Neural circuits are the underlying functional units of the human brain. By receiving glutamatergic (Glut) inputs through cortico-striatal pathway, the striatum acts as an integrative hub to coordinate multiple higher-order behavior and cognition. Dysfunction of the striatum and the associated neural circuitry development have been implicated in the pathogenesis of multiple neurodevelopmental disorders e.g., schizophrenia. Despite the functional importance, studies of such long-distance human cortical-subcortical network development and connectivity have been significantly hindered due to lack of suitable microphysiological platforms. A major unresolved hurdle in current human cells-based assays is that in vitro cultures weakly recapitulate the key biology of neural microphysiological system, especially the long-distance projections in both two and three dimensions (2D/3D). In this grant, we propose to fill this critical gap, by reconstructing human cortico-striatal circuits on-a- chip to recapitulate and monitor long-range brain circuitry development and connectivity in vitro, in response to PAR-20-082. We will reconstruct human cortico-striatal circuits by developing a novel 2D/3D microfluidic microelectrode arrays (MEA) chip together with the co-culture of human pluripotent stem cells (hPSCs)-derived region-specific neurons or brain organoids. Coupling with MEA allows us to monitor brain circuit dynamics in a high-throughput manner. Further, our innovative implementation of microfluidic channels and arrays of surface and probe electrodes in 3D configurations enables resemblance of 3D brain circuits for high-order brain function studies. We hypothesize that human cortico-striatal circuits on a microfluidic-MEA chip can reconstitute striatal synchrony, a key striatal physiology which is absent in unconnected striatal neurons, and the reconstructed 3D neural circuits between cortical and striatal organoids can resemble high-order brain function e.g., brain waves like in vivo. We will reconstruct human cortico-striatal circuits by using our novel microfluidic MEA chip together with the co-culture of hPSCs-derived cortical and striatal neurons in Aim 1. We will monitor axon projections and neural network dynamics during circuitry development and determine whether striatal synchrony is driven by cortical Glut inputs by pharmacological manipulation of Glut transmission. In Aim 2, we will develop a 3D microfluidic MEA chip with microchannels and microelectrodes integrated in 3D configurations. We will reconstruct 3D cortico-striatal circuits by assembling cortical and striatal organoids on-a-chip. We expect 3D brain circuits on-a-chip approach will resemble brain waves (a.k.a. large-scale neural oscillation) like human brains. This proposal presents a novel approach to reconstitute well-defined long-range human circuit in vitro. Our model can be benchmarked against existing human and rodent in vitro brain circuitry systems and exceed state-of-the-art b...