| Description |
Human intelligence and higher cognitive functions arise from the unique and highly organized structure of the human brain. With 86 billion interconnected neurons, the complexity of the human brain surpasses almost all observed structures in the known universe. Nevertheless, all these complexities unfold from relatively simple embryonic structures, following widely conserved developmental programs, such as neural differentiation and synapse formation. During recent evolutionary history, human-specific genetic alterations of neurodevelopmental programs have given rise to the dramatic expansion and rewiring of the cerebral cortex. These changes are thought to underlie the rise of human-specific intelligence and cognitive functions. Moreover, genetic mutations of key developmental regulators often cause severe cognitive changes in neuropsychiatric disorders, such as autism and schizophrenia. Thus, to understand the mechanisms of human intelligence and cognitive functions, we need a comprehensive knowledge of both the conserved and human-specific programs of neural development. Such knowledge is crucial for understanding the mechanisms of neuropsychiatric disorders and may provide hints for the future paths of human brain evolution. Animal models have provided invaluable insights into the conserved mechanisms of neural development. However, the study of the human-specific neurodevelopmental programs has been challenging due to the lack of experimental models. In this dissertation, I focus on using neurons derived from human pluripotent stem cells (hPSCs) to study human cortical neural differentiation and the formation of neuronal connectivity. In Chapter 2, I used cerebral organoids to study the neural differentiation of the human cerebral cortex. Cerebral organoids are a novel 3D model of cortex-like structure (minibrains) derived from hPSCs. Our lab developed a novel method for producing cerebral organoids with greatly improved reproducibility. Using high-throughput single-cell RNA sequencing, I surveyed the cell type composition of our organoid model during neural differentiation. The data show that our cerebral organoids contain multiple pools of neural progenitors, separate lineages of cortical excitatory and inhibitory neurons, as well as multiple glial cell types. Using patch-clamp electrophysiology, I discovered functional subtypes of mature and immature spiking neurons, and observed disease-associated intrinsic excitability deficits in organoids that lack SHANK3, an autism and intellectual disability gene. To understand the genetic programs underlying cell type specification in organoids, I performed computational mapping of developmental trajectory and uncovered genetic factors involved in different neuronal lineages. In Chapter 3, I developed novel methods for studying connectivity neuronal deficits associated with neurodevelopmental disorders in hPSC-derived human neurons. I focused on Phelan-McDermid Syndrome (PMDS), a genetic disorder associated with autism and intellectual disability, and created a CRISPR/Cas9-engineered human hPSC cell line that harbors PMDS-associated deletion of SHANK3. To characterize neuronal connectivity deficits, I transplanted control and mutant human neurons into the developing mouse cerebral cortex and performed trans-synaptic tracing. The tracing revealed both local and long-range synaptic inputs to transplanted human neurons, validating the method's ability to map brain-wide synaptic connectivity. To understand how the functional properties of synapses are affected by the deletion of SHANK3, I performed patch-clamp electrophysiology recordings, and observed significantly reduced AMPA/NMDA current ratios in the excitatory synapses of SHANK3-deficient neurons. |
