Opioid receptors are central to the development of tools that can be used to manage and fight against the opioid crisis that is prevalent in North America. They are part of a large protein family called G-protein-Coupled Receptors (GPCRs), which are the most therapeutically targeted receptors within the human body. Once activated, the receptors lead to the activation of multiple different signaling pathways such the -arrestin and G-protein signaling pathways. The -Arrestin pathway is usually associated with the side effects of opioid analgesics. An allosteric site that binds to sodium was identified within the delta-opioid receptor (DOR). Previous studies have found that the sodium cavity can regulate the activation of different signaling pathways and thus act at the functional selectivity level. Our lab has identified a subset of small molecules targeting this cavity. This finding supports the druggability of this site and thus opens the door for the development of a novel pharmacological entity to control opioid receptor activities. This thesis focuses on the characterization of the sodium cavity by performing structure activity relationship (SAR) studies on the delta opioid receptor with three allosteric modulators: MIA, HMA and zoniporide. We report that, through site-directed mutagenesis and functional studies, mutations in the allosteric sodium site has an impact on the receptor functionalities including ligand recognition, efficacy and also allosterism by small molecules; however, the mutations do not prevent the binding of the allosteric modulators to the receptor. We also report the development of a novel biomedical tool that can be used to study the recruitment of the G-protein subtypes as well as the arrestin subtypes. Our data suggest it is possible to design drugs that will target the sodium pocket and this site has a major role within DOR and could be used to design novel modulators with unique pharmacological properties. My work will serve as a platform to study other members of the opioid receptor family and for the future rational design of novel modulators.

Opioids are the most powerful pain relievers currently known (Huang & Mallet, 2013) and opioid abuse is considered a significant public health crisis (Pezalla, Rozen, Erensen, Haddox, & Mayne, 2017). Older adults face elevated risks for opioid abuse given the unique pain reported by many late adults (Wilder-Smith, 2012) and the potential for overuse of opioids is especially high given the growing population of older adults in the U.S. (Centers for Disease Control and Prevention, 2003). This project aims to explore the trends of opioid use and abuse by older adults and how to manage this growing epidemic by examining a multitude of options including the marketing of opioids, different addiction treatments, and other solutions.

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Opioid-linked overdose death rates have reached unprecedented levels in the United States. The growing incidence of Opioid Use Disorder (OUD) is concomitant with elevated rates of OUD in women during pregnancy and through parturition. Neonates born to mothers with active OUD can develop opioid dependence in utero and display various signs of postnatal withdrawal, a condition termed Neonatal Opioid Withdrawal Syndrome (NOWS). Common symptomatic features of NOWS include sleep disturbances, low birth weight, altered heart and respiratory rates, increased irritability, high-pitched crying, feeding difficulties. Many of these symptomatic presentations are driven by dysregulated function of the autonomic nervous system and hyperirritability of the sympathetic nervous system. Given the increasing incidence of NOWS, there is an alarming lack of knowledge regarding the long-term effects of perinatal opioid exposure on behavioral and neurodevelopmental outcomes. Murine models provide efficient means to understand the neurobiological adaptations impacted by opioid exposure during perinatal neurodevelopment that drive long-term effects on cognitive, social, affective, and reward-related behaviors. We describe a rodent model of third-trimester-equivalent opioid exposure which produces replicable, opioid withdrawal-related phenotypes including robust thermal hyperalgesia and altered ultrasonic vocalization (USV) profiles. We present results from two drug regimens of the model, differing in the schedule of opioid administration (once or twice daily injections of morphine from postnatal day (P) 1-14; 15.0 mg/kg). Beyond hyperalgesia and altered USV profiles, both drug regimens lead to weight loss. Furthermore, both models resulted in transcriptional adaptations within brain regions relevant to opioid dependence and withdrawal. Twice-daily exposure resulted in sex-specific changes in metabolic gene expression in the brainstem, while once-daily exposure resulted in down-regulation of genes related to myelin and dopaminergic circuitry development in the nucleus accumbens. We found minimal evidence for behavioral consequences associated with once-daily morphine exposure during adulthood; there were no significant effects of perinatal morphine on cognitive, reward-related, or fear learning tasks. This could potentially indicate compensatory mechanisms that mitigate the adverse effects of third trimester-equivalent morphine exposure over time. Lastly, we identified epigenetic mechanisms potentially driving dysregulation of normal development within the central nervous system following pre-natal opioid exposure in humans. We analyzed placental samples from pregnancies with opioid exposure and identified gene networks containing altered DNA methylation patterns. Notably, we found enrichment within the ‘integral component of the plasma membrane’ and ‘synapse assembly’ functional networks, indicating potential effects of prenatal opioid exposure on neural connectivity and transmission. Together, the transcriptional adaptations identified in rodent brain tissue and the epigenetic modifications identified in human placental tissue provide novel mechanistic insight as to how perinatal opioid exposure impacts neural and fetal development.

The COVID-19 pandemic adversely affected the lives of most Americans. People with Substance Use Disorder (SUD) were particularly vulnerable to the negative effects brought on by the pandemic. This study explored the increase in deaths due to opioid overdose during the pandemic exacerbated by factors such as increased stress, decrease in treatment options due to social distancing requirements and facility closures, social isolation, and an increase in spare time. Access to treatment for opioid use disorder (OUD) was interrupted by the measures meant to mitigate the spread of COVID-19. Through a systematic review of current literature, it was demonstrated that existing patients were able to maintain access to care, while few new patients were able to initiate treatment. Telehealth proved to be a vital means of assuring PWUD were able to access life-saving treatment amid a pandemic. Further research is needed to determine whether SUD treatment measures during the COVID-19 pandemic warrants changing the policies long term.

Background The lack of methadone pharmacokinetic data in children and neonates restrains dosing to achieve the target concentration in these populations. A minimum effective analgesic concentration of methadone in opioid naïve adults is 0.058 mg·l-1, while no withdrawal symptoms were observed in neonates suffering opioid withdrawal if plasma concentrations of methadone were above 0.06 mg·l-1. The racemate of methadone which is commonly used in pediatric and anesthetic care is metabolized to 2-ethylidine-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) and 2-ethyl-5-methyl-3,3-diphenylpyrroline (EMDP). Methods Data from four studies (age 33-week PMA-15 years) were pooled (n = 56) for compartment analysis using nonlinear mixed effects modeling. Parameter estimates were standardized to a 70-kg person using an allometric model approach. Investigation was made of the racemate and metabolite (EDDP and EMDP) dispositions. In addition, neonatal data (n = 7) allowed further study of R- and S-enantiomer pharmacokinetics. Results A three-compartment linear disposition model best described the observed time-concentration profiles with additional compartments for metabolites. Population parameter estimates (between-subject variability) were central volume (V1) 21.5 (29%) l·70 kg-1, peripheral volumes of distribution V2 75.1 (23%) l·70 kg-1 and V3 484 (8%) l·70 kg -1, clearance (CL) 9.45 (11%)l·h-1·70 kg-1, and intercompartment clearances Q2 325 (21%) l·h -1·70 kg-1 and Q3 136 (14%) l·h -1·70 kg-1. EDDP formation clearance was 9.1 (11%) l·h-1·70 kg-1, formation clearance of EMDP from EDDP 7.4 (63%)l·h-1·70 kg-1, elimination clearance of EDDP was 40.9 (26%) l·h-1·70 kg-1 and the rate constant for intermediate compartments 2.17 (43%) h-1. Conclusions Current pharmacokinetic parameter estimates in children and neonates are similar to those reported in adults. There was no clearance maturation with age. Neonatal enantiomer clearances were similar to those described in adults. A regimen of 0.2 mg·kg-1 per 8 h in neonates achieves a target concentration of 0.06 mg·l-1 within 36 h. Infusion, rather than intermittent dosing, should be considered if this target is to be achieved in older children after cardiac surgery.

The Fecal Fermentation Profile of Twins and Infants with Opioid Exposure Dawson Brown-Ezell Michelle Johnson PhD, RD, LDN W. Andrew Clark PhD, RD Amy Wahlquist, PhD Introduction: The gut microbiome is believed to have a significant impact on health throughout the lifespan, and the influence of infant nutrition and other environmental factors are of particular interest in its development. The aim of this research project was to learn more about the microbiome and short chain fatty acid (SCFA) composition of toddlers of differing weights, considering birth history, environment, and diet. In East Tennessee, opioid misuse is a growing issue, and a number of participants in this study were exposed in utero. We also hoped to identify related effects on infant’s SCFA composition. Finally, it has been concluded that twins share a variety of traits, but much about their microbiome is unknown. With several pairs of twins in the sample, we aimed to identify any associations with SCFAs in this group. Methods: With informed consent, the child’s history was obtained, including age, birth length and weight, delivery type (C-section or vaginal), and feeding method (breast, bottle fed, or both). The child’s current weight, height, and BMI %ile were determined. Caregivers completed the 90-question Block Questionnaire for Ages 2-7 Kids food frequency questionnaire, and results were analyzed by Berkeley Analytics Inc (dba NutritionQuest). Participant-provided stool samples were freeze-dried and ground, and SCFAs were extracted and analyzed by content and concentration. Data analysis was generated using SAS software, Version 9.4 of the SAS System, Copyright © 2013 SAS Institute Inc. Results: Nine SCFAs were measured in duplicate, and the concentrations averaged. Statistical analysis included comparisons of SCFAs related to factors including weight status, infant feeding modality, twin status, and intrauterine drug exposure, and significance determined with a p value < 0.05. Results did not identify significant differences in individual SCFA concentrations between obese and non-obese toddlers, however concentrations of isobutyrate, isovaleric acid, and octanoic acid were greater in toddlers who were formula fed as infants versus toddlers who were breastfed, and those fed a combination of breastmilk, and formula. Analysis further revealed a higher mean concentration of caproic and propionic acid in twin subjects. Of particular interest, toddlers with a history of opioid exposure had higher mean concentrations of isovaleric and octanoic acids, but less isocaproic acid when compared to those who were not drug exposed. Further analysis will help determine if these findings may be related to nutrient intake, in particular dietary fiber intake.

Death caused by opioid abuse has increased in recent years, and women in the state of Alaska have been significantly impacted by this opioid crisis. Previous researchers have indicated a possible connection between opioid use and sub-clinical PTSD criteria. The purpose of this grounded theory study was to develop a theory regarding the presence of PTSD diagnostic criteria in this population, to identify patterns in past traumas and other life stressors, and to investigate coping strategies in 43 Alaskan women who sought treatment for opioid use. Archived data in the form of therapy notes were analyzed using grounded theory techniques such as coding information, categorizing the codes, and comparing patterns that were discovered to previous research. Hyperarousal was the most commonly reported criterion of PTSD, becoming the basis of the theory that it plays an important role connecting lived experiences and coping in these women. The most commonly reported experiences included substance use by parents, parental divorce, domestic violence, employment issues, mental health issues, partner substance use, and legal issues. Coping strategies included medicating, seeking support from nonprofessionals, and compliance. Recommendations for applying findings included using trauma-informed care, and implementing therapeutic workplaces, to reinforce abstinence with the ability to work as part of treatment. This data can be used for social change by improving assessment and treatment through addressing what might not have previously been considered trauma in these clients. Thus, providers may provide more effective treatment for opioid use disorders, and implement strategies to help prepare clients for longer term success and reduced prevalence of relapse.

Prescription opioid abuse in the United States is an alarming health issue. In 2015, approximately 2 million people abused prescription opioids, and 12 million individuals misused their prescription opioid pain relievers. The percentage of individuals who died as a result of opiate abuse increased from 22% in 2013 to 76% in 2014. The purpose of this project was to evaluate the influence of an inner-city drug treatment (DTBF) program on opioid users' behavior. The practice question addressed whether knowledge of signs and symptoms of opioid withdrawal obtained from the DTBF program resulted in a significant behavioral change in opioid use in 45 adults ages 18 to 25. The Centers for Disease Control and Prevention framework for program evaluation was used to guide the study. Data were collected using a pretest and posttest with the Clinical Opiate Withdrawal Scale (COWS) over a 6-month period. Results of t-test analysis indicated a significant change in drug use (p = .000). Recommendations to clinic administrators included encouraging all staff to use the COWS in screening individuals and observing them at each clinic visit. The implication of this study for social change is that findings may be used to reduce drug abuse and misuse among prescription opioid users.

Prescription drug abuse has been a growing problem in Indiana and around the nation for almost two decades. In recent years, prescription drug overdoses have pushed drug poisonings ahead of motor vehicle crashes as the leading cause of injury death. However, deaths due to overdoses of prescription drugs are only the tip of the iceberg when it comes to the much larger problem of abuse. This study has characterized prescription drug abuse in Indiana and taken an in-depth look at how it is and can be addressed both through organizational policies and state legislation. Opioid painkillers such as hydrocodone, oxycodone, and methadone are the most commonly abused prescription drugs, and most of these prescriptions are written by primary care physicians. Because more than 70% of Indiana’s family medicine residents will remain in the state to practice medicine following the conclusion of their residencies, it is worthwhile to take a look at how these residents are being educated during their training. St. Vincent’s Family Medicine Residency program in Indianapolis is one of several residency programs in Indiana training their residents on best practices of prescribing controlled substances. A review of residents’ prescribing patterns before and after training on the subject went into effect showed significant reductions in the number of opioid painkillers being prescribed, and showed the same reductions for alprazolam, a benzodiazepine anxiolytic.