NMDA receptor mediates proliferation and CREB phosphorylation in postnatal Müller glia-derived retinal progenitors

[摘要] Purpose: Postnatal retinal Müller gliaare considered to be retinal progenitors as they retain the ability todedifferentiate, proliferate, and differentiate to new retinal glia andneurons after injury. The proliferation and differentiation processesare coordinated by several extrinsic factors and neurotransmitters,including glutamate. Thus, the appropriate numbers and proportions ofthe different cell types are generated to form a functional retinaduring development and during injury repair. Here we analyze thechanges in the proliferation of postnatal Müller glia-derivedprogenitors after activation of the N-methyl-D-aspartate (NMDA)glutamate receptors. Methods: Müller glia-derived progenitorcell cultures were characterized by immunocytochemistry with antibodiesagainst the NR1 subunit of the NMDA receptor and the progenitor cellmarker nestin. The effect of glutamate receptor agonists andantagonists on cell proliferation was analyzed by BrdU incorporation orKi67 immunostaining, cell counting, and by immunolabeling ofphosphorylated cAMP response element binding protein (P-CREB)transcription factor. The effect of NMDA receptor activation wasanalyzed in vivo by P-CREB immunohistochemistry in retinal sections ofLong-Evans NMDA injected rats. Results: We show that NMDA receptoractivation significantly increases the proliferation rate ofMüller-glia derived progenitor cells and that this increase can beblocked by NMDA receptor antagonists. Furthermore, we show that CREBphosphorylation is induced in NMDA-treated Müller-glia derivedprogenitor cells in culture and that specific pharmacologicalinhibition of CREB phosphorylation results in a decreased number ofproliferating cells. We confirmed the relevance of these observationsby the analysis of retinal sections after NMDA injection in vivo whereimmunoreactivity to phosphorylated CREB is also increased aftertreatment. Conclusions: In the present study weshow that NMDA receptor activation induces postnatal Müllerglia-derived retinal cell progenitor proliferation and transcriptionfactor CREB phosphorylation both in culture and in vivo. Theidentification of the molecular determinants of mature retinalprogenitors such as transcription factor CREB and NMDA receptor-inducedplayers should facilitate the control of growth and manipulation ofprogenitor cell cultures and the possible identification of themolecular mechanisms involved in progenitor self-renewal.

[学科分类] 生物化学/生物物理