Restoration of Visual Function and Cortical Connectivity After Ischemic Injury Through NeuroD1-Mediated Gene Therapy

Abstract

Neural circuits underlying brain functions are vulnerable to damage, including ischemic injury, leading to neuronal loss and gliosis. Recent technology of direct conversion of endogenous astrocytes into neurons in situ can simultaneously replenish the neuronal population and reverse the glial scar. However, whether these newly reprogrammed neurons undergo normal development, integrate into the existing neuronal circuit, and acquire functional properties specific for this circuit is not known. We investigated the effect of NeuroD1-mediated in vivo direct reprogramming on visual cortical circuit integration and functional recovery in a mouse model of ischemic injury. After performing electrophysiological extracellular recordings and two-photon calcium imaging of reprogrammed cells in vivo and mapping the synaptic connections formed onto these cells ex vivo, we discovered that NeuroD1 reprogrammed neurons were integrated into the cortical microcircuit and acquired direct visual responses. Furthermore, following visual experience, the reprogrammed neurons demonstrated maturation of orientation selectivity and functional connectivity. Our results show that NeuroD1-reprogrammed neurons can successfully develop and integrate into the visual cortical circuit leading to vision recovery after ischemic injury.

In vivo glia‐to‐neuron conversion: pitfalls and solutions

“…The answers to these questions, unfortunately, are a resounding no. By prelabeling astrocytes with a genetically encoded reporter in genetic lineage tracing mouse lines and using a different reporter for the later injected AAVs (Wang et al, 2020;Wang et al, 2021), our recent study unambiguously demonstrated that resident astrocytes are not the cell origin for those viral reporter-labeled "new" neurons that were observed in the studies of NEUROD1 (Y. C. Chen et al, 2020;Tang et al, 2021;Wu et al, 2020) or PTBP1 (Qian et al, 2020;Zhou et al, 2020). This conclusion is further supported by the results of several independent studies on either NEU-ROD1 (Leib et al, 2022) or PTBP1 (W. Chen et al, 2021;Hoang et al, 2021;Leib et al, 2022).…”

“…For example, AAV‐mediated ectopic expression of NEUROD1 or knockdown of PTBP1 was reported to induce highly efficient astrocyte‐to‐neuron conversion, with rapid appearance of viral reporter‐labeled mature neurons exhibiting perfect regional identity, electrophysiology, axonal projections, and connectivity (Y. C. Chen et al., 2020; Qian et al., 2020; Wu et al., 2020; Zhou et al., 2020). Astonishingly, studies with NEUROD1 or PTBP1 also reported greatly improved behavioral functions in many animal disease models including ischemic brain injury (Y. C. Chen et al., 2020; Tang et al., 2021), retinal injury (Zhou et al., 2020), Hungtington's disease (Wu et al., 2020), temporal lobe epilepsy (Zheng et al., 2022), and Parkinson disease (Qian et al., 2020; Zhou et al., 2020). Based on these remarkable discoveries, a cure‐all, regeneration‐based, therapeutic strategy is on the horizon for neural injuries, neurological diseases or symptoms.…”

p38 MAPK Endogenous Inhibition Improves Neurological Deficits in Global Cerebral Ischemia/Reperfusion Mice

“…Gene therapy for stroke is a promising and new treatment method for patients who are ineffective in drug treatment and unable to undergo surgery. It has been shown that gene therapy can help self-repair vision after cerebral ischemia injury; it proves that gene therapy is an effective and promising treatment for cerebral ischemia injury in a mouse model [ 66 ].…”

Enhancing NeuroD1 Expression to Convert Lineage-Traced Astrocytes into Neurons

“…We were alerted by the apparent lower conversion efficiency in the lineage-tracing Aldh1l1-CreERT2 mice compared to our previous results obtained in WT mice Ge et al, 2020;Guo et al, 2014;Liu et al, 2020;Puls et al, 2020;Tang et al, 2021;Wu et al, 2020;Zhang et al, 2020;Zheng et al, 2022). To increase the conversion rate in lineage-traced astrocytes without increasing AAV titre, we employed AAV vector with CMV enhancer (GFAPCE::NeuroD1) engineered to increase NeuroD1 expression in astrocytes as shown in Figure 3.…”

“…An alternative approach for regenerating new neurons is to convert internal glial cells into functional neurons (Barker et al, 2018;Bocchi et al, 2021;Lei et al, 2019;Li and Chen, 2016;Qian and Fu, 2021). Such direct in vivo reprogramming technology has been demonstrated repeatedly in my lab (Chen et al, 2020;Ge et al, 2020;Guo et al, 2014;Liu et al, 2020;Puls et al, 2020;Wu et al, 2020;Xiang et al, 2021;Zheng et al, 2022), as well as in many other labs around the world (Gascon et al, 2016;Heinrich et al, 2014;Jiang et al, 2021;Lentini et al, 2021;Liu et al, 2015;Niu et al, 2015;Niu et al, 2013;Qian et al, 2020;Su et al, 2014;Tang et al, 2021;Torper et al, 2015). In vivo reprogramming is not only achieved in the CNS and but also in other organ systems such as pancreas, heart, and liver (Larouche and Aguilar, 2019;Nam et al, 2014;Qian et al, 2012;Rezvani et al, 2016;Song et al, 2016;Zhou et al, 2008).…”