Research demonstrates that substance-use disorder (SUD) affects one’s decision-making and reward processing resulting in impaired goal-directed behaviors. However, our understanding of the intrinsic neurobiology underlying reward-seeking behaviors is lacking. Addressing this gap in knowledge requires approaches measuring behavioral phenotypes associated with persistent reward-seeking. My first introduction to SUD-related research was at the Undergraduate Research Symposium my freshman year. While attending, I realized the majority of presentations I found compelling were associated with Dr. Michael Bruchas’ laboratory, a neuropharmacology lab specializing in unraveling the intricacies of G-coupled protein receptor (GPCR) systems within the contexts of stress, depression, SUD, and pain. In particular, I was drawn to the idea of elucidating the neuropharmacological mechanisms underlying complex medical conditions like SUDs; an interest cultivated by my own personal experiences with family members that suffered from SUD. I then promptly contacted the lab and joined their team as their newest undergraduate researcher. Over the years I have engaged in various projects within Dr. Bruchas’ lab. My first project explored the neuropharmacological underpinnings that drive stress-induced binge eating. Currently, I work with my postdoctoral mentor, Dr. Kasey Girven designing behavioral paradigms mirroring cue-induced drug relapse and elucidating neuropeptidergic targets implicated in the modulation of drug-seeking behaviors.
In 2020, more than 9.5 million people misused opioid prescriptions and 2.7 million individuals were diagnosed with an opioid use disorder1. This resulted in 80,411 people losing their lives from opioid overdose in 2021 alone2. Despite research into the underlying pharmacological mechanisms generating the analgesic and euphoric effects associated with opioid use, the neurobiology driving the persistent urge to seek drugs and how environmental cues exacerbate these symptoms remains largely unexplored. Therefore, a rigorously validated fentanyl self-administration task is essential to accurately model the behavioral patterns observed in individuals with substance abuse disorder. As well as developing a task that can be combined with techniques that have fine spatio-temporal resolution to measure neuronal ensemble activity like with two photon single cell imaging.
Operant conditioning is a paradigm that utilizes reinforcements and punishments to respectively strengthen or weaken the likelihood of a particular behavior recurring. Within the paradigm of operant conditioning is a delivery schedule known as fixed ratio one (FR1). FR1 denotes that every desired action is reciprocated by one positive reinforcer on a 1:1 ratio.  During each session, wild-type (WT) mice undergo the FR1 task by way of entering their nose into a small indentation in the experimental box, colloquially known as a “nose poke”. Following a 2-second delay, a cue in the form of a light is elicited. After six seconds, the cue ceases and a sipper containing a 10 ug/ml fentanyl solution is extended for 10 seconds of unrestricted access to the drug (Figure 1a). Both our water-restricted male and female cohorts exhibited a correlated increase in active nose pokes and licks per session with a plateau of inactive nose pokes (Figure 1b,1c). This indicates a sufficient understanding of the task and proficient execution. As sessions advance raster plots used to measure the individual subject’s licking frequency indicate a rising proportion of licks occurring within the initial 30-40 minutes (Figure 1d). This attribute denotes that the subjects are learning the task and are eager to nosepoke for fentanyl delivery but have their own self-monitored stopping point, similar to what can be observed in humans with SUD. I have contributed to this experiment by performing this oral fentanyl self-administration task across multiple cohorts comprised of both male and female sexes, and learned how to analyze their behavioral responses, such as: lick frequency, nose poking, and individual fentanyl consumption to confirm their consistent participation in the task.
While monitoring licks and active nose pokes provides a reliable indicator of reward-seeking, sufficient fentanyl intoxication must be measured behaviorally. These behavioral assays include tail flick to measure changes in nociception, the open field locomotor assay, and the elevated zero maze anxiety assay to quantify exploratory behaviors.  In our current preliminary stages, a singular female cohort (n=4) has underwent these assays immediately following a 30-40 minute period of FR1 fentanyl self-administration, noting this increment of the task is when a majority of the consumption occurs. All behavioral assays were also conducted and analyzed by myself using a personally novel software known as Ethovision. This software allowed me to quantify their exploration in the arenas as well and generate a movement heat map using techniques designed to track the mouse throughout the session. 

In the upcoming phases, our plan involves extending the self-administration sessions and initiating behavioral assays for our current male fentanyl cohort while initiating new cohorts to enhance the statistical power of our fentanyl self-administration studies. Furthermore, we intend to delineate quantifiable behavioral markers of fentanyl intoxication by comparing these cohorts to a control cohort undergoing the same trials with water instead of fentanyl. Presently, we have discerned a trend towards less licking activity compared to the fentanyl cohorts already undergoing operant conditioning. In the upcoming months, these additional groups of mice will undergo the same behavioral assays as previously described alongside newly implemented protocols for examining rectal body temperature and opiate blood levels. Novel techniques such as these not only refine my animal handling skills but also allow me to apply my certified phlebotomic knowledge, which, I initially acquired for humans but eagerly anticipate adapting to mice. The autonomy granted for this project and its subsequent phases has been immensely beneficial and will continue to contribute to my growth as a researcher. Throughout this process, I've gained the ability to troubleshoot various assays and technical aspects of experiments with increasing ease. I've also developed skills in independently managing, analyzing, and presenting data—integral experiences shaping me into a self-sufficient and confident graduate researcher.