PROJECT SUMMARY Millions of children undergo general anesthesia each year, and animal and human studies have indicated that exposure to anesthesia at an early age can impact neuronal development, leading to behavioral and learning impairments that manifest later in childhood and adolescence. However, little is known about the methods which can be used to evaluate these impairments. Our preliminary and published data suggest that changes in regional resting state functional MRI (rsfMRI) can reflect the status of local neuronal networks and can have a predictive value for anesthesia-induced learning deficiency. Regional rsfMRI depends on the regulation of local brain tissue oxygen. Based on published and preliminary data we hypothesize that brain tissue oxygen and regional rsfMRI regulation are modulated by GABAergic neurons (interneurons) but neonatal anesthesia disrupts the development of this modulation and produces a specific signature on the rsfMRI signal. In Aim 1 we will determine the mechanisms responsible for regional rsfMRI and brain tissue oxygen regulation under normal conditions and after neonatal anesthesia exposure by using brain tissue oxygen, BOLD fMRI, electrophysiological recordings, local pharmacological and optogenetic approach in adults. In Aim 2 we will determine how the development of regional rsfMRI and brain tissue oxygen fluctuations is affected by neonatal anesthesia exposure and find time points when rsfMRI can predict the future behavioral deficiency. Behavioral tests, local rsfMRI, brain tissue oxygen, and neuronal activity will be recorded in across multiple time points after neonatal anesthesia exposure and in the control group. The results of the proposed work will provide a clear picture of how the regulation of brain tissue oxygen is affected by neonatal anesthesia exposure and will offer strong translational value of resting-state fMRI to predict learning deficiency.