2015 — 2021 |
Pasca, Sergiu |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Gaining Insight Into Psychiatric Disease by Engineering Piece by Piece the Human Brain in Vitro.
? DESCRIPTION (provided by applicant): Most therapeutic advances in medicine are concentrated in accessible tissue disorders, such as coagulopathies and oncology, where diseased tissue can be directly examined and manipulated. To systematically and comprehensively study neuropsychiatric disorders at the molecular level, direct access to neurons and glia from patients is indispensable. Here, we propose to use an innovative approach that leverages a novel tridimensional (3D) neural differentiation method of human pluripotent stem cells (hiPSC), to separately generate neural spheroids that include either excitatory neurons of the cortex (subpallium-like) or ventral telencephalic interneurons (subpallium-like). Using regional-specific genetic enhancers, we plan to fluorescently label pallium-like and subpallium-like forebrain spheroids and fuse them to engineer mixed neural structures in suspension. Using a series of assays in two-region human forebrain spheroids engineered from hiPSC derived from patients with genetic forms of schizophrenia and autism spectrum disorders (ASD), we aim to identify the key pathophysiological processes underlying these disorders.
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0.915 |
2017 — 2021 |
Pasca, Sergiu |
U01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Cross Modal Integration of Molecular and Physiological Networks in Asd (2/2)
Genetic approaches have been successful in identifying causal genetic factors, both common and rare, that contribute to risk for autism spectrum disorder (ASD), providing a crucial starting point for mechanistic neurobiological investigations. However, moving towards an integrated mechanistic understanding of ASD at a molecular, cellular, and circuit level faces substantial challenges, such as extreme genetic heterogeneity and the lack of causal frameworks with which to connect different levels of analysis of nervous system function in model systems or patients. Nearly a decade ago, we reasoned that gene and protein networks would provide an organizing framework for understanding heterogeneous psychiatric disease genetic risk in a unified context and inform disease modeling; indeed there is now substantial evidence supporting convergence of major effect risk genes during mid-fetal cortical development. Furthermore, related functional genomic studies, including in those with a major gene form of ASD (dup)15q11-13, show shared patterns of transcriptional and chromatin dysregulation in post-mortem ASD brain, further supporting biological convergence. Where and how this occurs, and what biological mechanism(s) it reflects is not known. To address this, we propose an ambitious project that addresses several major challenges in establishing causal linkages between genetic risk and CNS structure and function in ASD. The work proposed in this multi-PI U01 involves a team of four principal investigators and co-investigators from UCLA and Stanford with the expertise necessary to perform this work using state of the art methodologies, ranging from developing and characterizing in vitro models of human brain development, stem cells, physiology, genomics, physics, and behavior. Through close collaboration, we will develop and analyze in vitro human stem cell based models that are differentiated from induced pluripotent stem cells and assembled into organized 3D brain cultures called human forebrain spheroids (hFS). These hFS contain the major cell classes of the developing forebrain, including progenitors, radial glia, cortical interneurons, glutamatergic neurons, and non-reactive astrocytes, and form functional synapses. We will model the effects of six major effect ASD risk loci in hFS with molecular, genomic, and physiological analyses to assess convergence at each level of analysis. We will also conduct comparisons of physiology using three rodent models based on the same genes modeled in vitro with the aim of integrating phenotypes to develop predictive models and compare with in vivo rodent models. We will analyze the relationship of molecular alterations and basic cellular and synaptic features with potential emergent or dynamic network features in control-derived hFS and compare these features with hFS harboring ASD risk mutations and test a subset of causal relationships based on network model predictions. Completion of these aims will lead to a more clear understanding of the power and limitations of model systems and computational models, while uncovering potential areas of convergence in different genetic forms of ASD.
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0.915 |
2018 — 2021 |
Pasca, Sergiu |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Role of L-Type Calcium Channels in Human Interneuron Migration and Integration
ABSTRACT Gene variants and mutations in voltage-gated L-type calcium channels (LTCCs) genes are among the most replicable findings in genetic studies of autism spectrum disorders, bipolar disorder and schizophrenia. The mechanisms by which these genetic events lead to disease are not currently known. Previous work has indicated that these LTCss play a critical role in cortical interneuron migration and functional integration into cortical circuits. Here, we propose to leverage a novel tridimensional (3D) neural differentiation of human induced pluripotent stem cells (hiPSC) that we developed to generate functional neural spheroids resembling the laminated excitatory cerebral cortex (pallial spheroids) and, separately, subpallial spheroids giving rise to cortical interneurons. Using state-of-the-art live imaging, transcriptional profiling, genetic-engineering, pharmacology and electrophysiological methods, we plan to assemble two-region human forebrain structures (pallial-excitatory/subpallial-inhibitory) and investigate the functional role of LTCCs mutations in migration and functional synaptic integration of cortical interneurons. !
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0.915 |