Maturation of Spinal Motor Neurons Derived from Human Embryonic Stem Cells

Hynek Wichterle,Lorenz Studer,Gist F Croft,Tomonori Takazawa,Mackenzie W Amoroso,Shawn Hochman,Amy B Macdermott

doi:10.1371/journal.pone.0040154

Hynek Wichterle, Lorenz Studer + Show 5 more

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https://doi.org/10.1371/journal.pone.0040154

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Abstract

Our understanding of motor neuron biology in humans is derived mainly from investigation of human postmortem tissue and more indirectly from live animal models such as rodents. Thus generation of motor neurons from human embryonic stem cells and human induced pluripotent stem cells is an important new approach to model motor neuron function. To be useful models of human motor neuron function, cells generated in vitro should develop mature properties that are the hallmarks of motor neurons in vivo such as elaborated neuronal processes and mature electrophysiological characteristics. Here we have investigated changes in morphological and electrophysiological properties associated with maturation of neurons differentiated from human embryonic stem cells expressing GFP driven by a motor neuron specific reporter (Hb9::GFP) in culture. We observed maturation in cellular morphology seen as more complex neurite outgrowth and increased soma area over time. Electrophysiological changes included decreasing input resistance and increasing action potential firing frequency over 13 days in vitro. Furthermore, these human embryonic stem cell derived motor neurons acquired two physiological characteristics that are thought to underpin motor neuron integrated function in motor circuits; spike frequency adaptation and rebound action potential firing. These findings show that human embryonic stem cell derived motor neurons develop functional characteristics typical of spinal motor neurons in vivo and suggest that they are a relevant and useful platform for studying motor neuron development and function and for modeling motor neuron diseases.

Highlights

Motor neurons are the final connecting link between the central nervous system and skeletal muscles
If hESMNs are to be useful for modeling human motor neuron development and function, it is important that they acquire mature functional characteristics similar to motor neurons in vivo
We provide evidence that hESMNs follow a time dependent course of maturation in vitro that can be measured by changes in morphology and electrophysiology

Summary

Introduction

Motor neurons are the final connecting link between the central nervous system and skeletal muscles. They are typically large neurons with extensive dendritic fields, primarily located in the ventral horn of the spinal cord. In humans, these neurons are essentially inaccessible for study. Most of our understanding of motor neuron development and function is based on studies in a variety of mammalian model systems such as cats and rodents, extensively reviewed in [1]. Membrane properties change developmentally with a decrease in input resistance, increasingly hyperpolarized resting membrane potentials and appearance of a repetitive firing response to a sustained depolarizing stimulus [5,7,8,9]

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