Neuroendocrine control of anterior pituitary hormones

描述（申请人提供）：生长激素（GH）促进蛋白质合成和脂肪分解。反过来，代谢紊乱与生长激素产生的改变有关，这有助于临床相关疾病的病理生理，如营养不良、神经性厌食症、肥胖和糖尿病。尽管代谢与生长激素之间有很强的联系，但对于代谢途径的扰动导致生长激素合成和释放变化的基本机制知之甚少。因此，这种应用将确定营养有效性变化的机制；1)调节下丘脑正常垂体GH分泌所必需的神经肽（GH-释放激素[GHRH]和生长抑素[SRIF]）的表达，2)改变垂体对GH刺激肽、GHRH和ghrelin的敏感性。有人提出，循环瘦素（一种脂肪细胞因子）和胃饥饿素（胃中产生的一种gh释放肽）的变化通过激活神经肽Y （NPY）神经元介导下丘脑GHRH和SRIF的表达。为了验证这一假设，禁食对瘦素合成（ob/ob）、瘦素组织来源（AZIP-F1）、NPY合成（NPY-/-）、SRIF合成（smst-/-）和SRIF信号传导（ss11 /2-/-）缺陷小鼠神经肽mRNA水平的影响将通过核糖核酸酶保护实验和原位杂交检测。此外，将测试在外源性激素替代（增加NPY、瘦素或胃饥饿素）、药物治疗（阻止内源性NPY产生）或被动免疫中和（阻止胃饥饿素的作用）后，正常小鼠禁食的效果。禁食不仅改变gh调节神经肽的表达，还通过增加GHRH- r和GHS-R mRNA水平，增强垂体对GHRH和ghrelin的敏感性。在体外，FFAs单独或与糖皮质激素联合使用可增加GHS-R的合成。因此，我们假设空腹诱导的FFA和糖皮质激素的升高是增强垂体受体合成和敏感性的必要条件，从而补偿空腹诱导的中枢信号的改变。为了验证这一假设，研究将检验在阻断FFA形成（通过抗脂溶性药物acpimox）或糖皮质激素作用（通过糖皮质激素受体拮抗剂RU-486）后，禁食改变垂体受体表达（通过定量RT-PCR）和动态GH释放（通过连续血液样本的RIA）的能力。此外，原代大鼠和非人灵长类动物（狒狒）垂体细胞培养将用于确定FFA和糖皮质激素是否通过转录或转录后过程介导其对受体合成的影响，以及FFA的作用是否可以通过激活假定的FFA核受体，过氧化物酶体增殖激活受体（PPAR）来模拟。这是RO1应用竞争性更新的第二次修订，该应用继续研究GH轴的调节。目前的应用主要集中在代谢功能变化与GH轴之间的相互关系。

DESCRIPTION (provided by applicant): Growth hormone (GH) promotes protein synthesis and lipolysis. In turn, metabolic disturbances are associated with alteration in GH production which contribute to the pathophysiology of clinically relevant disorders such as malnutrition, anorexia nervosa, obesity and diabetes. Despite the strong association between metabolism and GH, little is known regarding the basic mechanisms by which perturbations in metabolic pathways bring about changes in GH synthesis and release. Therefore, this application will determine the mechanisms by which alterations in nutrient availability; 1) regulate hypothalamic expression of neuropeptides essential for normal pituitary GH production (GH-releasing hormone [GHRH] and somatostatin [SRIF]), and 2) modify pituitary sensitivity to the GH-stimulatory peptides, GHRH and ghrelin. It has been proposed that changes in circulating leptin (an adipocyte factor) and ghrelin (a GH-releasing peptide produced in the stomach) mediates hypothalamic expression of GHRH and SRIF through activation of neuropeptide Y (NPY) neurons. To test this hypothesis, the effects of fasting on neuropeptide mRNA levels, in mice harboring defects in leptin synthesis (ob/ob), tissue source of leptin (AZIP-F1), NPY synthesis (NPY-/-), SRIF synthesis (smst-/-) and SRIF signaling (ss1l/2-/-) will be examined by ribonuclease protection assay and in situ hybridization. Also the effects of fasting in normal mice following exogenous hormone replacement (to increase NPY, leptin or ghrelin), pharmacological treatment (to block endogenous NPY production) or passive immunoneutralization (to block the actions of ghrelin) will be tested. Fasting not only alters expression of GH-regulatory neuropeptides but also enhances pituitary sensitivity to GHRH and ghrelin by increasing GHRH-R and GHS-R mRNA levels. In vitro, FFAs alone or in conjunction with glucocorticoids increase GHS-R synthesis. Therefore it is hypothesized that fasting induced elevations in FFA and glucocorticoids are required to enhance pituitary receptor synthesis and sensitivity, and thus compensate for fasting-induced alteration in central signals. To test this hypothesis, studies will examine the ability of fasting to alter pituitary receptor expression (by quantitative RT-PCR) and dynamic GH release (by RIA of serial blood samples) following blockade of FFA formation (by the anti-lipolytic Acipimox) or glucocorticoid actions (by the glucocorticoid receptor antagonist, RU-486). Also primary rat and non-human primate (baboon) pituitary call cultures will be used to determine if FFA and glucocorticoids mediate their effects on receptor synthesis via transcriptional or post-transcriptional processes and if the effects of FFA can be mimicked by activation of the putative FFA nuclear receptors, peroxisome proliferator-activated receptors (PPAR). This is the second revision of a competitive renewal of an RO1 application which continues to study the regulation of the GH axis. The current application centers on the interrelationship between changes in metabolic function and the GH axis.