Barnea A, Aguila-Mansilla N, Chute H T, Welcher A A
Department of Obstetrics and Gynecology, University of Texas Southwestern Medical Center at Dallas 75235-9032, USA.
Brain Res. 1996 Sep 2;732(1-2):52-60. doi: 10.1016/0006-8993(96)00486-6.
Previous studies established that brain-derived neurotrophic factor (BDNF) induces neuropeptide Y (NPY) production and accumulation of NPY-mRNA in cultures of rat fetal brain tissues. In this study, we addressed the question: Are cultured human NPY neurons regulated by BDNF and/or by another member of the neurotrophin (NT) family of growth factors? Using aggregate cultures derived from human fetal cortical hemispheres, we assessed the effect of BDNF on NPY production varying the following experimental conditions: fetal and culture age; medium composition (with and without serum), dose and duration of exposure to BDNF, and neurotrophin species tested (BDNF, NT-4/5, NT-3 or NGF). Under none of these conditions did BDNF, NT-4/5, NT-3 or NGF induce an increase in NPY production. This was in contrast to forskolin + phorbol 12 myristate 13-acetate (PMA) which were highly effective in inducing NPY production, verifying that expression of NPY is a regulated process in these cultures. None of these neurotrophins enhanced the response to forskolin + PMA. By comparison, using aggregate cultures derived from rat fetal cortices, BDNF and NT-4/5 were equipotent in inducing NPY production but NT-3 and NGF were essentially ineffective. Moreover, the effects of BDNF or NT-4/5 and forskolin + PMA on NPY production were additive, indicating the involvement of distinct intracellular signalling pathways. Western blot analyses of human- and rat-derived aggregates indicated the presence of full-length Trk receptors which are tyrosine-phosphorylated in response to either BDNF, NT-4/5 or NT-3. Primary cultures of astrocytes (rat as well as human) were devoid of a functional TrkB receptor, strongly suggesting a neuronal expression of TrkB in the aggregates. Thus, a functional TrkB receptor is expressed by both the human and rat aggregates, but only the rat aggregates responded to BDNF or NT-4/5. These results are consistent with a difference in a post TrkB-receptor event(s) mediating BDNF action in the cultured human and rat fetal NPY neurons.
先前的研究证实，脑源性神经营养因子（BDNF）可诱导大鼠胎儿脑组织培养物中神经肽Y（NPY）的产生以及NPY-mRNA的积累。在本研究中，我们探讨了以下问题：培养的人NPY神经元是否受BDNF和/或神经营养因子（NT）家族的另一种生长因子调控？利用源自人胎儿皮质半球的聚集体培养物，我们通过改变以下实验条件来评估BDNF对NPY产生的影响：胎儿及培养年龄；培养基成分（含血清和不含血清）、BDNF的剂量和暴露持续时间，以及所测试的神经营养因子种类（BDNF、NT-4/5、NT-3或NGF）。在这些条件下，BDNF、NT-4/5、NT-3或NGF均未诱导NPY产生增加。这与福斯高林+佛波醇12-肉豆蔻酸酯13-乙酸酯（PMA）相反，后者在诱导NPY产生方面非常有效，证实了NPY的表达在这些培养物中是一个受调控的过程。这些神经营养因子均未增强对福斯高林+PMA的反应。相比之下，利用源自大鼠胎儿皮质的聚集体培养物，BDNF和NT-4/5在诱导NPY产生方面效力相当，但NT-3和NGF基本无效。此外，BDNF或NT-4/5与福斯高林+PMA对NPY产生的作用是相加的，表明涉及不同的细胞内信号通路。对源自人和大鼠的聚集体进行的蛋白质免疫印迹分析表明存在全长Trk受体，其在对BDNF、NT-4/5或NT-3的反应中发生酪氨酸磷酸化。星形胶质细胞（大鼠和人的）原代培养物缺乏功能性TrkB受体，强烈提示聚集体中TrkB的神经元表达。因此，人和大鼠的聚集体均表达功能性TrkB受体，但只有大鼠的聚集体对BDNF或NT-4/5有反应。这些结果与介导BDNF在培养的人及大鼠胎儿NPY神经元中作用的TrkB受体后事件的差异一致。
