Small molecule (SM)‐mediated neuronal reprograming of Müller glia (MG): A, Schematic diagram showing experimental design for the reprogramming of MG. B‐F, Temporal analysis of morphology of MG and immunoreactivities corresponding to β‐tubulin in response to SMs demonstrate the acquisition of neuronal features. At day 0, GFAP+ MG do not express β‐tubulin immunoreactivities. Upon differentiation MG turn on and off β‐tubulin and GFAP expression, respectively, with residual GFAP immunoreactivities detected in few cells' processes (arrow F). G‐H, Differentiated MG display both bipolar (arrowheads) and multipolar (arrow) neuronal morphologies, the former being predominant (H). I‐K, Double immunocytochemical analyses of cells at day 18 reveal co‐localization of Map2 (I)/Tau1 (J) in cells expressing β‐tubulin, suggesting the acquisition of mature neuronal phenotype. FIBD = Forskolin, ISX9, I‐BET, and DAPT. Scale bar = 50 μm.
Abstract
Müller glia (MG), cells that maintain homeostasis in the retina, are dormant stem cells that can regenerate neurons upon injury. However, the regenerative property of MG, which is reproducibly displayed in the lower vertebrates, is not readily observed in the mammals even upon forced expression of regulatory genes or exposure to growth factors. Here, we demonstrate a reproducible unmasking of the neurogenic properties of enriched rodent MG by serial exposure to different combinations of small molecules. The enriched MG, in response to changing culture conditions, silenced glia‐specific genes and acquired transcriptional signature of neurons, accompanied by upregulation of genes known to regulate neuronal potential of MG. The MG‐derived neurons expressed immunoreactivities corresponding to neuronal proteins and displayed electrophysiological features of immature neurons. Our study presents a proof of principle of pharmacological activation of neurogenic properties of mammalian MG, which may be utilized for therapeutic regeneration.