Roseboom P H, Klein D C
Section of Neuroendocrinology, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892.
Mol Pharmacol. 1995 Mar;47(3):439-49.
Norepinephrine (NE) regulates melatonin production and many other aspects of pineal function through actions involving cAMP. In the present study the effects of NE on the phosphorylation of the cAMP response element-binding protein (CREB) were studied to determine whether CREB phosphorylation might be involved in cAMP signal transduction in this tissue. CREB was detected using gel mobility-shift analysis with the radiolabeled Ca2+/cAMP response element of the c-fos promoter. CREB phosphorylation was estimated in the gel mobility-shift assay using an antiserum specific for phosphorylated CREB. This antiserum generates a supershifted CREB signal with protein extracts obtained from glands treated with NE (EC50 approximately equal to 10 nM) in organ culture, demonstrating that NE stimulates CREB phosphorylation. CREB phosphorylation peaks 30-45 min after NE treatment is initiated and then gradually returns to base-line values. Pharmacological studies show that NE-stimulated CREB phosphorylation is mediated primarily through beta 1-adrenergic receptor-stimulated increases in cAMP. Activation of alpha 1-adrenergic receptors, which is known to elevate the intracellular free Ca2+ concentration, does not cause CREB phosphorylation. However, it is possible to produce CREB phosphorylation with certain pharmacological agents that elevate the intracellular free Ca2+ concentration. In vivo studies show that CREB phosphorylation can be induced by treatment with isoproterenol (1 mg/kg), demonstrating that phosphorylation of pineal CREB occurs in intact animals. These studies indicate that cAMP-dependent CREB phosphorylation could play a role in the adrenergic regulation of gene expression in pinealocytes.
去甲肾上腺素（NE）通过涉及环磷酸腺苷（cAMP）的作用来调节褪黑素的产生以及松果体功能的许多其他方面。在本研究中，研究了NE对环磷酸腺苷反应元件结合蛋白（CREB）磷酸化的影响，以确定CREB磷酸化是否可能参与该组织中的cAMP信号转导。使用凝胶迁移率变动分析，结合c-fos启动子的放射性标记的Ca2+/cAMP反应元件来检测CREB。在凝胶迁移率变动分析中，使用针对磷酸化CREB的抗血清来估计CREB磷酸化。在用NE（半数有效浓度约等于10 nM）处理的器官培养腺体的蛋白质提取物中，这种抗血清产生超迁移的CREB信号，表明NE刺激CREB磷酸化。NE处理开始后30 - 45分钟，CREB磷酸化达到峰值，然后逐渐恢复到基线值。药理学研究表明，NE刺激的CREB磷酸化主要通过β1 - 肾上腺素能受体刺激cAMP增加来介导。已知可提高细胞内游离Ca2+浓度的α1 - 肾上腺素能受体的激活不会导致CREB磷酸化。然而，使用某些可提高细胞内游离Ca2+浓度的药剂可以产生CREB磷酸化。体内研究表明，用异丙肾上腺素（1 mg/kg）处理可诱导CREB磷酸化，表明松果体CREB磷酸化发生在完整动物体内。这些研究表明，cAMP依赖性CREB磷酸化可能在松果体细胞基因表达的肾上腺素能调节中起作用。
