Functional Study of Hippocampal Neurons in Mice Deficient with Collapsin Response Mediator Protein-1

• Main Authors: Yun-Chieh Tsai, 蔡昀潔
• Other Authors: Shu-Rung Lin
• Format: Others
• Language: zh-TW
• Published: 2010
• Online Access: http://ndltd.ncl.edu.tw/handle/6y8fh7

碩士 === 中原大學 === 生物科技研究所 === 98 === Collapsing response mediator protein-1 ( CRMP-1 ) belongs to a family of cytosolic phosphorproteins that mediate the extracellular signals from semaphorin 3A. It was initially identified in brain and has been implicated in plexin-dependent neuronal function. In order to study the physiological function of CRMP-1 in hippocampus, primary cultures of hippocampal neurons were prepared from wild-type ( WT ) and crmp-1 knockout ( KO ) neonatal mice. In the studies of pre-synaptic functions, hippocampal neurons of crmp-1 KO mice showed a marked increase in glutamate secretion when treating with 75 mM KCl solution, however there was no difference between WT and crmp-1 KO neurons in glutamate secretion that leaked out at the nerve terminals. In addition, intracellular Ca2+ concentration ( [Ca2+]i ) was also measured after the addition of 75 mM KCl. At transient stage, both WT and crmp-1 KO neurons have similar [Ca2+]i . But at 150 seconds, crmp-1 KO neurons exhibit a higher [Ca2+]i than WT neurons. This result indicated that crmp-1 KO neurons still maintain in a higher activation of voltage-sensitive Ca2+ channel after a few minutes. On the other hand, for studying the function of NMDA receptor in post-synapses, 100 and 500 μM glutamate were added, and a higher [Ca2+]i was detected in WT neurons, but not in crmp-1 KO neurons. This indicates that the number or sensitivity of NMDA receptor in crmp-1 KO neurons is less or lower then in WT neurons. Furthermore, neuronal stem cells ( NSC ) from WT and crmp-1 KO mice were also isolated and their migration rate and differentiation was determined. Our results showed that neuronal stem cells from WT mice have higher migration rate and differentiation ability than those from KO mice. Thus, in conclusion, our results demonstrate that neurons disrupted with crmp-1 gene exhibit significant differences in both presynaptic and postsynaptic functions of differentiated neurons and migration and differentiation of neuronal stem cells, when compared to WT neurons.