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

描述（由申请人提供）：先前定义阿片类药物抗痛觉性依赖的研究并未准备好揭示相关的机制基础和生物学基础。本研究旨在建立脊髓阿片抗痛感药物的性别依赖性药理特征与(1)脊髓阿片受体的性别二型性性质和(2)鞘内吗啡对脊髓阿片的性别依赖性释放之间的因果关系。脊髓间两性二形关系的假设？-阿片受体（MOR），内啡肽2 (EM2), ？大量的初步数据支持了-阿片样受体（KOR）和肌啡肽1-17 （Dyn）的作用，我们之前的研究表明，在男性中，吗啡主要引起dor偶联的脊髓抗痛觉作用，而在女性中，吗啡招募了一个更广泛的综合系统，需要脊髓MOR和KOR的同时活动。脊髓MOR剪接变异体和MOR KOR异源二聚体的两性二态表达水平被认为是女性对脊髓吗啡表型反应的分子基础。与假定的脊髓阿片受体的性别二形组织平行，我们假设性别依赖的脊髓阿片释放和镇痛反应的调节是性别依赖的脊髓阿片抗痛觉的组成部分。具体来说，Aim 1将测试由吗啡产生的抗性感觉需要从雄性脊髓释放EM2，而不是雌性脊髓的假设。目的2将验证一种假设，即相对于雌性而言，雌性吗啡对脊髓EM2的性别依赖性释放会导致男性更强的抗性感觉，这加剧了吗啡对脊髓EM2的性别依赖性释放的后果。我们假设这至少在一定程度上是由脊柱MOR剪接变体的性别依赖性表达介导的。证实吗啡释放脊髓EM2是男性产生脊髓抗痛觉的先决条件，而不是女性，并不能完全揭示脊髓吗啡抗痛觉的性别依赖性。要做到这一点，需要了解吗啡在雌性体内的镇痛底物。因此，Aim 3将检验假设，在女性中，吗啡镇痛是通过Dyn的释放介导的，而不是通过EM2。目的4将检验这样一个假设，即女性脊髓中MOR和KOR同时激活对脊髓吗啡抗痛性的要求是明显的，这反映了女性脊髓中MOR和KOR异源二聚体的表达水平明显高于男性。我们将采用多维度的方法，整合行为、药理学、分子和基于免疫的生化水平分析，来追求这些相互关联的目标。这些将用作交叉验证发现的补充措施。由于疼痛和成瘾有共同的底物，预期的结果将揭示新的药物靶点，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.