ABSTRACT Layer 2/3 projection neurons (PNs) are critical nodes in association cortico-cortical (Ac-c) circuits, and likely targets of neurodevelopmental disorder (NDD) pathology. In mouse models of 22q11.2 Deletion Syndrome (22q11DS), which in humans confers risk for multiple clinically defined NDDs, Ac-c circuits are under- connected, in part due to diminished layer 2/3 PN growth accompanied by mitochondrial oxidative stress. We will use mouse 22q11DS genetic models to test the hypothesis that Ac-c circuit pathology in NDD reflects selective, mitochondria-dependent developmental dysregulation of layer 2/3 PN long-range connections, leading to disordered synaptic organization, disconnection, and dysfunction. We combine genetic, physiological, transcriptomic, proteomic, and behavioral approaches to assess how 22q11 deletion alters identity and function of layer 2/3 PNs to result in individually variable behavior and circuit pathology. In Specific Aim 1 we determine whether excitatory (E) vs. inhibitory (I) synaptic sites on layer 2/3 PNs that make long-range connections are altered by NDD pathogenesis, including that due to 22q11 deletion, leading to Ac-c circuit disconnection and dysfunction. Our data will provide foundational insight into cellular, physiological and transcriptional identities of typical layer 2/3 Ac-c vs. 5/6 PNs, as well as consequences of NDD pathology for E/I balance in circuits defined by long-range Ac-c connections. In Specific Aim 2, we assess mitochondrial function in developing layer 2/3 PNs as they generate pre- and post-synaptic domains to define mature microcircuits. We analyze how NDD pathogenesis due to 22q11 deletion disrupts developing and mature mitochondrial regulation of Ac-c connectivity. Our data will provide fundamental characterization of mitochondria deployment, bioenergetics, and proteomic identities in typical developing and mature layer 2/3 vs. 5/6 PNs, including mitochondria at layer 2/3 PN synapses. In parallel, we will define how mitochondrial identity and function is compromised by 22q11 deletion. In Specific Aim 3, we analyze how mitochondrial antioxidant defense influences behaviors that rely upon long range Ac-c connections made by layer 2/3 PNs. We assess individual variability in cognitive, mnemonic, sensory gating and social behaviors. Our data will resolve whether a specific dimension of mitochondrial function—clearance of reactive oxygen species generated by oxidative phosphorylation—varies individually to disrupt Ac-c circuit dependent behaviors and circuits. Data collected from these Specific Aims will provide a framework for understanding optimal mitochondrial regulation of Ac-c circuit development and function and its disruption in NDDs. We will resolve contributions of mitochondrial function and antioxidant defense to 22q11 deletion-mediated Ac-c disconnection and related behavioral deficits. Our work will define whether emerging therapies, including antioxidants and other metabolic int...