Sex-dependent expression and utilization of spinal mu- and kappa-opioid systems

描述（由申请方提供）：定义阿片类镇痛作用的性别依赖性的既往研究尚未准备好揭示相关的机制基础和生物学底物。本申请提出建立脊髓阿片类抗伤害感受的性别依赖性药理学特征与（1）脊髓阿片类受体的性别二态性性质和（2）通过鞘内（i.t.）吗啡假设的性二态关系在脊柱？-阿片受体（莫尔）、内吗啡肽2（EM 2）、？阿片受体（KOR）和强啡肽1-17（Dyn）的研究得到了大量初步数据的支持，这得到了我们以前的证明的支持，即：在雄性大鼠中，吗啡主要引起MOR偶联的脊髓镇痛作用，而在雌性大鼠中，i.t.吗啡募集一个更包容的整合系统，其需要脊髓莫尔和KOR的伴随活性。脊髓莫尔剪接变体和莫尔KOR异二聚体的性二态表达水平被假定为女性对脊髓吗啡的表型反应的分子基础。与脊髓阿片受体的性二态组织假设平行，我们假设，脊髓阿片类药物的释放和镇痛反应的性别依赖性调节是性别依赖性脊髓阿片类镇痛作用的组成部分。具体而言，目标1将检验由i.t.吗啡需要从雄性而不是雌性的脊髓释放EM 2。目标2将检验i.t.与雌性相比，EM 2在雄性中引起更大的抗伤害感受，这加剧了通过i.t.吗啡我们推测，这是介导的，至少部分，由性别依赖性表达的脊髓莫尔剪接变异体。通过i.t.吗啡是其在雄性而不是雌性中产生的脊髓抗伤害感受的先决条件，这不能完全揭示脊髓吗啡抗伤害感受的性别依赖性特征。要做到这一点，需要了解i.t.吗啡在女性因此，目标3将检验这一假设，即在女性中，i.t.吗啡镇痛是通过释放Dyn而不是EM 2来介导的。目的4将检验以下假设，即在雌性中需要脊髓莫尔和KOR的伴随激活以使脊髓吗啡抗伤害感受明显，这反映了与雄性相比，在雌性脊髓中存在显著更高表达水平的莫尔KOR异二聚体。我们将采用多维度的方法来实现这些相互关联的目标，该方法整合了行为，药理学，分子和基于免疫的生化水平的分析。这些将被用作交叉验证调查结果的补充措施。由于疼痛和成瘾具有共同的底物，预期结果将揭示新的药物靶点，这些靶点不仅与优化男性和女性的疼痛管理有关，而且与以性别依赖的方式治疗阿片类药物戒断和复发有关。

DESCRIPTION (provided by applicant): Previous studies that defined sex dependency of opioid antinociception were not poised to reveal relevant mechanistic underpinnings and biological substrates. The present application proposes to establish a causal association between sex-dependent pharmacological characteristics of spinal opioid antinociception and (1) the sexual dimorphic nature of spinal opioid receptors and (2) the sex-dependent release of spinal opioids by intrathecal (i.t.) morphine. Hypothesized sexual dimorphic relationships among spinal ?-opioid receptors (MOR), endomorphin 2 (EM2), ?-opioid receptors (KOR) and dynorphin 1-17 (Dyn) are supported by substantial preliminary data, which is undergirded by our previous demonstrations that i.t. morphine elicits predominantly MOR-coupled spinal antinociception in males, whereas in females i.t. morphine recruits a more inclusive integrative system that requires concomitant activity of spinal MOR and KOR. Sexual dimorphic expression levels of spinal MOR splice variants and MOR KOR heterodimers are postulated to be molecular underpinnings of the female phenotypic response to spinal morphine. In parallel with the postulated sexually dimorphic organization of spinal opioid receptors, we hypothesize that the sex-dependent regulation of the release of and analgesic responsiveness to spinal opioids are integral component of sex-dependent spinal opioid antinociception. Specifically, Aim 1 will test the hypothesis that the antinociception produced by i.t. morphine, requires the release of EM2 from the spinal cord of males, but not females. Aim 2 will test the hypothesis that i.t. EM2 elicits greater antinociception in males vs. females, which exacerbates the consequences of the sex-dependent release of spinal EM2 by i.t. morphine. We postulate that this is mediated, at lest in part, by the sex-dependent expression of spinal MOR splice variants. Validation that release of spinal EM2 by i.t. morphine is a prerequisite for the spinal antinociception it produces in males, but not females, would not fully reveal the sex-dependent landscape that underlies spinal morphine antinociception. To do so requires knowledge of the analgesic substrates recruited by i.t. morphine in females. Accordingly, Aim 3 will test the hypothesis that in females, i.t. morphine analgesia is mediated via the release of Dyn, not EM2. Aim 4 will test the hypothesis that the requirement in females for the concomitant activation of spinal MOR and KOR for spinal morphine antinociception to be manifest reflects the presence of significantly higher expression levels of MOR KOR heterodimers in spinal cord of females vs. males. We will pursue these interrelated Aims using a multi-dimensional approach that integrates behavioral, pharmacological, molecular and immuno-based biochemical levels of analyses. These will be used as complementary measures to cross validate findings. Since pain and addiction share common substrates, expected outcomes will reveal novel drug targets that are not only relevant to optimizing pain management in men and women but also to treating opioid withdrawal and relapse in a sex-dependent fashion.