Hormones, Neuronal Phosphoproteins and Behavior

描述(申请人提供)：卵巢类固醇激素，雌二醇和黄体酮，调节中枢神经系统的功能，导致生理和生殖行为的改变。雌二醇和孕酮对生殖行为发挥神经作用的一种方式是通过它们与同源的细胞内受体相互作用，作为配体依赖的转录因子。我实验室的研究表明，除了类固醇激素外，神经递质多巴胺还可以以一种不依赖配体的方式激活孕激素受体。多巴胺促进雌性大鼠和小鼠的生殖行为是通过多巴胺的膜受体和细胞内的孕激素受体之间的串扰来实现的。 多巴胺和孕酮启动的信号转导通路导致下丘脑中cAMP水平、PKA活性和苏氨酸34上DARPP-32磷酸化水平的增加。除了它在苏氨酸34上的磷酸化外，DARPP-32还可以在苏氨酸75以及两个丝氨酸残基Ser102和Ser137上被磷酸化。这种多点磷酸化通过将DARPP-32转化为更好的PKA磷酸化底物并增强其对下游蛋白磷酸酶-1(PP-1)级联的抑制作用来放大DARPP-32的作用。虽然孕酮和多巴胺都增强了下丘脑中的PKA活性，但这种增加调节DARPP-32抑制PP-1的机制(S)尚未得到评估。这项建议着重于孕酮和多巴胺对DARPP-32多位点磷酸化的潜在机制及其对雌性生殖行为的影响。具体目标1将评估孕酮和多巴胺对下丘脑Thr34和Thr75处DARPP-32磷酸化的影响，以及由此产生的PP-1在介导生物学效应中的调节。特异性目标2将确定多巴胺和孕酮对下丘脑Ser102和Ser137上DARPP-32磷酸化状态的调节作用，并检测它们对DARPP-32/PP-1级联扩增的影响。具体目标3将使用DARPP-32 Thr34、Thr75、Ser102和Ser137点突变的突变小鼠来研究黄体酮和多巴胺对生殖行为的影响。这些研究将为孕酮和多巴胺在调节生殖行为中对PKA/DARPP-32/PP-1的级联调节提供信息。

DESCRIPTION (provided by applicant): The ovarian steroid hormones, estradiol and progesterone, regulate functions in the central nervous system resulting in alterations in physiology and reproductive behavior. One means by which estradiol and progesterone exert their neural effects on reproductive behavior is via their interaction with cognate, intracellular receptors functioning as ligand-dependent transcription factors. Studies from my laboratory have shown that in addition to steroid hormones, neurotransmitter dopamine can activate progestin receptors in a ligand-independent manner. Dopamine facilitation of reproductive behavior in female rats and mice occurs by means of cross-talk between membrane receptors for dopamine and intracellular progestin receptors. Signal transduction cascades initiated by dopamine and progesterone, cause an increase in the level of cAMP, PKA activity and phosphorylation of DARPP-32, on Threonine34 in the hypothalamus. In addition to its phosphorylation on Threonine34, DARPP-32 can also be phosphorylated on Threonine75, as well as two Serine residues Ser102 and Ser137. This multisite phosphorylation amplifies the effects of DARPP-32 by converting it into a better substrate for PKA phosphorylation and potentiating its inhibitory effects on the downstream protein phosphatase-1 (PP-1) cascade. While both progesterone and dopamine enhance PKA activity in the hypothalamus, the mechanism(s) by which this increase regulates DARPP-32 inhibition of PP-1 have not been evaluated. This proposal focuses on the mechanisms underlying this multi-site phosphorylation of DARPP-32 by progesterone and dopamine and their effects on female reproductive behavior. Specific aim 1 will evaluate the effects of progesterone and dopamine on DARPP-32 phosphorylation at Thr34 and Thr75 in the hypothalamus and the resultant regulation of PP-1 in mediating the biological effects. Specific aim 2 will determine the modulatory role of dopamine and progesterone on the state of phosphorylation of DARPP-32 on Ser102 and Ser 137 in the hypothalamus and examine their effects on the amplification of DARPP-32/PP-1 cascade. Specific aim 3 will examine the effects of progesterone and dopamine on reproductive behavior using mutant mice with point mutations at Thr34, Thr75, Ser102 and Ser137 of DARPP-32. These studies will provide information on the PKA/DARPP-32/PP-1 cascade regulation by progesterone and dopamine in mediating reproductive behavior.