Kapcala L P
Department of Medicine, University of Maryland School of Medicine and Hospital, Baltimore 21201.
Brain Res. 1989 Jul 10;491(2):253-65. doi: 10.1016/0006-8993(89)90061-9.
Despite many in vivo studies, little is known about brain regulation of POMC synthesis or regulation of secretion of POMC-related peptides. To test the hypothesis that dissociated brain cells in culture can produce and release POMC-related peptides, immunoreactive (IR)-adrenocorticotropin (ACTH) and beta-endorphin were measured in cells and media of dissociated cell cultures incubated up to 38 days. Fetal rat hypothalamic and extrahypothalamic forebrain cells were maintained in serum free medium. IR-ACTH and beta-endorphin were measured by radioimmunoassay in concentrated cells and media after various incubation times using two ACTH (mid-portion = R4; carboxy-portion directed = KEND) antisera and a beta-endorphin antiserum. IR-ACTH and IR-beta-endorphin in hypothalamic and extrahypothalamic cells and in media (cumulative) were greater than quantities in cells before culture. Peak hypothalamic cellular content of IR-ACTH (5.3 fmol/10(6) cells-R4; 4.7 fmol/10(6) cells-KEND) and content of IR-beta-endorphin (32.0 fmol/10(6) cells) occurred on days 16, 9 and 23, respectively. Peak extrahypothalamic content of IR-ACTH (2.9 fmol/10(6) cells-R4; 1.0 fmol/10(6) cells-KEND) and content of IR-beta-endorphin (10.8 fmol/10(6) cells) was also seen on different days, was lower than hypothalamic content and was not always concurrent with peak hypothalamic content. Gel filtration chromatography revealed that the predominant forms of IR-ACTH and IR-beta-endorphin in hypothalamic cell extracts co-eluted with synthetic ACTH1-39 and beta-endorphin. Changes in molar ratios of IR-ACTH and IR-beta-endorphin also suggested a differential regulation of different POMC derivatives.
(1) IR-ACTH and IR-beta-endorphin are produced by hypothalamic and extrahypothalamic forebrain cells in culture: and (2) dissociated brain cell cultures can be used as a potential model for studying regulation of POMC-related peptides in brain.
尽管进行了许多体内研究，但对于脑对促黑素细胞激素（POMC）合成的调节或POMC相关肽分泌的调节了解甚少。为了验证培养的解离脑细胞能够产生并释放POMC相关肽这一假设，在培养长达38天的解离细胞培养物的细胞和培养基中测量了免疫反应性（IR）促肾上腺皮质激素（ACTH）和β-内啡肽。将胎鼠下丘脑和下丘脑外前脑细胞维持在无血清培养基中。在不同孵育时间后，使用两种ACTH抗血清（中部=R4；羧基末端定向=KEND）和一种β-内啡肽抗血清，通过放射免疫分析法在浓缩的细胞和培养基中测量IR-ACTH和β-内啡肽。下丘脑和下丘脑外细胞以及培养基中（累积）的IR-ACTH和IR-β-内啡肽含量高于培养前细胞中的含量。下丘脑细胞中IR-ACTH的峰值含量（5.3 fmol/10⁶细胞-R4；4.7 fmol/10⁶细胞-KEND）和IR-β-内啡肽的含量（32.0 fmol/10⁶细胞）分别出现在第16天、第9天和第23天。下丘脑外IR-ACTH的峰值含量（2.9 fmol/10⁶细胞-R4；1.0 fmol/10⁶细胞-KEND）和IR-β-内啡肽的含量（10.8 fmol/10⁶细胞）也出现在不同的日子，低于下丘脑含量，且并不总是与下丘脑峰值含量同时出现。凝胶过滤色谱显示，下丘脑细胞提取物中IR-ACTH和IR-β-内啡肽的主要形式与合成的ACTH1-39和β-内啡肽共洗脱。IR-ACTH和IR-β-内啡肽摩尔比的变化也表明对不同POMC衍生物的调节存在差异。
（1）培养的下丘脑和下丘脑外前脑细胞可产生IR-ACTH和IR-β-内啡肽；（2）解离的脑细胞培养物可作为研究脑中POMC相关肽调节的潜在模型。
