麩胺酸受體在神經分化過程中所扮演的角色
Glutamate Receptor in Neuronal Differentiation
中文摘要
麩胺酸受體可藉由受體離子通道，或連接次級訊息傳遞路徑，而造成細胞內鈣離
子濃度的上升；所以了解在發育早期神經元中，麩胺酸受體對鈣離子的調控作
用，對於探討其在神經發育過程中所扮演的角色是很重要的。在本篇論文的研究
主題中，我們利用源自胚胎的幹細胞，P19 所分化而來的發育早期的神經細胞，
來研究神經發育期中，麩胺酸受體所引發的細胞內鈣離子濃度的上升，及他們對
生長因子受體的表現與細胞的計劃性死亡之影響。結果如下： (1) AMPA 受體
的次單位 (subunits) 麩胺酸受 2/3 (GluR2/3)，在神經細胞開始形成的時期
就會有表現。 (2) 紅藻胺酸受體之次單位麩胺酸受體 5/6/7 (GluR5/6/7)，則
是在原始
P19 幹細胞中即已有表現，其後在神經細胞的分化過程中則逐漸減
少，而在神經元成熟之後
GluR5/6/7 的表現量又會增加。 (3) 而在 5DAR 的
P19 神經細胞膜上，也可看到 mGluR1 受體的表現。 (4) 麩胺酸受體的專
一性致效劑 (agonist) KA、AMPA 及 NMDA，在以維生素 A 酸處理後的 3DAR
P19-derived neuron 中，並不會影響細胞內鈣離子濃度變化的情形，而一直
到
5DAR 及 7DAR 時，才會造成細胞內鈣離子濃度的增加；然而，t-ACPD 則
一直到
7DAR 時，都不會去影響 P19-derived neuron 的細胞內鈣離子的濃度。
(5)而在 5DAR 的 P19-derived neuron 給予興奮性胺基酸致效劑刺激時，發
現會增加細胞內
p53 與 Bax 的表現。至於 Bcl-2 則是除了紅藻胺酸外，其餘的
興奮性胺基酸都會減少其表現。而
TrkA 也會因興奮性胺基酸的作用，而增加其
表現。 (6) 麩胺酸致效劑雖不會影響細胞的計劃性死亡，但可以透過與神經生
長因子之間的相互作用，使細胞的計劃性死亡降低，這作用亦已經由西方墨點法
證實。由以上的實驗結果可知，麩胺酸受體不只會調控細胞內鈣離子濃度的上
升，亦因受體
subtype 的不同而有所差異。同時，他們也會去調節與細胞自殺
性死亡相關蛋白質及神經營養因子受體
TrkA 的表現。由麩胺酸受體的特異性致
效劑所導致
p53 與 Bax 的減少及 Bcl-2 與 TrkA 表現增加，證明了興奮性胺基
酸在發育中的神經細胞所具有的保護作用當中，可能扮演著一個相當重要的角
色。
英文摘要
Glutamate receptors (GluRs) are known to mediate intracellular calcium elevation through receptor ion channels or coupling to second messenger pathways. Knowing their calcium regulation in early developing neurons may provide important clues for their developmental roles. In this study, we used early developing neurons differentiated from embryonic stem cell P19 cell line to investigate GluR-mediated
intracellular calcium surge, expression of growth factor receptors and programmed cell death during neuronal development. Results are summarized as following. (1) The expression of the AMPA receptor subunit GluR2/3 appeared at the same stage as the beginning of neurogenesis. (2) The expression of the KA receptor subunit GluR5/6/7 appeared as early as in the stem cell stage, decreased during
differentiation, and increased toward neuronal maturation. (3) In 5DAR P19 neurons, mGluR1 receptor also expressed on the membrane. (4) Specific GluR agonists KA, AMPA, NMDA had no increase of [Ca2+]i changes in 3DAR P19-derived neurons until 5DAR and 7DAR, whereas trans-ACPD had no effect at 7DAR. (5) EAAs resulted in induction of p53 and Bax protein expression in 5DAR P19 neurons. Other than KA, all EAAs were resulted in reduction of Bcl-2 expression. And the expression of TrkA receptors were induced by all EAAs. (6) Glutamate had no effect on
apoptosis of P19 neurons, but following NGF treatment, all EAAs could reduce their apoptosis. These results were also be proved by Western blot analysis, and EAA may cooperate with NGF on regulating neuronal differentiation and survival. These results suggest that glutamate receptors are not only acting differently in terms of mediating [Ca2+]i, but also regulating apoptotic or antiapoptotic protein and NGF receptor expression. The glutamate receptor-mediated profound decreased of p53 and Bax protein expression; induced Bcl-2 and TrkA expression by glutamate receptor agonists implies the EAA-mediated cell protection may play an important role in neural development.
