Project Summary. Epilepsy is the second most prevalent neurological disorder, affecting approximately 2 million people in the United States. While many patients achieve satisfactory seizure control with pharmacotherapy, a significant proportion (20-40%) have medically intractable seizures. For these patients, identification of novel methods for seizure control is a high priority. One such method may be to harness endogenous seizure suppressive circuits in the brain. Because seizures may have multiple or unknown initiation sites, focal stimulation approaches (e.g., deep brain stimulation [DBS]) that can control seizures originating in diverse brain networks are highly desirable. The circuitry of the basal ganglia has received particular attention in this regard. While four decades of research have elaborated the specific nuclei that can contribute to this effect, the precise pathways, projections, and circuit architecture remain obscure. Building on our findings in the prior project period, and a robust literature in preclinical epilepsy research, we aim to define the upstream regulators (basal ganglia direct and indirect pathways; Aim 1), midbrain local circuits (deep layers of superior colliculus [DLSC]-pedunculopontine nucleus [PPN] interactions; Aim 2), and PPN projection targets (Aim 3) that mediate the antiseizure effects of the basal ganglia. Our goal is to develop a comprehensive macro-network map and to determine how, at the circuit level, manipulations in this circuit lead to broad- spectrum resistance to seizures. We expect that a deeper understanding of this circuitry will lead to novel interventions to control seizures. In Specific Aim 1, we will monitor and modulate activity in the basal ganglia direct and indirect pathways through a combination of optogenetics, in vivo single-unit recording, and fiber photometry in models of temporal lobe epilepsy (TLE) and absence epilepsy. In Specific Aim 2, we will define the midbrain (DLSC-PPN) circuits engaged by limbic and absence seizure activity and test the functional relationship between DLSC and PPN in controlling seizures. We will address this by combining in vivo single- unit recording, slice electrophysiology, and optogenetics. In Specific Aim 3, we will define PPN output targets that mediate protection against limbic and absence seizures, focusing on the mediodorsal and reticular thalamic nucleus, using temporal lobe and absence epilepsy models. Together the proposed studies will provide proof of principle for optogenetic modulation of seizures in diverse networks from a single circuit. Moreover, this approach allows us to examine previously untestable hypotheses about the connections that mediate basal ganglia seizure control. Finally, the proposed experiments aim to uncover the circuit and neurotransmitter mechanism by which focal manipulations in motor control regions (i.e., SNpr/DLSC) translate into brain-wide changes in excitability.