Tetrahydrobiopterin and pain

两种四氢生物蝶呤（BH 4）合成的转录本 酶GTP环化水解酶1（GCH 1）和sepiapterin还原酶以及BH 4再循环酶醌式二氢蝶啶还原酶在外周神经损伤后在背根（DRG）神经元中上调。这种诱导与DRG中BH4的增加有关。BH4是必不可少的 产生5-羟色胺、去甲肾上腺素和多巴胺的芳香胺羟化酶和所有一氧化氮合酶的辅因子。神经损伤诱导的BH4增加可通过以下方式预防： 全身施用GCH 1抑制剂二氨基羟基嘧啶（DAHP）。DAHP在幼年动物中没有镇痛作用，但在具有外周神经损伤和炎症的啮齿动物中产生疼痛相关行为的显著减少。鞘内注射BH4本身会产生急性疼痛 超敏反应使用单核苷酸多态性的分析，我们还鉴定了人GCH 1中的单倍型，其在慢性背痛手术后的患者中具有疼痛保护作用，并且与健康受试者中疼痛敏感性降低相关。基于这些数据，我们假设BH4有助于神经性疼痛和炎性疼痛的启动和维持。本提案的目的是：1.研究疼痛相关啮齿动物模型中BH4诱导在DRG中发生的位置和时间以及何种刺激负责，2。选择性地表征GCH 1缺失、抑制或过表达的行为后果 在成年初级感觉神经元中，和3.识别BH4产生疼痛的感觉神经元机制。我们将研究响应于组织炎症和部分周围神经损伤（目的1）的DRG中BH 4合成和再循环酶的表达和活性变化的时间过程和细胞定位。为了阐明BH4在感觉神经元中的特定作用，我们将采用选择性过表达或在成年初级传入中缺失GCH 1的小鼠，以及过表达GCH 1的内源性抑制剂GFRP的小鼠，使用他莫昔芬诱导的DRG神经元特异性Cre重组酶技术（Aim 2）。nNOS的上调与 我们将测试BH4是否通过产生NO和钙内流的增加而产生疼痛。我们将使用DRG神经元的原代培养物来探索BH4对这些神经元的直接作用和下游效应器。该提案旨在探索负责疼痛的分子机制，并确定开发新镇痛药的新靶点。相关性：持续性疼痛是一个巨大的问题，因为患者经历的痛苦 以及对社会造成的高昂经济成本。由于目前的治疗通常无效或伴有不良副作用，因此需要更有效的镇痛药。这项补助金旨在了解 负责疼痛的机制，并验证一种特殊的酶，GCH 1作为开发新型镇痛药的目标。

Transcripts for two tetrahydrobiopterin (BH4) synthetic enzymes, GTP cyclohydrolase 1 (GCH1) and sepiapterin reductase, and a BH4 recycling enzyme quinoid dihydroypteridine reductase, are upregulated in dorsal root (DRG) neurons after peripheral nerve injury. This induction is associated with an increase in BH4 in the DRG. BH4 is an essential cofactor for the aromatic amine hydroxylases that produce 5- hyrdoxytryptamine, norepinephrine and dopamine, and for all nitric oxide synthases. The nerve injury-induced increase in BH4 is prevented by systemic administration of a GCH1 inhibitor, diamino hydroxypyrimidine (DAHP). DAHP has no analgesic effects in naove animals but produces a marked reduction in pain-related behavior in rodents with peripheral nerve lesions and inflammation. Intrathecal BH4 itself produces acute pain hypersensitivity. Using an analysis of single nucleotide polymorphisms we have also identified a haplotype in human GCH1 that is ¬pain protective¬ in patients after surgery for chronic back pain and associated with reduced pain sensitivity in healthy subjects. Based on these data we hypothesize that BH4 contributes to the initiation and maintenance of neuropathic and inflammatory pain. The aim of this proposal is to: 1. Study where and when BH4 induction occurs in the DRG in pain-related rodent models and what kinds of stimuli are responsible, 2. Characterize the behavioral consequences of deletion, inhibition or overexpression of GCH1 selectively in adult primary sensory neurons, and 3. Identify the mechanisms in sensory neurons by which BH4 produces pain. We will study the time course and cellular localization of changes in the expression and activity of BH4 synthetic and recycling enzymes in the DRG in response to tissue inflammation and partial peripheral nerve injury (Aim 1). To elucidate the specific role of BH4 in sensory neurons we will employ mice that selectively overexpress, or have a deletion of GCH1 in adult primary afferents, as well as mice that overexpress the endogenous inhibitor of GCH1, GFRP, using tamoxifen-inducible DRG neuron-specific Cre-recombinase technology (Aim 2). nNOS is upregulated in association with the increase in BH4 and we will test if BH4 produces pain by producing an increase in NO and calcium influx. We will use primary cultures of DRG neurons to explore the direct action of BH4 on these neurons and the downstream effectors responsible. The proposal is designed to explore the molecular mechanisms responsible for pain and identify novel targets for the development of new analgesics. Relevance: Persistent pain is an enormous problem due both to the suffering experienced by patients and the high economic cost to society. Because current therapy is often ineffective or associated withadverse side effects, more efficacious analgesics are required. This grant aims both to understand the mechanisms responsible for pain, and validate a particular enzyme, GCH1, as a target for developingnovel analgesics.