title: Large-scale generation of human iPSC-derived neural stem cells/early neural progenitor cells and their neuronal differentiation.
abstract: Induced pluripotent stem cell (iPSC)-based technologies offer an unprecedented opportunity to perform high-throughput screening of novel drugs for neurological and neurodegenerative diseases. Such screenings require a robust and scalable method for generating large numbers of mature, differentiated neuronal cells. Currently available methods based on differentiation of embryoid bodies (EBs) or directed differentiation of adherent culture systems are either expensive or are not scalable. We developed a protocol for large-scale generation of neuronal stem cells (NSCs)/early neural progenitor cells (eNPCs) and their differentiation into neurons. Our scalable protocol allows robust and cost-effective generation of NSCs/eNPCs from iPSCs. Following culture in neurobasal medium supplemented with B27 and BDNF, NSCs/eNPCs differentiate predominantly into vesicular glutamate transporter 1 (VGLUT1) positive neurons. Targeted mass spectrometry analysis demonstrates that iPSC-derived neurons express ligand-gated channels and other synaptic proteins and whole-cell patch-clamp experiments indicate that these channels are functional. The robust and cost-effective differentiation protocol described here for large-scale generation of NSCs/eNPCs and their differentiation into neurons paves the way for automated high-throughput screening of drugs for neurological and neurodegenerative diseases.
authors: a Department of Psychiatry ; Western Psychiatric Institute and Clinic ; University of Pittsburgh School of Medicine ; Pittsburgh , PA USA.
authors: Leonardo
authors: L
authors: D'Aiuto
authors: Yun
authors: Y
authors: Zhi
authors: Dhanjit
authors: D
authors: Kumar Das
authors: Madeleine R
authors: MR
authors: Wilcox
authors: Jon W
authors: JW
authors: Johnson
authors: Lora
authors: L
authors: McClain
authors: Matthew L
authors: ML
authors: MacDonald
authors: Roberto
authors: R
authors: Di Maio
authors: Mark E
authors: ME
authors: Schurdak
authors: Paolo
authors: P
authors: Piazza
authors: Luigi
authors: L
authors: Viggiano
authors: Robert
authors: R
authors: Sweet
authors: Paul R
authors: PR
authors: Kinchington
authors: Ayantika G
authors: AG
authors: Bhattacharjee
authors: Robert
authors: R
authors: Yolken
authors: Vishwajit L
authors: VL
authors: Nimgaonka
authors: Vishwajit L
authors: VL
authors: Nimgaonkar
mesh_terms: D061249:Batch Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation / Q000502:physiology; D049109:Cell Proliferation / Q000502:physiology; D002478:Cells, Cultured; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology* / Q000502:physiology; D058953:Neural Stem Cells / Q000166:cytology* / Q000502:physiology; D009474:Neurons / Q000166:cytology* / Q000502:physiology*
keywords: high-throughput screening; induced pluripotent stem cells (iPSCs); neural progenitor cells; neural stem cells; neuronal differentiation
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