Modeling Normal Human Neuron Development and Autism Spectrum Disorders Using Stem Cells

Human FXS forebrain neurons have reduced neurite initiation and outgrowth. A, B. Low magnification images of human forebrain neurospheres from control A) and Fragile X Syndrome (FXS) B) iPSCs cultured on laminin and labeled by immunocytochemistry for neural-specific βIII-Tubulin (red) and F-actin using fluorescent phalloidin (green). Note the decreased number and length of projections in FXS neurospheres as compared to control neurospheres after 2 DIV. Also note that many βIII-Tubulin positive neurons migrate away from the FXS neurosphere (arrows)

Modulation of mTOR-dependent protein synthesis by TSC1/TSC2.  Sequential phosphorylation events occur downstream of axon guidance cue receptors to activate or inhibit the synthesis of proteins that regulate axon outgrowth.  Many regulatory phospho-proteins can be detected by Western blot or ICC.

Current Open Questions

 

    1.Do human neurons use local protein synthesis in growth cones to regulate axon guidance?

 

    2.Do axons of human neurons with mutations in the Tuberous Sclerosis (TSC) related genes, TSC1 and TSC2, exhibit proper axon guidance behaviors in vitro?

 

    3.Is local protein synthesis defective in growth cones of neurons derived from iPSCs of patients with TSC?

 

    4.Do axons of human neurons with mutations in the Fragile X syndrome (FXS) gene exhibit proper axon guidance behaviors in vitro?

 

    5.Is local protein synthesis defective in growth cones of neurons derived from iPSCs of patients with FXS?

The discovery of human embryonic stem cells (hESCs) and the generation of human induced pluripotent stem cells (hiPSCs) has dramatically expanded our ability to understand the normal development of the human nervous system and the molecular basis of neurodevelopmental disorders.  In particular, reprogramming somatic cells into hiPSCs allows generation patient-specific lines of hiPSCs that carry disease causing mutations.  One of the most powerful features of hESCs and hiPSCs is their ability to be differentiated into a wide variety of cell types, including neurons.  Specific classes of neurons can be generated in abundance for use in biochemical, molecular and cell biological assays of development and function.  We are currently using human forebrain neurons, retinal ganglion cells and motorneurons to address both basic and disease related questions concerning the development of the human nervous system.