Biology subjects

Brennan, M.

Publications and source records attributed to Brennan, M..

At least 19 recordsLinked to original sources

bioRxiv2026

Replication of Host St6galnac1-Microbiome Association Across Distinct Experimental Conditions in Heterogeneous Stock Rats

The gut microbiome is shaped by both environmental and host genetic factors. Identifying specific host loci is essential for understanding the heritability of microbiome-associated traits. The gene St6galnac1 encodes a sialyltransferase involved in mucin O-glycosylation. Tonnele et al. (2025) identified significant associations between a locus containing St6galnac1 gene, and the abundance of genus Paraprevotella using cecal samples collected from fasted animals. This association was replicated across three cohorts and paralleled with association found in humans between a functionally related sialyltransferase gene ST6GAL1 and Paraprevotella. Given the sensitivity of microbiome analyses to factors including sampling site, feeding status, and analytical framework, independent replication of these findings is important and it is also important to know if the association generalizes across other conditions or is highly specific to certain environments. Here, we performed a replication analysis using stool (not cecal) samples from 174 HS rats that were not fasted prior to collection. Stool samples underwent 16S rRNA gene sequencing, and microbial community structure was analyzed using compositionally aware methods. The rats were genotyped at the top SNP that was associated with Paraprevotella in the Tonnele et al. (2025) study. The association between genotype and Paraprevotella abundance was tested using a linear mixed model accounting for sex, cohort, and genetic relatedness. We successfully replicated (p=0.015) St6galnac1 - Paraprevotella association. These findings support the robustness of the St6galnac1 - Paraprevotella association across different sampling strategies and experimental conditions.

genomics↗

bioRxiv2026

Addiction-Like Severity Predicts Prolonged Oxycodone Withdrawal-Induced Allodynia in Genetically Diverse Rats

Opioid withdrawal is associated with heightened pain sensitivity, including allodynia. Although opioid-induced allodynia is well-documented in humans and animal models, the relationship between the severity of opioid withdrawal-induced allodynia and individual addiction-like behaviors remains poorly understood. To address this gap, Heterogeneous Stock rats underwent long access (12 hours/day) intravenous oxycodone self-administration, followed by measurement of mechanical sensitivity at six timepoints across three weeks of abstinence. Rats were stratified by an Addiction Index derived from individual differences in the escalation of oxycodone intake, motivation to consume oxycodone, tolerance to oxycodones analgesic effects, and acute withdrawal-induced mechanical pain sensitivity. Here, we show that oxycodone withdrawal induces significant and prolonged allodynia for up to three weeks, with High Addiction Index rats exhibiting greater intensity and longer duration of pain sensitivity than Low Addiction Index rats. Results remained consistent even when excluding allodynia from the Addiction Index, highlighting the robustness of the association between addiction-like severity and protracted allodynia. Linear regression associations revealed that self-administration behaviors, particularly oxycodone intake escalation and motivation to seek oxycodone, predicted subsequent withdrawal-induced allodynia severity. These findings demonstrate that greater addiction-like severity is associated with more intense and prolonged withdrawal-induced pain, supporting mechanical allodynia as a marker of addiction severity. These results motivate future work to define the mechanisms linking addiction severity to protracted opioid withdrawal-induced pain, with the goal of informing targeted clinical interventions for individuals most susceptible to severe abstinence-related allodynia.

pharmacology and toxicology↗

bioRxiv2026

Spatial transcriptomics for gene discovery identifies Slc13a5 as a modulator of bone mechanoadaptation

Bone is a dynamic tissue that continuously adapts its structure in response to mechanical loading, an essential process for maintaining skeletal health. However, this adaptive capacity declines with aging, contributing to increased fragility and fracture risk. Developing therapeutic strategies that preserve or restore bone mechanoadaptation in patients with increased bone fragility requires identifying key molecular regulators of this process. We applied spatial transcriptomics (GeoMx, NanoString) to characterize gene expression changes induced by mechanical loading in the murine tibia, focusing on periosteal and bone compartments in regions under tension and compression. Spatial data were validated and cross-compared with previously published bulk RNA-seq and laser-capture microdissection datasets, identifying a set of 12 genes consistently regulated by loading across independent platforms and laboratories. As part of a functional analysis, we selected Slc13a5, a citrate transporter implicated in bone mineralization and metabolism. Conditional deletion of Slc13a5 in osteolineage cells using Osteocalcin-Cre significantly increased the loading-induced mineralizing surface in tensile regions compared with Cre- Slc13a5fl/fl littermates. In addition, Slc13a5 cKO mice exhibited lower resorption around the neutral axis after loading compared to controls. Together, these findings identify Slc13a5 as a regulator of bone adaptation in regions experiencing low mechanical stimulation and suggest it as a potential therapeutic target for conditions characterized by impaired mechanoadaptive responses. This study highlights spatial transcriptomics as a powerful gene discovery framework for bone, enabling identification of novel targets to understand mechanisms and develop therapies. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=181 SRC="FIGDIR/small/711126v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@14fc0e9org.highwire.dtl.DTLVardef@1023a16org.highwire.dtl.DTLVardef@133aa0eorg.highwire.dtl.DTLVardef@e19c86_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2026

Regulatory T cells modulate bone marrow stromal cell osteogenesis by activating ROCK-myosin axis and cell contractility

Bone regeneration depends not only on the intrinsic signaling in bone marrow-derived mesenchymal stromal cells (BMSC), but also on immune cell-derived cues. Regulatory T cells (Treg) are emerging as immune regulators of tissue repair, yet their direct impact on BMSC osteogenesis remains unexplored. Here, we showed that direct human BMSC-Treg co-culture induced more pronounced osteogenic gene expression and cytokine modulation in BMSC than indirect co-culture. This contact-dependent interaction temporally enhanced BMSC osteogenic gene expression, alkaline phosphatase activity and matrix mineralization by interacting with ROCK-myosin signaling pathway. Exploratory proteomic profiling further revealed enrichment of cytoskeletal, mechanosensitive and osteogenic regulators in BMSC after direct Treg co-culture. Pharmacological inhibition of cell contractility impaired osteogenesis yet Treg co-culture partially restored cytoskeletal integrity and differentiation. These findings identify novel mechanisms behind Treg as direct modulators of BMSC mechanobiology and osteogenesis, highlighting their therapeutic potential in bone regeneration.

cell biology↗

bioRxiv2026

Loss of E3 ligase Ube4A disrupts colon homeostasis and accelerates experimental colitis via altered lipid handling

BackgroundUbiquitin-dependent signaling is essential for maintaining intestinal homeostasis and its dysregulation contributes to chronic intestinal disorders, such as Inflammatory Bowel Disease (IBD). Ube4A is a U-box E3/E4 ubiquitin ligase involved in lipid metabolism and insulin signaling in metabolic tissues. Autoantibodies against Ube4A have been identified in patients with IBD and are associated with disease long-term complications. Despite these clinical associations, the physiological role of Ube4A in the gastrointestinal tract remains unknown. This study aimed to define the function of Ube4A in the colon and determine how its loss influences susceptibility to experimental colitis. MethodsUBE4A expression in human colonic tissue from healthy individuals and patients with IBD was analyzed using publicly available single-cell RNA sequencing datasets. The role of Ube4A in colonic homeostasis and colitis pathogenesis was examined using global Ube4A knockout (UKO) mice subjected to dextran sulfate sodium (DSS)-induced colitis. UKO colon phenotypes were characterized using transcriptomic analyses, immunofluorescence, and flow cytometry. ResultsUBE4A is highly expressed in human colonic epithelial cells, and its expression is reduced from healthy to IBD inflamed tissues. In mice, Ube4A deficiency significantly exacerbated DSS-induced colitis, as evidenced by increased weight loss, disease activity scores, shortened colon length, and more severe histological injury. Transcriptomic profiling revealed enhanced inflammatory signaling, alongside dysregulation of lipid transport and storage, as well as antimicrobial defense pathways. DSS-treated UKO mice also exhibited increased mast cell activation and elevated expression of matrix metalloproteinases. Importantly, colons from UKO mice displayed baseline transcriptional alterations indicative of epithelial stress and disrupted lipid metabolic programs, even in the absence of injury. ConclusionsUbe4A is a previously unrecognized regulator of colon homeostasis. Its loss induces existing epithelial stress and metabolic reprogramming that sensitize the colon to exaggerated inflammatory responses during injury such as experimental colitis.

pathology↗

bioRxiv2025

BDH1-Dependent Ketone Body Metabolism Maintains Muller Cell Homeostasis and Retinal Function

Ketone body metabolism serves as an auxiliary regulator of cellular energetics and redox balance, particularly during prolonged fasting and carbohydrate restriction, yet its role in retinal homeostasis under physiological conditions remains poorly defined. {beta}-hydroxybutyrate dehydrogenase 1 (BDH1) is a mitochondrial enzyme that interconverts acetoacetate and {beta}-hydroxybutyrate, and is required for efficient ketone utilization. Here, we investigated the impact of impaired endogenous ketone metabolism on retinal function using global and retinal pigment epithelium (RPE)-specific BDH1 knockout (KO) mice. Global BDH1 KO mice showed reduced circulating {beta}-hydroxybutyrate and blunted fasting-induced ketone elevations, accompanied by ganglion cell loss, structural abnormalities on fundus and OCT imaging, and diminished scotopic and photopic electroretinogram (ERG) a- and b-wave amplitudes, consistent with impaired photoreceptor responses and downstream bipolar and Muller cell signaling. In contrast, RPE-specific BDH1 KO mice exhibited no changes in ERG responses or retinal morphology. Transcriptomic and molecular analyses in global KO retinas revealed disrupted Muller cell homeostasis, including reduced CAMKII-CREB activation, which is required for EAAT1 glutamate transporter expression. Administration of exogenous {beta}-hydroxybutyrate, in vitro and in vivo, restored CAMKII-CREB-EAAT1 signaling, glutamate uptake, and antioxidant gene expression in BDH1 KO mice, demonstrating a central role for ketone bodies in Muller cell metabolic support, glutamate homeostasis, and redox balance. Together with reduced BDH1 expression in human AMD retinas, these findings identify the BDH1-{beta}-hydroxybutyrate axis as a critical metabolic pathway for Muller cell function and retinal integrity, and highlight ketone metabolism as a potential therapeutic target in degenerative retinal diseases.

neuroscience↗

bioRxiv2025

Incentive Salience, not Psychomotor Sensitization or Tolerance, Drives Escalation of Cocaine Self-Administration in Heterogeneous Stock Rats

Sensitization and tolerance are two phenomena often studied independently despite overlapping neurobiological substrates. Each has extensive research showing their influence on the development and maintenance of addiction, but the degree to which they drive escalation in cocaine self-administration is poorly understood. Using self-administration, intravenous noncontingent infusions, and pose-estimation machine vision, we find that incentive salience, not psychomotor sensitization or tolerance, drives the escalation of cocaine self-administration in heterogenous stock rats. Individual differences in psychomotor sensitization or tolerance were found to have no effect on cocaine intake. Incentive salience as measured by locomotion and active lever entrances per meter traveled occurring before the self-administration session began (pre-lever activity measures) during Short Access (2-hours) was found to predict intake during Long Access (6-hours). Both pre-lever locomotion and active lever entrances per meter were found to increase during Long Access and after two-to-three days of abstinence. Critically, rats with low pre-lever activity during Short Access escalated both their intake and pre-lever measures by the end of Long Access to levels comparable with high pre-lever activity rats who maintained their elevated responding. These findings support the notion that incentive salience during Short Access is a catalyst to escalated use and an early marker of addiction vulnerability. Moreover, they suggest that individuals initially resistant to incentive salience can, with sufficient exposure, become sensitized and escalate cocaine use to the same level as more susceptible individuals. Analysis of pre-lever activity offers a novel longitudinal behavioral marker to predict vulnerability and provides a framework for understanding individual trajectories of addiction.

neuroscience↗

bioRxiv2025

Cellular and Spatial Drivers of Unresolved Injury and Functional Decline in the Human Kidney

Building upon a foundational Human Kidney resource, we present a comprehensive multi-modal atlas that defines spatially resolved versus unresolved repair states and mechanisms in human kidney disease. Homeostatic interactions between injured kidney epithelium and its surrounding milieu determine successful repair outcomes, while pathogenic signaling promotes unresolved inflammation and fibrosis leading to chronic disease. We integrated multiple single-cell and spatial modalities across [~]700 samples from >350 patients ([~]250 research biopsies), analyzing [~]1.7 million cells alongside complementary mouse multi-omic profiles spanning acute-to-chronic injury and aging (>300,000 cells) and spatial transcriptomic analysis of >150 human biopsies. This cross-species atlas delineates functional pathways and druggable targets across the nephron and defines gene regulatory networks and chromatin landscapes governing tubular, fibroblast, and immune cell transitions from injury to either recovery or failed repair states. We identified distinct cellular states associated with specific pathological features that show dynamic distributions between acute kidney injury (AKI) and chronic kidney disease (CKD), organized within unique spatial niches that reveal progression mechanisms from early injury to unresolved disease. Gene regulatory analyses prioritized key transcription factor activities (SOX4, SOX9, NFKB1, REL, KLFs) and their target networks establishing disease states and tissue microenvironments. These regulatory programs were directly linked to clinical outcomes, identifying molecular signatures of recovery and secreted biomarkers predictive of AKI-to-CKD progression, providing a key resource for therapeutic development and precision medicine approaches in kidney disease.

molecular biology↗

bioRxiv2025

Genome-wide association study of cocaine self-administration behavior in Heterogeneous Stock rats

BackgroundCocaine use disorder (CUD) is a major public health crisis with detrimental individual and societal effects. The specific genes mediating CUD remain largely unknown. MethodsWe conducted a genome-wide association study (GWAS) using outbred N/NIH Heterogeneous Stock (HS; n = 836, female = 415, male = 421) rats. We examined multiple CUD-related phenotypes that captured acquisition of self-administration, escalation of intake, and compulsive-like responding. ResultsConsistent with the existing literature, these traits were phenotypically and genetically correlated and exhibited modest heritability (h2 = 0.07 - 0.16). We identified six genome-wide significant associations. One locus on chromosome 19 was associated with the variable time between cocaine infusions (post infusion interval) and contains several carboxylesterase genes that are orthologous to the human CES1 gene; notably, carboxylesterases metabolize cocaine. Three non-synonymous coding variants in the genes Ces1c and Ces1d were in perfect linkage disequilibrium with this locus, suggesting that one or more of them might be the causal SNP. The other 5 loci also contained promising coding and expression variants, including Trak2, a gene previously associated with CUD in human GWAS and Slc10a7, Plcl1, and Satb2 which have been associated with alcohol and tobacco use disorder. ConclusionsThis is the largest genetic study of cocaine self-administration ever conducted in any species. Our results replicate previous loci associated with CUD in humans and provide several novel biological insights including the potential of pharmacological strategies targeting carboxylesterases for the treatment of CUD.

genetics↗

bioRxiv2025

High Incidence of Estrous Cycle Irregularities in Heterogeneous Stock (HS) Rats is Associated with Severe Cocaine Addiction-like Behaviors

Hormonal fluctuations throughout the estrous cycle have been hypothesized to influence drug-related behaviors. Preclinical models show that some cocaine-related behaviors are influenced by the estrous cycle. However, the extent to which the estrous cycle modulates cocaine self-administration in outbred heterogeneous stock (HS) rats, a population that captures human genetic diversity, is unknown. This study aimed to examine the relationship between estrous phases and cocaine self-administration behavior in HS rats using a model of extended access to cocaine self-administration. We focused on the escalation of intake, breaking point, and resistance to foot shock. Using vaginal swabbing and lavage techniques, we first characterized the relationship between estrous phase and cocaine intake. We then comprehensively evaluated estrous cycling patterns in young adult and adult HS rats, comparing them with Wistar rats. Contrary to our hypothesis, estrous phase showed no association with cocaine self-administration in HS rats. HS rats exhibited irregular estrous cycling with variability to the phase length, even in the absence of drug exposure, a phenomenon not observed in the Wistar strain. Irregular estrous cycle was associated with high cocaine-related behaviors. This study provides the first evidence that some female HS rats exhibit irregular estrous cycling. Moreover, rats with severe addiction-like behaviors had more instances of irregular cycling. These results demonstrate that, in HS rats, the estrous phase per se has no major influence on cocaine self-administration, but that the severity of addiction-like behaviors are associated with more irregularity of the estrus cycle. As HS rats gain popularity in behavioral and genome-wide studies, understanding these cycle disruptions is crucial as they may reveal genetic links into female vulnerability to drugs.

neuroscience↗

bioRxiv2024

Loss of mitochondrial pyruvate transport initiates cardiac glycogen accumulation and heart failure

Heart failure involves metabolic alterations including increased glycolysis despite unchanged or decreased glucose oxidation. The mitochondrial pyruvate carrier (MPC) regulates pyruvate entry into the mitochondrial matrix, and cardiac deletion of the MPC in mice causes heart failure. How MPC deletion results in heart failure is unknown. We performed targeted metabolomics and isotope tracing in wildtype (fl/fl) and cardiac-specific Mpc2-/- (CS-Mpc2-/-) hearts after in vivo injection of U-13C-glucose. Failing CS-Mpc2-/- hearts contained normal levels of ATP and phosphocreatine, yet these hearts displayed increased enrichment from U-13C-glucose and increased glycolytic metabolite pool sizes. 13C enrichment and pool size was also increased for the glycogen intermediate UDP-glucose, as well as increased enrichment of the glycogen pool. Glycogen levels were increased [~]6-fold in the failing CS-Mpc2-/- hearts, and glycogen granules were easily detected by electron microscopy. In young, non-failing CS-Mpc2-/- hearts, increased glycolytic 13C enrichment occurred, but glycogen levels remained low and unchanged compared to fl/fl hearts. Inhibiting glycogen synthase with MZ-101 reduced cardiac glycogen levels and improved heart failure. Feeding a ketogenic diet to CS-Mpc2-/- mice reversed the heart failure and normalized the cardiac glycogen and glycolytic metabolite accumulation. Cardiac glycogen levels were also elevated in mice infused with angiotensin-II, and both the cardiac hypertrophy and glycogen levels were improved by ketogenic diet. Thus, loss of MPC in the heart causes glycogen accumulation and heart failure, while inhibition of glycogen synthesis or a ketogenic diet can reverse both the glycogen accumulation and heart failure.

physiology↗

bioRxiv2023

Deletion of NuRD component Mta2 in nephron progenitor cells causes developmentally programmed FSGS

Low nephron endowment at birth is a risk factor for chronic kidney disease. The prevalence of this condition is increasing due to higher survival rates of preterm infants and children with multi- organ birth defect syndromes that affect the kidney and urinary tract. We created a mouse model of congenital low nephron number due to deletion of Mta2 in nephron progenitor cells. Mta2 is a core component of the Nucleosome Remodeling and Deacetylase (NuRD) chromatin remodeling complex. These mice developed albuminuria at 4 weeks of age followed by focal segmental glomerulosclerosis (FSGS) at 8 weeks, with progressive kidney injury and fibrosis. Our studies reveal that altered mitochondrial metabolism in the post-natal period leads to accumulation of neutral lipids in glomeruli at 4 weeks of age followed by reduced mitochondrial oxygen consumption. We found that NuRD cooperated with Zbtb7a/7b to regulate a large number of metabolic genes required for fatty acid oxidation and oxidative phosphorylation. Analysis of human kidney tissue also supported a role for reduced mitochondrial lipid metabolism and ZBTB7A/7B in FSGS and CKD. We propose that an inability to meet the physiological and metabolic demands of post-natal somatic growth of the kidney promotes the transition to CKD in the setting of glomerular hypertrophy due to low nephron endowment.

developmental biology↗

bioRxiv2023

Antagonists of the stress and opioid systems restore the functional connectivity of the prefrontal cortex during alcohol withdrawal through divergent mechanisms.

Chronic alcohol consumption leads to dependence and withdrawal symptoms upon cessation, contributing to persistent use. However, the brain network mechanisms by which the brain orchestrates alcohol withdrawal and how these networks are affected by pharmacological treatments remain elusive. Recent work revealed that alcohol withdrawal produces a widespread increase in coordinated brain activity and a decrease in modularity of the whole-brain functional network using single-cell whole-brain imaging of immediate early genes. This decreased modularity and functional hyperconnectivity are hypothesized to be novel biomarkers of alcohol withdrawal in alcohol dependence, which could potentially be used to evaluate the efficacy of new medications for alcohol use disorder. However, there is no evidence that current FDA-approved medications or experimental treatments known to reduce alcohol drinking in animal models can normalize the changes in whole-brain functional connectivity. In this report, we tested the effect of R121919, a CRF1 antagonist, and naltrexone, an FDA-approved treatment for alcohol use disorder, on whole-brain functional connectivity using the cellular marker FOS combined with graph theory and advanced network analyses. Results show that both R121919 and naltrexone restored the functional connectivity of the prefrontal cortex during alcohol withdrawal, but through divergent mechanisms. Specifically, R121919 increased FOS activation in the prefrontal cortex, partially restored modularity, and normalized connectivity, particularly in CRF1-rich regions, including the prefrontal, pallidum, and extended amygdala circuits. On the other hand, naltrexone decreased FOS activation throughout the brain, decreased modularity, and increased connectivity overall except for the Mu opioid receptor-rich regions, including the thalamus. These results identify the brain networks underlying the pharmacological effects of R121919 and naltrexone and demonstrate that these drugs restored different aspects of functional connectivity of the prefrontal cortex, pallidum, amygdala, and thalamus during alcohol withdrawal. Notably, these effects were particularly prominent in CRF1-and Mu opioid receptors-rich regions highlighting the potential of whole-brain functional connectivity using FOS as a tool for identifying neuronal network mechanisms underlying the pharmacological effects of existing and new medications for alcohol use disorder.

neuroscience↗

bioRxiv2023

The chromatin landscape of healthy and injured cell types in the human kidney

There is a need to define regions of gene activation or repression that control human kidney cells in states of health, injury, and repair to understand the molecular pathogenesis of kidney disease and design therapeutic strategies. However, comprehensive integration of gene expression with epigenetic features that define regulatory elements remains a significant challenge. We measured dual single nucleus RNA expression and chromatin accessibility, DNA methylation, and H3K27ac, H3K4me1, H3K4me3, and H3K27me3 histone modifications to decipher the chromatin landscape and gene regulation of the kidney in reference and adaptive injury states. We established a comprehensive and spatially-anchored epigenomic atlas to define the kidneys active, silent, and regulatory accessible chromatin regions across the genome. Using this atlas, we noted distinct control of adaptive injury in different epithelial cell types. A proximal tubule cell transcription factor network of ELF3, KLF6, and KLF10 regulated the transition between health and injury, while in thick ascending limb cells this transition was regulated by NR2F1. Further, combined perturbation of ELF3, KLF6, and KLF10 distinguished two adaptive proximal tubular cell subtypes, one of which manifested a repair trajectory after knockout. This atlas will serve as a foundation to facilitate targeted cell-specific therapeutics by reprogramming gene regulatory networks.

molecular biology↗

bioRxiv2022

Inherent envelope fluctuations in forward masking: Effects of age and hearing loss

Forward masking is generally greater for Gaussian noise (GN) than for low-fluctuation noise maskers, i.e., GN disruption. Because the minimal hearing loss that is associated with older age may affect GN disruption differently than more significant hearing loss, the current study explored the contribution of minimal hearing loss associated with older age to GN disruption. GN disruption was measured using three masker-signal delays (25, 75, and 150 ms) for three adult groups: younger participants with normal hearing, older participants with minimal hearing loss, and older participants with sensorineural hearing loss. The role of underlying mechanisms was tested using a computational model for midbrain neurons. The primary result suggests that older listeners with mild threshold elevations that typically occur with age may be more susceptible to the deleterious effects of masker-envelope fluctuations than younger listeners with normal hearing. Results from the computational model indicate that there may be a larger influence of efferent feedback and saturation of inner hair cells on forward masking and GN disruption than previously thought.

neuroscience↗

bioRxiv2022

Individual differences in oxycodone addiction-like behaviors in a large cohort of heterogeneous stock (HS) rats

Family and twin studies demonstrate that genetic factors determine 20-60% of the vulnerability to opioid use disorder. However, the genes/alleles that mediate the risk of developing addiction-related behaviors, including the sensitivity to the analgesic efficacy of opioids, the development of tolerance, dependence, and escalation of oxycodone taking and seeking, have been ill-defined, thus hindering efforts to design pharmacological interventions to enable precision medicine strategies. Here we characterized oxycodone addiction-like behaviors in heterogeneous stock (HS) rats, that show high genetic diversity that mimics the high genetic variability in humans. HS rats were allowed to self-administer oxycodone for two h/daily for four days (ShA) and then moved to 12h/daily (LgA) for 14 days. Animals were screened for motivation to self-administer oxycodone using a progressive-ratio (PR) schedule of reinforcement and for the development of withdrawal-induced hyperalgesia and tolerance to the analgesic effects of oxycodone using the von-Frey and tail immersion tests, respectively. To reduce cohort-specific effects, we used cohorts of 46-60 rats and normalized the response level within cohorts using a Z-score. To take advantage of the four opioid-related behaviors and further identify subjects that are consistently vulnerable vs. resilient to compulsive oxycodone use, we computed an Addiction Index by averaging normalized responding (Z-scores) for the four behavioral tests. Results showed high individual variability between vulnerable and resilient rats, likely to facilitate the detection of gene variants associated with vulnerable vs. resilient individuals. Such data will have considerable translational value for designing follow-up studies in humans.

neuroscience↗

bioRxiv2022

Identification of individual differences in response to methadone, buprenorphine, and naltrexone in animal models of opioid use disorder

RationaleCurrent medications for opioid use disorder include buprenorphine, methadone, and naltrexone. While these medications show significant efficacy in reducing craving and opioid use, there are substantial individual differences in response to these treatments in humans. The reason for such difference is poorly known. ObjectivesHere, we tested the hypothesis that similar individual differences may be observed in a large population of heterogenous stock rats, that have been bred to maximize genetic diversity, using a behavioral paradigm relevant to opioid use disorder. MethodsOver 500 rats were given intermittent (4d/week) and extended access (12h/day) to oxycodone self-administration for 14 sessions to establish oxycodone dependence and escalation of intake. We then measured the effect of buprenorphine (0.5mg/kg), methadone (3mg/kg) and naltrexone (3mg/kg) on the motivation to self-administer oxycodone by using a progressive ratio schedule of reinforcement. ResultsWe found that naltrexone and buprenorphine significantly decreased motivation to oxycodone rewards. While naltrexone reduced oxycodone intake in both males and females, systemic administration with buprenorphine reduced progressive ratio responses only in males. Methadone reduced motivation to oxycodone self-administration in nearly 25% of the population, without reaching statical significance. Our results showed that the efficacy of these medications depends on the severity of addiction like behaviors, indicated by the addiction index. ConclusionsThese results demonstrate individual differences in response to medications to treat opioid use disorder in a genetically diverse population of rats.

neuroscience↗

bioRxiv2022

Identification of pre-existing microbiome and metabolic vulnerabilities to escalation of oxycodone self-administration and identification of a causal role of short-chain fatty acids in addiction-like behaviors.

The gut brain axis is thought to play a role in behavior and physiological responses through chemical, immunological, and metabolite signaling. Antibiotics, diet, and drugs can alter the transit time of gut contents as well as the makeup of the microbiome. Heterogeneity in genetics and environment are also well-known factors involved in the initiation and perpetuation of substance use disorders. Few viable genetic or biological markers are available to identify individuals who are at risk of escalating opioid intake. Primarily, the addiction field has focused on the nervous system, limiting the discovery of peripheral factors that contribute to addiction. To address this gap, we characterized the microbiome before and after drug exposure, and after antibiotics depletion in male and female heterogenous stock rats to determine if microbiome constituents are protective of escalation. We hypothesized that individuals that are prone to escalation of opioid self-administration will have distinct microbial and metabolic profiles. The fecal microbiome and behavioral responses were measured over several weeks of oxycodone self-administration and after antibiotic treatment. Antibiotic treatment reduces circulating short-chain fatty acids (SCFA) by depleting microbes that ferment fiber into these essential signaling molecules for the gut-brain axis. Depletion of the microbiome increased oxycodone self-administration in a subpopulation of animals (Responders). Supplementation of SCFAs in antibiotic depleted animals decreased elevated oxycodone self-administration. Phylogenetic functional analysis reveals distinct metabolic differences in the subpopulations of animals that are sensitive to antibiotic depletion and animals rescued by SCFA supplementation. In conclusion, this study identifies pre-existing microbiome and metabolic vulnerabilities to escalation of oxycodone self-administration, demonstrates that escalation of oxycodone self-administration dysregulates the microbiome and metabolic landscape, and identifies a causal role of short-chain fatty acids in addiction-like behaviors.

neuroscience↗