PROJECT SUMMARY/ABSTRACT Individuals who overdose on opioids die from respiratory depression due to activation of the mu opioid receptors, but the precise mechanism remains elusive. Respiratory output is generated and controlled by nuclei in the brainstem, including the Kölliker-Fuse (KF) in the pons and the Bӧtzinger Complex, preBötzinger Complex and rostral ventral respiratory group in the ventrolateral medulla. All of these structures express postsynaptic and/or presynaptic mu opioid receptors and therefore are vulnerable to opioid modulation. The work of this proposal focuses on understanding if and how opioids modulate respiratory-controlling KF neurons that project to respiratory nuclei in the ventrolateral medulla, namely the Bӧtzinger Complex, preBötzinger Complex and rostral ventral respiratory group. Opioids could modulate KF neurons by hyperpolarization and/or inhibition of neurotransmitter release from axon terminals. Recently, it was found that mu opioid receptor agonists hyperpolarize a population of KF neurons, but it is unknown how these opioid-sensitive neurons integrate into the pontomedullary respiratory circuitry. I will determine if these opioid-sensitive KF neurons project to respiratory nuclei (Aim 1), and if KF projections to respiratory nuclei are vulnerable to presynaptic opioid modulation (Aim 2). I hypothesize that opioid-sensitive KF neurons project to and synapse onto neurons within the Bӧtzinger Complex, preBötzinger Complex and/or rostral ventral respiratory group, thereby changing respiratory output. In Aim 1, the projection target of opioid-sensitive KF neurons will be determined using brain-slice electrophysiology and retrograde tracing. In Aim 2, brain-slice electrophysiology, in combination with optogenetics, will be used to determine if opioids inhibit neurotransmitter release from KF axon terminals. Results from this project will 1) identify respiratory neurocircuitry vulnerable to opioid modulation and 2) provide insight into the mechanism by which opioids change respiratory output. My work will be supervised by Drs. Levitt (sponsor) and Mitchell (co- sponsor), two NIH-funded investigators with extensive publications in their respective fields. Together, we developed a training plan that will help me learn new experimental techniques important to neuropharmacology and neuroscience research, develop my writing and oral dissemination skills, and foster my teaching and career development. This work and Drs. Levitt and Mitchell’s mentorship will give me the necessary skills to pursue a competitive post-doctoral fellowship and eventual faculty position.