3D cultured human medium spiny neurons functionally integrate and rescue motor deficits in Huntington’s disease mice
Yuting Mei, Yuan Xu, Xinyue Zhang, Ban Feng, Yingying Zhou, Qian Cheng, Yuan Li, Xingsheng Peng, Mengnan Wu, Lianshun Xie, Lei Xiao, Wenhao Zhou, Yuejun Chen, Man Xiong
Yuting Mei, Yuan Xu, Xinyue Zhang, Ban Feng, Yingying Zhou, Qian Cheng, Yuan Li, Xingsheng Peng, Mengnan Wu, Lianshun Xie, Lei Xiao, Wenhao Zhou, Yuejun Chen, Man Xiong
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Research Article Cell biology

3D cultured human medium spiny neurons functionally integrate and rescue motor deficits in Huntington’s disease mice

Abstract

Dysfunction of striatal medium spiny neurons (MSNs) is implicated in several neurological disorders, including Huntington’s disease (HD). Despite progress in characterizing MSN pathology in HD, mechanisms underlying MSN susceptibility remain unknown, driving the need for MSNs derived from human pluripotent stem cells (hPSCs), especially subtypes in research and therapy. Here, we established a scalable 3D-default culture system to produce striatal MSNs efficiently from hPSCs by activation of the endogenous sonic hedgehog (SHH) pathway. These cells expressed canonical markers of striatal progenitors and dopamine D1 (D1) and dopamine D2 (D2) MSNs and presented dynamic specification and transcriptional signatures that closely resemble endogenous MSNs at single-cell resolution, both in vitro and post-transplantation in HD mice with quinolinic acid (QA) lesions. Grafted human cells survived and matured into D1-/D2-like MSNs and projected axons to endogenous targets including globus pallidus externus, globus pallidus internus, and substantia nigra pars reticulata to reconstruct the basal ganglia pathways. Functionally, they displayed spontaneous synaptic currents, received regulation from host cortex and thalamus, and were modulated by dopamine to either enhance or reduce neuronal excitability, similar to the endogenous D1-/D2-MSNs, subsequently improving behavior in QA-lesioned HD mice. Our study presents a method for generating authentic MSNs, providing a reliable cell source for HD cell therapy, mechanistic studies, and drug screening.

Authors

Yuting Mei, Yuan Xu, Xinyue Zhang, Ban Feng, Yingying Zhou, Qian Cheng, Yuan Li, Xingsheng Peng, Mengnan Wu, Lianshun Xie, Lei Xiao, Wenhao Zhou, Yuejun Chen, Man Xiong

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Figure 7

Grafted striatal neuron integrated into host neural circuit and alleviated motor defects of HD mice.

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Grafted striatal neuron integrated into host neural circuit and alleviat...
(A) Experimental design for cell transplantation (transpl.) and behavioral tests. (B–E) Schematic of electrophysiological recordings and representative sIPSC/sEPSC traces of grafted neurons 5MPT, with quantified amplitude and frequency. Data were analyzed by Student’s t test. (G) The strategy for whole-cell patch-clamp recording of synaptic regulation from host cortex. (H) Representative image showing host cortical GFP+ fibers around mCherry+ grafted neurons. (I) Typical traces showing light-evoked EPSCs in grafted neurons 5MPT (top) and the subsequent blockade by CNQX (bottom). (J) Quantified amplitudes of optogenetically induced EPSCs (n = 9 cells from 3 mice). (K) The strategy for whole-cell patch-clamp recording of synaptic regulation from host thalamus. (L) Representative image of host thalamic tdTomato+ fibers around GFP+ grafted neurons. (M and F) Light-evoked EPSCs in grafted neurons (blocked by CNQX) with amplitude quantification (n = 8 cells from 3 mice). (N and O) Representative open-field movement traces and total distance covered by WT, ACSF-injected, and striatal neuron-grafted HD model animals at 0MPT, 2MPT, and 5MPT. (P) The rotarod test showing time of latency to fall in different groups at 0MPT, 2MPT, and 5MPT. n = 8–10 mice for HD+ASCF, 8–13 mice for HD+Cell, and 6–13 mice for WT. Two-way ANOVA was used for comparison between different groups. Data are presented as the mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.