Abstract
A key objective of stem cell biology is to create physiologically relevant cells suitable for modeling disease pathologies in vitro. Much progress towards this goal has been made in the area of motor neuron (MN) disease through the development of methods to direct spinal MN formation from both embryonic and induced pluripotent stem cells. Previous studies have characterized these neurons with respect to their molecular and intrinsic functional properties. However, the synaptic activity of stem cell-derived MNs remains less well defined. In this study, we report the development of low-density co-culture conditions that encourage the formation of active neuromuscular synapses between stem cell-derived MNs and muscle cells in vitro. Fluorescence microscopy reveals the expression of numerous synaptic proteins at these contacts, while dual patch clamp recording detects both spontaneous and multi-quantal evoked synaptic responses similar to those observed in vivo. Together, these findings demonstrate that stem cell-derived MNs innervate muscle cells in a functionally relevant manner. This dual recording approach further offers a sensitive and quantitative assay platform to probe disorders of synaptic dysfunction associated with MN disease.
Highlights
All motor functions from locomotion to respiration depend on the communication between motor neurons (MNs) in the spinal cord and muscle cells in different regions of the body
Recent studies have further shown that MNs can be produced from induced pluripotent stem cells (IPSC) including those derived from amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy (SMA) patients [10,11,12], and through transcription factor-mediated reprogramming of fibroblasts [13]
Our study provides critical evidence that ESC-derived MNs can exhibit robust synaptic communication with muscle cells under simplified in vitro culture conditions
Summary
All motor functions from locomotion to respiration depend on the communication between motor neurons (MNs) in the spinal cord and muscle cells in different regions of the body. This vital activity is susceptible to many neurodegenerative diseases, most notably amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy (SMA), resulting in MN dysfunction and death [1,2]. A remaining challenge, is to establish methods to evaluate the function of normal and diseased MNs obtained from these sources in a physiologically relevant setting
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