Project summary Cortical excitatory and inhibitory (E/I) imbalance in the brain has been commonly observed in psychiatric disorders. However, neither circuit mechanisms or cell types responsible for the E/I imbalance in the disease are clearly specified. We focus on layer 6b (L6b) neurons as a strong candidate for cortical inhibitory gain controller. Although L6b neurons, also called subplate neurons, are previously known to show abnormal distribution in postmortem brain tissues of schizophrenia (SZ) and autism-spectrum disorders (ASD), the anatomy and physiology of the neurons are poorly understood. Our optogenetics-based slice electrophysiology and in vivo two-photon axonal calcium imaging demonstrate that these neurons persist in adult brains, project axons toward up to layer 1 by crossing all six cortical layers, innervate inhibitory interneurons, form functional synaptic connection within cortical circuit, and respond to external sensory stimuli. More interestingly, our preliminary data found that the number of neurons in layer 6b (L6b) was decreased in a mouse model of a CNV, 16p11.2 duplication which is known to show deficient GABAergic synaptic transmission and highly associated with SZ and ASD. These data suggest that L6b neurons may play a key role in cortical gain control via feed-forward inhibition and the dysregulation of these neurons may cause psychological and behavioral symptoms. We will determine this innovative hypothesis, L6b neuron as a novel inhibitory gain controller in the neocortex and they are responsible for cortical imbalance in psychiatric disorders. We will combine multiple approaches including optogenetics-based multiple whole-cell patch clamp recordings, in vivo calcium imaging, and pharmacogenetic manipulation of the L6b neurons to determine the roles of the L6b neurons and their implications in pathological conditions. Our proposed work will provide a new conceptual understanding of dysregulated cortical inhibition in psychiatric disorders, by presenting L6b neuron as a key cortical gain controller.