一氧化氮对体外培养的胚鼠脑干5-羟色胺能神经元生长发育的影响

研究体外培养条件下一氧化氮合酶(NOS)对胚鼠脑干5-羟色胺(5-HT)能神经元生长发育的影响,进一步探讨一氧化氮在胚脑发育过程中的作用。将孕14d SD胚鼠脑干细胞悬液接种于24孔培养板,实验分两组:即常规培养液组及NOS竞争性抑制剂L-NAME处理组。两组均在接种5、10、15、20d后终止培养,部分盖片行NADPH-d组织化学染色,余盖片行5-HT免疫组织化学染色。观察并计数各组NOS阳性神经元和5-HT样免疫阳性神经元的形态和数量。结果显示:经L-NAME处理的胚鼠脑干细胞在培养10d后出现NOS阳性神经元减少。虽在整个过程中未观察到5-HT样免疫阳性神经元胞体的变化,但其突起的密度及突起上膨体和生长锥的数量明显减少。以上结果表明:在生长发育的关键时期,应用NOS抑制剂会影响5-HT能神经元的形态发育和功能,提示NO参与调节神经元的发育与成熟过程。

EFFECT OF NITRIC OXIDE ON THE DEVELOPMENT OF SEROTONERGIC NEURONS IN THE EMBRYONIC RAT BRAIN STEM IN VITRO

In the present study, we observed the effect of interference with nitric oxide (NO) synthesis on the development of serotonergic neurons in the embryonic rat brain stem in vitro and realized further the possible roles of NO during embryonic development. Brain stem cell suspension obtained from E 14 rat were plated at 24 well culture plate, were kept in defined medium or in defined medium plus the competitive inhibitor N-nitric-L-arginine methyl ester(L-NAME). Cultures were terminated, respectively, at 5,10,15,20 days in vitro. Some slips were stained for NADPH-d activity, while the remains were stained by 5-HT immunocytochemical method. The number and shape about the NOS-positive neurons and 5-HT-like immunoreactive (LI)cells of each group were observed. The results were as follows: chronic exposure of brain stem cells to L-NAME resulted in slight decrease of surviving neurons after 10 d in culture. In the whole experiment, the variation of the 5-HT-LI cell bodies was not observed, but the density of neuropile and the quantities of varicosities and growth cones obviously decreased. Our study demonstrated that nitric oxide synthesis inhibition may have effect on serotonergic neuronal morphological development and function during the critical period of brain development. These results indicated that NO could be involved in neuronal development and maturation.