Illuminating Neuropeptide Modulation of Substance Abuse Withdrawal: Novel Photochemical Tools for Precision Control

About This Grant

Project Summary/Abstract Methamphetamine use disorder afflicts millions of Americans and precipitates a debilitating withdrawal syndrome marked by hypodopaminergia, anxiety, and somatic distress, yet our ability to interrogate its neuropeptidergic underpinnings is hindered by a lack of versatile, high precision chemical probes in neuroscience. We hypothesize that corticotropin releasing factor (CRF) mediates negative affect and stress responsivity, neuropeptide Y (NPY) promotes resilience within reward circuits, and orexin (ORX) governs motivational and somatic outputs during withdrawal. To test this, we will engineer a modular library of cyclic, photolabile caging scaffolds that render CRF, NPY, and ORX both inert and proteolysis resistant (enhancing in vivo stability and bioavailability), thereby enabling multiplexed interrogation of multiple peptide systems reducing the need to generate or maintain multiple mouse lines to modulate each peptidergic system. Each probe will undergo a rigorous validation pipeline: biophysical photolysis and binding assays to quantify release kinetics and receptor engagement, cell culture studies to confirm receptor specificity and downstream signaling, and acute brain slice experiments in both naïve and methamphetamine-withdrawn mice (focused uncaging on genetically identified cell types with whole cell electrophysiology and two photon calcium imaging) to verify functional efficacy and altered peptide responsiveness. We will then deploy genetically encoded peptide sensors to dynamically monitor CRF in the bed nucleus of the stria terminalis (BNST), NPY in the ventral tegmental area (VTA), and ORX in the nucleus accumbens (NAc) both before methamphetamine exposure and during withdrawal in behaving mice, an approach that will reveal how peptide activity in discrete circuits drives specific withdrawal features. Separate experiments using time locked uncaging manipulations will test causality by modulating peptide signal and assessing effects on withdrawal phenotypes. Behavioral endpoints will include quantitative somatic withdrawal scoring (for example, jumps and tremors), validated anxiety assays (elevated plus maze and open field), and reward related paradigms (sucrose preference and conditioned place preference), analyzed in both male and female mice to uncover sex specific peptide mechanisms. Because CRF, NPY, and ORX, along with their respective brain circuits, are conserved across mammals, these mechanistic insights will be directly relevant to human neurobiology and withdrawal physiology. Leveraging these findings will pave the way to rationally design peptide-based agonists and antagonists as prototype therapeutics to restore neuropeptide balance and ameliorate withdrawal symptoms. Finally, we will disseminate our chemical reagents, protocols, and data through open access repositories, empowering the broader neuroscience community to apply these spatiotemporal control strategies across diverse research domains and accelerate development of targeted treatments for substance use disorders.