Directed Differentiation and Functional Maturation of Cortical Interneurons from Human Embryonic Stem Cells

Maroof, Asif; Keros, Sotirios; Tyson, Jennifer A.; Ying, Shui‐Wang; Ganat, Yosif; Merkle, Florian T.; Liu, Becky; Goulburn, Adam L; Stanley, Edouard G.; Widmer, HR; Eggan, Kevin; Goldstein, Peter A.; Anderson, Stewart A.; Studer, Lorenz

doi:10.1016/j.stem.2013.04.008

Cited by 531 publications

(595 citation statements)

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“…We observed no differences in neuronal induction efficiency and yield among LiR, LiNR, or unaffected patient-derived hiPSCs, all of which showed similar expression of neuronal markers [Tau, βIII-tubulin (Tuj1), MAP2, vGLUT, GABA] and produced neural progenitor cells (NPCs) and electrophysiologically active (17) glutamatergic (vGLUT + ) and GABAergic (GABA + ) neurons that initially expressed markers consistent with a dorsal anterior forebrain cortical phenotype (SI Appendix, Figs. S2 E-L and S3 A-J) (17)(18)(19)(20). For our in vitro studies, we elected to preserve the distinction made by clinicians between LiR and LiNR patients (5)(6)(7)(8) and to probe lithium's protein targets within LiR BPD neurons (SI Appendix, Figs.…”

Section: Resultsmentioning

confidence: 99%

“…S1) were reprogrammed to hiPSCs via nonintegrating episomalmediated (42), lentivirus-mediated (1), or retrovirus-mediated (19) gene transfer, characterized (43), and differentiated to NPCs and cortical interneurons, as per our routine and as previously described (17)(18)(19)(20). Protein isolation, 2D-DIGE and SILAC, Western blotting, and coimmunoprecipitation were performed as described previously (17,28).…”

Section: Methodsmentioning

confidence: 99%

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Probing the lithium-response pathway in hiPSCs implicates the phosphoregulatory set-point for a cytoskeletal modulator in bipolar pathogenesis

Tobe

Crain

Winquist

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

136

121

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The molecular pathogenesis of bipolar disorder (BPD) is poorly understood. Using human-induced pluripotent stem cells (hiPSCs) to unravel such mechanisms in polygenic diseases is generally challenging. However, hiPSCs from BPD patients responsive to lithium offered unique opportunities to discern lithium's target and hence gain molecular insight into BPD. By profiling the proteomics of BDP-hiPSCderived neurons, we found that lithium alters the phosphorylation state of collapsin response mediator protein-2 (CRMP2). Active nonphosphorylated CRMP2, which binds cytoskeleton, is present throughout the neuron; inactive phosphorylated CRMP2, which dissociates from cytoskeleton, exits dendritic spines. CRMP2 elimination yields aberrant dendritogenesis with diminished spine density and lost lithium responsiveness (LiR). The "set-point" for the ratio of pCRMP2:CRMP2 is elevated uniquely in hiPSC-derived neurons from LiR BPD patients, but not with other psychiatric (including lithium-nonresponsive BPD) and neurological disorders. Lithium (and other pathway modulators) lowers pCRMP2, increasing spine area and density. Human BPD brains show similarly elevated ratios and diminished spine densities; lithium therapy normalizes the ratios and spines. Consistent with such "spine-opathies," human LiR BPD neurons with abnormal ratios evince abnormally steep slopes for calcium flux; lithium normalizes both. Behaviorally, transgenic mice that reproduce lithium's postulated site-of-action in dephosphorylating CRMP2 emulate LiR in BPD. These data suggest that the "lithium response pathway" in BPD governs CRMP2's phosphorylation, which regulates cytoskeletal organization, particularly in spines, modulating neural networks. Aberrations in the posttranslational regulation of this developmentally critical molecule may underlie LiR BPD pathogenesis. Instructively, examining the proteomic profile in hiPSCs of a functional agent-even one whose mechanism-of-action is unknownmight reveal otherwise inscrutable intracellular pathogenic pathways. have proven valuable for studying the molecular pathology of monogenic diseases, one of the technique's greatest challenges has been to offer similar insights into the molecular pathogenesis of polygenic, multifactorial disorders for which the underlying pathophysiology is unknown. The struggle has been to go beyond phenotypic description to discerning underlying molecular mechanisms. Neuropsychiatric illnesses are a prototype for such complex conditions (1-3). They are difficult to model not only because of the likelihood of polygenic influences, but also because of the subjectivity with which these diseases must often be diagnosed, the empirical fashion with which drugs are prescribed, and the heterogeneity of patient response. Of such maladies, bipolar disorder

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Probing the lithium-response pathway in hiPSCs implicates the phosphoregulatory set-point for a cytoskeletal modulator in bipolar pathogenesis

Tobe

Crain

Winquist

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

136

121

View full text Add to dashboard Cite

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“…Hence, generation of MGE-and LGE-like progenitors from hESCs and hiPSCs through directed differentiation approaches is critical. The methods to generate MGE-like cells from hESCs and hiPSCs are already available Maroof et al, 2013;Liu et al, 2013aLiu et al, , 2013bNicholas et al, 2013). Fig.…”

Section: Discussionmentioning

confidence: 99%

Potential of GABA-ergic cell therapy for schizophrenia, neuropathic pain, and Alzheimer׳s and Parkinson׳s diseases

Shetty

Bates

2016

Brain Research

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Several neurological and psychiatric disorders present hyperexcitability of neurons in specific regions of the brain or spinal cord, partly because of some loss and/or dysfunction of gammaamino butyric acid positive (GABA-ergic) inhibitory interneurons. Strategies that enhance inhibitory neurotransmission in the affected brain regions may therefore ease several or most deficits linked to these disorders. This perception has incited a huge interest in testing the efficacy of GABA-ergic interneuron cell grafting into regions of the brain or spinal cord exhibiting hyperexcitability, dearth of GABA-ergic interneurons or impaired inhibitory neurotransmission, using preclinical models of neurological and psychiatric disorders. Interneuron progenitors from the embryonic ventral telencephalon capable of differentiating into diverse subclasses of interneurons have particularly received much consideration because of their ability for dispersion, migration and integration with the host neural circuitry after grafting. The goal of this review is to discuss the premise, scope and advancement of GABA-ergic cell therapy for easing neurological deficits in preclinical models of schizophrenia, chronic neuropathic pain, Alzheimer's disease and Parkinson's disease. As grafting studies in these prototypes have so far utilized either primary cells from the embryonic medial and lateral ganglionic eminences or neural progenitor cells expanded from these eminences as donor material, the proficiency of these cell types is highlighted. Moreover, future studies that are essential prior to considering the possible clinical application of these cells for the above neurological conditions are proposed. Particularly, the need for grafting studies utilizing medial ganglionic eminence-like progenitors generated from human pluripotent stem cells via directed differentiation approaches or somatic cells through direct reprogramming methods are emphasized.

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“…To characterize human interneuron development, we developed a protocol to generate MGElike cells in vitro based on previous methods (Maroof et al, 2013). Telencephalic neural induction of hESCs was initiated with dual SMAD inhibition, combined with a smallmolecule inhibitor of the WNT pathway ( Figure 1A) (Chambers et al, 2009).…”

Section: In Vitro Generation Of Mge Progenitors and Neuronsmentioning

confidence: 99%

Single-Cell Profiling of an In Vitro Model of Human Interneuron Development Reveals Temporal Dynamics of Cell Type Production and Maturation

Close¹,

Yao²,

Miller³

et al. 2017

Neuron

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SummaryGABAergic interneurons are essential for neural circuit function and their loss or dysfunction is implicated in human neuropsychiatric disease. In vitro methods for interneuron generation hold promise for studying human cellular and functional properties and ultimately therapeutic cell replacement. We describe here a protocol for generating cortical interneurons from hESCs and analyze the properties and maturation timecourse of cell types using single-cell RNAseq. We find that the cell types produced mimic in vivo temporal patterns of neuron and glial production, with immature progenitors and neurons observed early and mature cortical neurons and glial cell types produced late. By comparing the transcriptomes of immature interneurons to more mature neurons, we identified genes important for human interneuron differentiation. Many of these genes were previously implicated in neurodevelopmental and neuropsychiatric disorders. Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. HHS Public Access

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Directed Differentiation and Functional Maturation of Cortical Interneurons from Human Embryonic Stem Cells

Cited by 531 publications

References 54 publications

Probing the lithium-response pathway in hiPSCs implicates the phosphoregulatory set-point for a cytoskeletal modulator in bipolar pathogenesis

Probing the lithium-response pathway in hiPSCs implicates the phosphoregulatory set-point for a cytoskeletal modulator in bipolar pathogenesis

Potential of GABA-ergic cell therapy for schizophrenia, neuropathic pain, and Alzheimer׳s and Parkinson׳s diseases

Single-Cell Profiling of an In Vitro Model of Human Interneuron Development Reveals Temporal Dynamics of Cell Type Production and Maturation

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