认为，MSN可分为两条独立的通路：1)D1MSN，表达多巴胺D1受体(D1Rs)，形成“直接通路”，仅靶向BG的输出核团；(Ii)D2-MSN，表达D2受体(D2Rs)，形成“间接通路”，通过苍白球(GPE)间接靶向BG输出核团。然而，随后的解剖学研究发现，来自D1-MSN的轴突形成侧支，这些侧支也投射到GPE。因此，这些“桥接络脉”允许D1-MSN影响直接和间接途径的靶点。我们最近发现，这些桥接侧支的终末密度是高度可塑性的，并随着DA信号的变化而变化。D-1MSN桥联侧支因慢性D2R阻断而减少，但因D2R过度表达而几乎加倍。重要的是，桥接侧支密度的变化显著改变了d1-msn对运动输出的影响。虽然D1MSN的光遗传刺激通常会增加运动行为，但通过D2R过度表达增加桥接侧支会导致同样的刺激抑制运动行为。通过慢性D2R阻断使桥接侧支末端密度正常化，这种运动抑制被逆转为运动激活。本文提出的实验将验证这一假说，即PD和LID中异常的DA信号也会改变桥接侧支。我们将使用在Drd1a启动子的控制下表达绿色荧光蛋白(GFP)的转基因小鼠，这使得D1-MSN的轴突投射可以通过针对GFP的免疫组织化学来可视化。因此，D1MSN桥联侧支的终末密度可以通过GFP在GPE中的染色强度来确定。用6-羟基多巴胺去神经将被用来诱导偏侧帕金森综合征。肢体使用偏差将被用来评估行动不便，而慢性L-多巴治疗将会导致LID。我们认为，DA耗竭将逐渐降低桥接侧支末端密度，并与D1R激动剂逆转运动障碍的渐进能力平行。此外，我们认为，慢性L-多巴治疗导致盖子严重程度的增加，将与桥接侧支末端密度的进一步变化有关。帕金森病桥接侧支异常改变的发现将证明未来逆转这些改变的治疗方法的发展是合理的。

 DESCRIPTION (provided by applicant): In Parkinson's disease (PD), degeneration of dopaminergic neurons leads to symptoms including slowness of movement, rigidity, tremor, and gait abnormality. The dopamine (DA) precursor levodopa (L-DOPA) dramatically alleviates motor symptoms, but can cause side effects such as L-DOPA-induced dyskinesia (LID) after prolonged treatment, thereby limiting its therapeutic use. A critical mediator of motor deficit in PD and excessive movement in LID is the striatal medium spiny neurons (MSNs), which form the sole output of the striatum and receive DA input under normal conditions. The lack of DA input in PD, and the excessive, non-physiological DA input after chronic L-DOPA leads to abnormal MSN function and contribute to slowness of movement and LID respectively. The classical model of basal ganglia (BG) proposes that MSNs can be divided into two segregated pathways: i) D1-MSNs, which express dopamine D1 receptors (D1Rs) and form the "direct pathway", targeting only the output nuclei of BG; ii) D2-MSNs, which express D2 receptors (D2Rs) and form the "indirect pathway", targeting the output nuclei of BG indirectly via the external globus pallidus (GPe). However, subsequent anatomical studies found that axons from D1-MSNs form collaterals that also project to the GPe. These "bridging collaterals" thus allow D1-MSNs to influence the targets of both direct and indirect pathway. We recently found that the terminal density of these bridging collaterals is highly plastic and changes in response to alterations in DA signaling. D-1MSN bridging collateral is reduced by chronic D2R blockade but is almost doubled by D2R overexpression. Importantly, changes in bridging collateral density dramatically alter D1-MSN's influence on motor output. While optogenetic stimulation of D1-MSNs normally increases locomotor behavior, increasing bridging collaterals by D2R overexpression caused the same stimulation to inhibit locomotor behavior. This motor inhibition was reversed back to motor activation by normalizing bridging collateral terminal density via chronic D2R blockade. Experiments proposed here will test the hypothesis that bridging collaterals are also altered by abnormal DA signaling in PD and LID. We will use transgenic mice that express green fluorescent protein (GFP) under the control of the Drd1a promoter, which allows the axonal projections of D1-MSNs to be visualized by immunohistochemistry against GFP. The terminal density of D1- MSN bridging collateral can therefore be determined by GFP staining intensity in the GPe. DA denervation by 6-hydroxydopamine will be used to induce hemi-parkinsonism. Limb-use bias will be used to assess akinesia, and LID will be induced by chronic L-DOPA treatment. We propose that DA depletion will progressively reduce bridging collateral terminal density, and that it would parallel the progressive ability of a D1R agonist to reverse akinesia. Furthermore, we propose that chronic L-DOPA treatment, which causes increases in LID severity, will be associated with further alterations in the terminal density of bridging collaterals. The identification of abnormal bridging collateral changes in PD will justify future development of therapy to reverse these changes.