PROJECT SUMMARY Prenatal exposure to cigarette smoke increases the risk of sudden infant death syndrome as well as developmental deficits in the brain that persist into adulthood. Sensory impairments have been observed, notably in the auditory system, following prenatal cigarette exposure. Due to the increasing popularity of e-cigarettes, the need for research into prenatal exposure to nicotine alone is becoming increasingly important. The critical period of mouse auditory development occurs after birth, allowing for perinatal nicotine exposure to accurately model prenatal nicotine exposure on auditory system phenotypes. In PNE models, disrupted glutamatergic signaling in the auditory cortex and impaired temporal processing in an auditory startle test has been reported. However, the cellular mechanisms whereby perinatal nicotine exposure (PNE) impairs auditory development and central auditory processing are currently unknown. In the auditory system, cholinergic signaling is necessary for peripheral and central auditory processes. Recent work demonstrates the importance of nicotinic acetylcholine receptors in signal-in-noise detection in the medial nucleus of the trapezoid body (MNTB) of the auditory brainstem. The alpha 7 nicotinic acetylcholine receptor (α7 nAChR), essential for glutamatergic synapse development in the hippocampus and cortex, is highly expressed in the MNTB during early postnatal development. Utilizing the large glutamatergic Calyx of Held synapse of the MNTB, the proposed studies aim to investigate the effect of PNE on structural and functional development of the calyx terminal, MNTB synapse, and central auditory processing. The central hypothesis is that PNE increases α7 nAChR expression and its chronic activation impairs glutamatergic synapse development in the MNTB resulting in central auditory deficits. Due to the crucial role of MNTB in binaural processing, it is important to examine how the MNTB is affected by PNE during auditory development to understand auditory processing disorders in children prenatally exposed to nicotine. Aim 1 examines developmental expression of nAChRs at the calyx-MNTB synapse and the effect of PNE on nAChR expression using patch-clamp electrophysiology, immunohistochemistry, and western blot. Aim 2 investigates the developmental impact of PNE on the structure and function of calyx of Held synapse. Direct presynaptic recordings of the calyx terminal will measure vesicular glutamate release followed by 3D reconstruction of the calyx to quantify structural development. Patch clamp recordings of the MNTB neuron with afferent fiber stimulation will be done to measure glutamate-mediated currents, and immunohistochemistry will visualize glutamate receptors in the calyx synapse. Aim 3 tests auditory processing following PNE using in vivo auditory tests. The proposed aims will reveal cellular and circuit-level mechanisms underlying developmental nicotine exposure-induced auditory deficits that will be us...