Pharmacological Regulation of Neural Circuit Formation in hIPSC-Derived Neurons and 'Mini-Brains'

Emerging evidence suggests that altered neural connectivity , particularly at the level of synaptic connections , contributes to the pathology of many neurodevelopmental and neurodegenerative diseases. For instance , post-mortem Autistic patient brain samples have increased numbers of excitatory to inhibitory synaptic connections , referred to as an E/I imbalance [42]. Contrastingly , post-mortem brain samples from patients diagnosed with Alzheimer's disease have decreased numbers of synaptic connections [42]. In order to understand the mechanisms that underlie the formation of these synaptic circuits , we develop 3-D human cortical organoids ('mini-brains') from human-induced pluripotent stem cells (hIPSCs). Previous research demonstrates that rearrangements of the actomyosin cytoskeleton drive neural circuit formation , in particular the development and maturation of actin-enriched spines at excitatory synapses. This thesis work investigates how pharmacological regulation of actomyosin activity affects neuronal connectivity during neurite formation in 2-D and excitatory synapse formation in 3-D 'mini-brains'. The Rho-Kinase (ROCK) inhibitor , Y-27632 , both inhibits non-muscle myosin II (NM-II) and leads to a corresponding increase in Rac-driven actin polymerization. In 2-D , Y-27632 promotes neurite formation. Specifically , Y-27632 increases the number , length , and branching of neurites in hIPSC-derived neurons. Furthermore , Y-27632 increases neurite persistence , while decreasing neurite protrusion and retraction rates. However , in 3-D , acute Y-27632 treatment increases excitatory synapse area , consistent with an increase in Rac-driven actin polymerization [39]. Thus , Y-27632 increases both neurite outgrowth and excitatory synapse formation and may serve as a potential therapeutic for neurodegenerative diseases associated with synapse loss such as Alzheimer's disease. This study demonstrates the need for physiologically-relevant brain models , such as 3-D cortical organoids , to assess the impact of drug therapies on developing neural circuits to potentially treat neurodevelopmental and neurodegenerative disorders