Mechanisms of postnatal cutaneous afferent development during inflammation

描述（由申请人提供）：许多儿科疾病和对儿童进行的外科手术导致疼痛状态，但众所周知，目前的疼痛药物治疗对儿童有不良影响，如轻度恶心或呕吐，但也有更严重的副作用，如呼吸抑制，甚至死亡。因此，为了开发更合适的治疗儿童疼痛，更好地了解如何特定的感觉传入人群可能有助于儿科疼痛状态是至关重要的。虽然对成人外周损伤后皮肤伤害感受器的功能特性和可塑性了解很多，但对发育过程中感觉传入的功能特性和外周损伤后致敏机制知之甚少。这项建议的主要目标是在功能上表征皮肤感觉传入在整个发展过程中在幼稚小鼠和产后炎症模型，并开始，以确定一些与感觉传入的变化所涉及的机制。为了确定这一点，我们开发了一种离体毛状皮肤，隐神经，背根神经节（DRG），脊髓记录制备新生儿/出生后的小鼠，使我们能够全面表型感觉纤维之前和之后的皮肤损伤。在具体目标1中，我们将首先分析这些传入的功能，解剖和神经化学表型在不同的时间在出生后的发展，使用离体记录和免疫细胞化学分析的组合。然后，功能的变化将与被认为参与DRG中感觉功能的各种受体/通道的mRNA和蛋白质表达的改变相关。接下来，在具体目标2中，我们将进行与SA 1中所述类似的研究，不同之处在于我们将分析出生后发育期间不同时间毛状皮肤注射角叉菜胶后感觉传入的综合表型，并将这些数据与DRG中基因表达的变化相关联，以确定观察到的皮肤传入功能变化的潜在机制。这些实验将使我们能够表征所有类型的皮肤感觉神经元在整个发育过程中和外周炎症期间的变化，并可能确定与发育中的感觉神经元如何对损伤做出反应相关的独特年龄依赖性机制。这些研究将建立未来实验的基础，使用体内siRNA介导的单个外周神经中的基因敲除结合离体记录制备物，其中我们将能够分析在正常（未受伤）发育期间和在出生后皮肤炎症的不同时间期间特定受体/通道的变化的作用。这些和未来的研究将使我们更好地了解感觉纤维的变化如何影响儿童疼痛，也可能导致建立更合适的儿童疼痛治疗方法。

DESCRIPTION (provided by applicant): Numerous pediatric disorders and surgical procedures performed on children lead to pain states yet it is well known that current pharmacological treatments for pain have adverse effects in children such as mild nausea or vomiting but also more severe side effects like respiratory depression and even death. Thus in order to develop more appropriate therapies for pain in children, a better understanding of how particular sensory afferent populations may contribute to pediatric pain states is of utmost importance. While much is known about the functional properties and plasticity of cutaneous nociceptors following peripheral injuries in adults, relatively little is known about the functiona properties of sensory afferents during development and the mechanisms of sensitization after peripheral injury. The main goal of this proposal is to functionally characterize cutaneous sensory afferents throughout development in naive mice and in models of postnatal inflammation, and begin to determine some of the mechanisms involved with the changes in sensory afferents. In order to determine this, we developed an ex vivo hairy skin, saphenous nerve, dorsal root ganglion (DRG), spinal cord recording preparation in neonatal/ postnatal mice that enables us to comprehensively phenotype sensory fibers before and after cutaneous injury. In Specific Aim 1, we will first analyze the functional, anatomical, and neurochemical phenotypes of these afferents at various times during postnatal development using combinations of ex vivo recording and immunocytochemical analyses. Changes in function will then be correlated with alterations in mRNA and protein expression of various receptors/ channels thought to be involved in sensory function in the DRGs. Next, in Specific Aim 2, we will perform similar studies as described in SA1 except we will analyze the comprehensive phenotypes of sensory afferents after hairy skin injection of carrageenan at different times during postnatal development and correlate these data with changes in gene expression in the DRGs to determine the potential mechanisms of the observed functional changes in cutaneous afferents. These experiments will enable us to characterize the changes in all types of cutaneous sensory neurons throughout development and during peripheral inflammation, and potentially identify unique age-dependent mechanisms associated with how developing sensory neurons respond to injury. These studies will establish the foundation of future experimentation using in vivo siRNA- mediated knockdown of genes in single peripheral nerves in conjunction with ex vivo recording preparations where we will be able to analyze the roles of changes in specific receptors/ channels during normal (uninjured) development and during different times of postnatal cutaneous inflammation. These and future studies will allow us to better understand how changes in sensory fibers impact pediatric pain and may also lead to the establishment of more suitable treatments for pain in children.