NEUROCHEMICALS IN HYPOTHALAMIC SIGNALING AFTER COITUS

在雌性新西兰白兔身上，交配会导致大量的释放 下丘脑促性腺激素释放激素(GnRH)触发 黄体生成素(LH)激增，从而导致排卵。性交 由外周神经通过脊髓传递的信号不会 突触直接分布在GnRH神经元上。相反，这些信号通过 分泌刺激GnRH释放的中间神经元。的性质，以及 这些中间神经元之间的相互作用尚不完全清楚。 此外，男性的促黄体生成素和雄激素水平也有所增加， 但性交活动是否会引起男性的神经内分泌反应 下丘脑与女性的相似之处尚不清楚。我们的假设是 性交引起去甲肾上腺素(NE)和/或神经肽Y的变化 (NPY)基因表达、分泌和突触传递导致 雄性和雌性兔下丘脑GnRH的戏剧性释放。 具体目的1是找出性交是否激活GnRH/NPY/NE分泌 在雄性和去卵巢的雌性兔中，雌二醇-17β处理； 不间断的下丘脑灌流(即微透析和推拉 灌注)和静脉血样将从远程采集 拴住的、自由活动的兔子，每隔2.5至10分钟在5小时前和 与异性交配后4h。浓缩物 灌流液中的GnRH、NPY和NE以及血液中的黄体生成素和性腺激素 用特异性放射免疫分析(RLA)和高效液体测定 高效液相色谱法(去甲肾上腺素)。目标2是观察下丘脑是否释放出 NE和/或NPY介导性交诱导的GnRH变化；哌唑嗪是一种α-受体激动剂。 肾上腺素能拮抗剂和抗NPY血清将用于阻断 去甲肾上腺素和神经肽Y在中枢神经系统的兴奋作用 性交诱发的GnRH峰。目标3是检查封锁是否 下丘脑神经末梢突触前NE转运蛋白可以(A) 通过增加局部去甲肾上腺素和(B)促性腺激素释放激素 随后降低下丘脑去甲肾上腺素，从而释放促性腺激素释放激素 酪氨酸羟化酶(TH)基因表达的调节；急性， 下丘脑内注射地塞帕明(DMI)，一种特异性阻滞剂 一种运输器，将被给予动物，其中一些将被预先- 全身用DMI或生理盐水治疗(每天两次，连续14天)。效应 急性和慢性DMI治疗的评估将分别通过以下方式进行 测定下丘脑NE/GnRH释放(用高效液相色谱法和RIA法)以及TH和TH 蓝斑和外侧核胞体NPY mRNA的含量 脑干被盖(通过核糖核酸酶保护试验) 接受类固醇治疗的女性。目标4是评估(A)脑干是否以及何时 TH和NPY信使RNA的变化发生在性交后和(B)如果 下丘脑DMI长时间灌注抑制TH和NPY基因的表达 脑干去甲肾上腺素神经元的表达；脑干基因表达的改变 神经元将在与NE和NPY变化平行的时间进行测量 分泌物。克隆了兔TH特异性探针和人NPY探针 已经在我们实验室的兔子身上得到了验证。这些发现 将是第一个解决神经转运体分子在 繁殖。研究的结果应该会增加对 影响正常内分泌过程的神经因素和 与性相关的神经回路故障有关 男性和女性的精神障碍。

In the female New Zealand White rabbit, coitus induces a massive release of hypothalamic gonadotropin-releasing hormone (GnRH) that triggers the surge of luteinizing hormone (LH), and therefore ovulation. The coital signals, relayed by peripheral nerves through the spinal cord, do not synapse directly on GnRH neurons. Instead, these signals are transduced by interneurons whose secretions stimulate GnRH release. The nature of, and interactions between these interneurons are not fully understood. Furthermore, increases of LH and androgens in males have been suggested, but whether coital activities induce neuroendocrine responses in the male hypothalamus similar to those in the female are unknown. Our hypothesis is that coitus provokes changes in norepinephrine (NE) and/or neuropeptide Y (NPY) gene expression, secretion and synaptic transmission that result in a dramatic release of hypothalamic GnRH in both male and female rabbits. Specific aim 1 is to find out if coitus activates GnRH/NPY/NE secretions in male and Ovariectomized, estradiol-17beta-treated female rabbits; uninterrupted hypothalamic perfusion (i.e., microdialysis and push-pull perfusion) and venous blood samples will be collected remotely from tethered, freely moving rabbits at 2.5- to 10-min intervals 5 h before and 4 h after mating with a partner of the opposite sex. Concentrations of GnRH, NPY and NE in perfusate and LH and gonadal steroids in blood will be measured by specific radioimmunoassays (RlA) and high-performance liquid chromatography (HPLC, for NE). Aim 2 is to see if hypothalamic releases of NE and/or NPY mediate coitally-induced GnRH changes; prazosin, an alpha- adrenergic antagonist, and anti-NPY serum will be used to block the stimulatory action of NE and NPY, respectively, during initiation of the coitally-induced GnRH surge. Aim 3 is to examine if blockade of presynaptic NE transporter protein at hypothalamic nerve terminals can (a) initially enhance GnRH secretion by increasing local NE and (b) subsequently reduce hypothalamic NE, and therefore GnRH, release by down- regulation of tyrosine hydroxylase (TH) gene expression; acute, intrahypothalamic treatment with desipramine (DMI), a specific blocker for NE transporter, will be given to animals, some of which will be pre- treated systemically with DMI or saline (twice-daily for 14 days). Effects of acute and chronic DMI treatment will be assessed, respectively, by measuring NE/GnRH release in the hypothalamus (by HPLC and RIA) and TH and NPY mRNA content in cell bodies located in locus coeruleus and lateral tegmentum of the brainstem (by RNase protection assay) in either male or steroid-treated females. Aim 4 is to assess (a) if and when brainstem changes in TH and NPY messenger RNAs occur after coitus and (b) if prolonged intrahypothalamic DMI infusion suppresses TH and NPY mRNA expression in brainstem NE neurons; altered gene expression in brainstem neurons will be measured at times parallel to changes in NE and NPY secretions. Specific rabbit TH probe has been cloned and human NPY probe has been validated for use in the rabbit in our laboratory. These findings will be the first to address the role of neurotransporter molecules in reproduction. The outcome of the studies should increase understanding of neural factors that influence normal endocrine processes and those associated with malfunctioning neural circuitry in sexually-related disorders in men and women.