Snyder G L, Girault J A, Chen J Y, Czernik A J, Kebabian J W, Nathanson J A, Greengard P
Laboratory of Molecular and Cellular Neuroscience, Rockefeller University, New York, New York 10021.
J Neurosci. 1992 Aug;12(8):3071-83. doi: 10.1523/JNEUROSCI.12-08-03071.1992.
The molecular mechanisms underlying regulation of fluid production by secretory epithelia such as the choroid plexus are poorly understood. Two cAMP-regulated inhibitors of protein phosphatase-1, inhibitor-1 (I1) and a dopamine- and cAMP-regulated phosphoprotein, M(r) = 32,000 (DARPP-32), are enriched in the choroid plexus. We show here that these two phosphoproteins are colocalized in choroid plexus epithelial cells. We have developed a novel method for studying the phosphorylation state of DARPP-32 and I1 in intact cells, using a phosphorylation state-specific monoclonal antibody. Several drugs and hormones that are known to alter fluid secretion and that increase cAMP levels (forskolin, isoproterenol, vasoactive intestinal peptide) or cGMP levels (atrial natriuretic peptide) or that may use additional second messenger pathways (5-HT), increase the phosphorylation of I1 and DARPP-32 in rat choroid plexus. In contrast, dopamine does not alter cAMP and cGMP levels, or I1 and DARPP-32 phosphorylation. Our results indicate that DARPP-32, known to be regulated by dopamine in a number of tissues, can be phosphorylated in response to non-dopaminergic factors, including hormones acting through non-cAMP-dependent pathways. Our results also raise the possibility that inhibition of phosphatase-1, as a result of I1 and DARPP-32 phosphorylation, might be part of a final common pathway in the action of several factors that are known or thought to alter cerebrospinal fluid production.
诸如脉络丛等分泌上皮调节液体生成的分子机制仍知之甚少。两种受环磷酸腺苷（cAMP）调节的蛋白磷酸酶-1抑制剂，即抑制剂-1（I1）和一种多巴胺及cAMP调节的磷蛋白，分子量为32,000（DARPP-32），在脉络丛中含量丰富。我们在此表明，这两种磷蛋白在脉络丛上皮细胞中共定位。我们开发了一种新方法，使用磷酸化状态特异性单克隆抗体来研究完整细胞中DARPP-32和I1的磷酸化状态。几种已知可改变液体分泌且能提高cAMP水平（福斯可林、异丙肾上腺素、血管活性肠肽）或cGMP水平（心房利钠肽）或可能利用其他第二信使途径（5-羟色胺）的药物和激素，会增加大鼠脉络丛中I1和DARPP-32的磷酸化。相比之下，多巴胺不会改变cAMP和cGMP水平，也不会改变I1和DARPP-32的磷酸化。我们的结果表明，已知在许多组织中受多巴胺调节的DARPP-32，可响应非多巴胺能因子而被磷酸化，这些因子包括通过非cAMP依赖途径起作用的激素。我们的结果还提出了一种可能性，即由于I1和DARPP-32磷酸化导致的蛋白磷酸酶-1抑制，可能是几种已知或被认为会改变脑脊液生成的因子作用的最终共同途径的一部分。
