Mechanisms of ATP Release from Glial Progenitors

神经胶质细胞的核苷酸释放在CMSand中起重要的自分泌和旁分泌信号作用 PNS。最近的研究表明，神经胶质细胞ATP的非溶解性释放可能涉及受调节的 胞吐作用或通过核苷酸渗透通道流出。细胞外ATP作用于质膜 亲离子型P2 X和亲代谢型P2 Y受体调节神经元活性，诱导钙瞬变， 星形胶质细胞和产生关键的细胞内第二信使， 脑外伤后神经胶质增生和神经干细胞的增殖和迁移。最近我们 获得的证据表明，神经祖细胞显示自发钙振荡， 依赖于P2 R的激活。嘌呤能受体拮抗剂或ATP降解剂的浴用 腺苷三磷酸双磷酸酶可防止自发性钙波动的发生。此外，封锁 P2 Rs抑制了这些祖细胞的增殖和迁移率。这些结果表明 神经前体细胞具有释放ATP的能力，ATP随后发挥关键的自分泌、旁分泌信号传导作用 在早期中枢神经系统发育过程中的作用。虽然识别的途径涉及递质释放， 神经胶质细胞被认为是成熟的星形胶质细胞，但对它的发育方面还一无所知。 ATP从前体释放的机制。因此，本申请的目的是 为了评估是否以及在星形胶质细胞发育的哪个阶段， 递质释放在功能上是有能力的（胞吐/分泌和通过离子型P2 XR的扩散 和连接蛋白半通道）并确定它们对钙振荡和细胞迁移的贡献。 免疫细胞化学、生物化学和药理学方法结合发光和 荧光成像将用于识别ATP释放途径。这种识别对于 了解CNS发育和修复过程中的信号传导机制。

Nucleotide releasefrom glial cells plays important autocrine and paracrine signaling roles in the CMSand PNS. Recentstudies suggest that the non-lytic releaseof ATP from glial cells can either involve regulated exocytosis or efflux through nucleotide-permeable channels. Extracellular ATP acting onplasmalemmal ionotropic P2X and metabotropic P2Y receptors modulates neuronal activity, induces calcium transients in astrocytes and generatescritical intracellular second messengers implicated for instance in the development of gliosis following brain trauma and in the proliferation and migration of neural stem cells. Recently, we have obtained evidencethat neural progenitor cells display spontaneous calcium oscillations that are highly dependent on activation of P2Rs. Bath application of purinergic receptor antagonists or of the ATPdegrading enzyme apyrasepreventedthe occurrence of spontaneous calcium fluctuations. Moreover,blockade of P2Rs reduced the proliferation and the migration ratesof these progenitors. These results suggest that neural progenitors have the ability to release ATP,which then plays a crucial autocrine, paracrine signaling role during early CNS development.Although identification of pathways involvedon transmitter release from glial cells have been suggested for mature astrocytes, nothing is known about the developmental aspectsof the mechanism(s) by which ATP is released from precursors. Therefore, the goal of this grant application is to evaluate whether and at which stage of astrocyte development three of the putative pathways for transmitter release are functionally competent (excocytosis/secretion and diffusion through ionotropic P2XRs and connexin hemichannels) and to determine their contribution to calcium oscillations and cell migration. Immuocytochemical, biochemical and pharmacological approaches combined with luminescence and fluorecence imaging will be used to identify ATP release pathways. This identification is fundamental for the understanding of the signaling mechanisms ongoing during CNS development and repair.