Device-free, long-term monitoring of opioid circuit activity in individual mice exposed to fentanyl

Abstract While the incidence of Opioid Use Disorder (OUD) continues at epidemic levels, a necessary step in the development of improved OUD therapeutics is to understand the adapting molecular architecture and functional dynamics neural circuits responding to opioids. Despite extensive research to date, it remains unclear how opioidergic circuits in various brain regions modulate neural activities and opioid gene expression patterns across stages of the addiction cycle (e.g., drug naïve, acute exposure, chronic exposure, and abstinence) within the same animals. To better capture the molecular and activity underpinnings of OUD, our collaborative team of bioengineers and circuit physiologists will develop a new approach for long-term monitoring of opioid-induced changes in neural population activity and opioid receptor gene expression with a simple blood test, called Released Markers of Activity—RMAs—that translocate from neurons into the peripheral blood stream. Thus, the breakthrough potential of RMAs for OUD discoveries are illustrated by their device-free and within-subjects measurement features with around-the-clock, weeks-long monitoring, as well as modular use for both neural activity and genetic alterations as provided by our novel viral promoters that give genetic access to active populations and opioid receptor-expressing cell-types. In Aim 1, we will clone our 2nd generation “erasable” RMAs (eRMAs; protease cleavable variants for higher signal-to-noise detection amplification across time) under an activity-dependent viral promoter system to measure increased and/or decrease neural activity in two key brain regions for OUD—before, during and after abstinence from morphine and fentanyl exposure, within the same animal. In Aim 2, we will clone eRMAs under a new opioid receptor promoter system that would permit a new class of long-term gene expression measurement related to the up- or down-regulation of the mu-opioid receptor following chronic morphine and fentanyl exposure. Across both aims we will target this new blood-based monitoring system to key brain regions implicated in OUD: ventral tegmental area, nucleus accumbens, and central amygdala. The RMAs for OUD approach will enable us to understand how opioid exposure influences neural population functions and opioid receptor expression that are related to OUD-related behavior in mice. Beyond our lab, this innovative and team-driven project will result in the creation of an easy-to-implement and fast, device-free blood monitoring system for researchers to accelerate the pace of research in opioid addiction and future development of new OUD treatments. In principle, RMAs for OUD are not limited to mice but any genetically intractable model system, such as rats, human iPSC brain organoids, pigs, and non-human primates, which we will explore after successful completion of the CEBRA R21 goal: To develop Release Markers of Activity for detecting neural activities and opioid gene expression across the OUD cycle in individual mice.