Effects of orexin on catecholamine synthesis and release in adrenal medulla

食欲素-A和食欲素-B，也称为下丘脑泌素-1和下丘脑泌素-2，最近被鉴定为下丘脑肽，其来源于位于大鼠脑中下丘脑外侧和后部内和周围的神经元中的前体食欲素原。为了确定食欲素-A在外周代谢过程中的作用，我们研究了食欲素-A对培养的牛肾上腺髓质细胞中儿茶酚胺合成和分泌的直接影响。1)将细胞与食欲素-A（100 pM）孵育20分钟，引起<14>由[13 <14>C]酪氨酸而不是由L-3，4-二羟基苯基[3- 13 <14>C]丙氨酸合成13 C-儿茶酚胺的少量但显著的增加。食欲素-A（100 pM）增强了乙酰胆碱（0.3 mM）13 <14>C-儿茶酚胺合成的刺激作用。2)食欲素-A显著增加酪氨酸羟化酶活性，这在1 pM时是明显的，在100 pM时最大。蛋白激酶C激活剂4β-佛波醇-12 β-肉豆蔻酸酯-13 α-乙酸酯不能增强orexin-A对酪氨酸羟化酶活性的刺激作用，而蛋白激酶C抑制剂H-7和staurosporine则使orexin-A的作用无效。3)食欲素-A对细胞分泌儿茶酚胺的影响不大。4)用逆转录-聚合酶链反应（RT-PCR）技术在牛肾上腺髓质细胞中检测到食欲素受体1（OX_1R）mRNA，但未检测到食欲素受体2（OX_2R）mRNA，提示食欲素A可能通过激活OX_1R-蛋白激酶C途径激活肾上腺髓质细胞酪氨酸羟化酶，进而促进儿茶酚胺合成。这些信息有助于了解食欲素对成人和儿童肥胖症代谢稳态影响的生化基础。

Orexin-A and orexin-B, also known as hypocretin-1 and hypocretin-2, have recently been identified as hypothalamic peptides derived from a precursor, prepro-orexin, in neurons located within and around the lateral and posterior hypothalamus in the rat brain. To determine the role of orexin-A in peripheral metabolic processes, we examined direct effects of orexin-A on catecholamine synthesis and secretion in cultured bovine adrenal medullary cells. 1) Incubation of cells with orexin-A (100 pM) for 20 min caused a small but significant increase in ^<14>C-catecholamine synthesis from [^<14>C] tyrosine, but not from L-3,4-dihydroxyphenyl [3-^<14>C] alanine. Orexin-A (100 pM) potentiated the stimulatory effects of acetylcholine (0.3 mM) ^<14>C-catecholamine synthesis. 2) Orexin-A significantly increased tyrosine hydroxylase activity, which was evident at 1 pM and maximal at 100 pM. 4β-Phorbol-12β-myristate-13α-acetate, an activator of protein kinase C, did not enhance the stimulatory effects of orexin-A on tyrosine hydroxylase activity, while H-7 and staurosporine, inhibitors of protein kinase C, nullified the effects of orexin-A. 3) Orexin-A had little effect on catecholamine secretion from the cells. 4) Orexin receptor 1 (OX_1R) but not orexin receptor 2 (OX_2R) mRNA was detected in bovine adrenal medullary cells by reverse transcriptase-polymerase chain reaction.These findings suggest that orexin-A activates tyrosine hydroxylase and then stimulates catecholamine synthesis, probably via activation of the OX_1R-protein kinase C pathway in adrenal medullary cells. This information helps to understand the biochemical basis of the effect of orexins on metabolic homeostasis in the obesity of adults as well as children.