The Research Of Galanin's Effects Of Proliferation And Differentiation Of Neural Stem Cell

Galanin, a neuropeptides of 29-30 amino acids is widely distributed in the central and peripheral nervous system. Numerous studies have shown that galanin mediates a variety of physiological activities such as cognition in memory, the modulation of neurotransmitter and hormone release, the nociception in the spinal cord and the promotion of neuroregenesis and neurite outgrow. The biological effects of galanin depend on the specific galanin receptors found at the cell surface, to date, three galanin receptor subtypes, GalRl, GalR2, and GalR3 have been cloned and they all belong to the G-protein-coupled receptor superfamily, but it is certainly possible that other subtypes exist. However, there are big differences between the distribution of the galanin receptors in CNS which suggests specific selectivity of galanin-mediated effects. Recent reports have described that galanin and its receptors are expressed in embryonic stem cells of mammalian and in neural stem/progenitor cells located in the proliferation zones of SVZ in the brain of adult rat, which indicates that galanin may be involved in the neurodevelopment. In order to validate this speculation, for the first time we detected the expression of three galanin receptors in cultured neural stem cells using the method of RT-PCR, after got some useful information about receptor expression, we then studied the effects of galanin and its receptors' specific agonist and antagonist on the proliferation and differentiation of neural stem cells. The experiment received some primary results. In this experiment:1.The neural stem cells were isolated from the SVZ of the embryonic 14 day rats brain and were cultured and proliferated in the no-serum cell culture medium supplemented with bFGF and B27. The different cell types were identified by the specific antigen using the method of immunohistochemistry.2.A RT-PCR reaction was performed to detect the expression of three galanin receptors in cultured neural stem cells using the specific primer aimed at different receptor subtypes. 3.The effects of galanin on the proliferation of neural stem cells was investigated bythe means of MTT, after that, double lable immunohistochemistry was performed to analysis the influences of these substances on the differentiation-induced cells and the two cell phenotypes(neurons and astrocyte) was determined by their specific marker , finally the proportion of glial cells was compared between the different groups. The results of this study showed that:1 .The cells isolated from SVZ of embryonic rats can proliferate continuously in vitro, they have the ability of self-renewal and show the muiti-differentiation potential and can be identified as neural stem cells of CNS.2.There is expression of GalRl and GalR2 mRNA in cultured neural stem cells. 3-Galanin, galaninl-11, galanin2-ll can inhibit the proliferation of neural stem cells and also can increase the astrocyte proportion after the differentiation of neural stem cells and these two effects can both be blocked by M35.The present experiment demonstrated that there was expression of type-1,2 galanin receptors in cultured neural stem cells. Galanin, galanin receptor non-selective agonist (galaninl-11) and type-2 receptor selective agonist (galanin2-ll) can inhibit the proliferation of neural stem cells effectively and this action can be blocked by the galanin receptors non-selective antagonist M35. Moreover, galanin, galaninl-11, galanin2-ll also have a tendency to promote glial cells differentiation , and this action can be blocked by M35 too.