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To characterize disclosures of conflicts of interest in review articles in psychiatry, we identified 285 reviews from 10 high-impact journals in psychiatry and 2 in general medicine. Disclosures were reliably coded as biotechnology/pharmaceutical/other material interests, nonprofit/government, communication companies, or other. The authors in both types of journals frequently reported industry ties. However, the reviews in the psychiatric journals were significantly less likely to include industry-related disclosures (32% of the reviews; 18% of the authors) compared with the general medical journals (64% of the articles; 40% of the authors). The most common types of industry-related disclosures were for consulting, research support, and speaking fees. Disclosures seemed to be of limited utility in helping readers assess possible biases because the nature and the extent of the relationships being disclosed were often unclear. Efforts to screen out authors with significant financial relationships pertaining to the topic under review may be more effective than are disclosures in protecting the integrity of the medical literature. [ABSTRACT FROM AUTHOR]

Substance use is significantly associated with physical injury, yet relatively little is known about the prevalence of specific substance use disorders among trauma patients, or their associated sociodemographic characteristics. We evaluated these issues in an unselected sample of 1,118 adult inpatients at the University of Maryland Shock Trauma Center, Baltimore, MD, who were interviewed with the psychoactive substance use disorder section of the Structured Clinical Interview for DSM-III-R. Among trauma inpatients, lifetime alcohol users (71.8% of subjects) were more likely male; users of illegal drugs (45.3%) were also more likely to be younger, unmarried, and poor. Patients with current drug abuse/dependence (18.8%) were more likely to be non-white, less educated, and poor; those with current alcohol abuse/dependence (32.1%) were also more likely male, unmarried, and older. These findings highlight the need for screening for substance use disorders in trauma settings and referral of patients to substance abuse treatment programs. [ABSTRACT FROM AUTHOR]

One measure of a substance's addictive risk is the proportion of users who become dependent. This study evaluates the lifetime and current risk of substance dependence among lifetime substance users' among trauma inpatients and provides a relative ranking of addictive risk among the substances. Data on use of 8 substance groups (alcohol, opiates, marijuana, cocaine, other stimulants, sedative-hypnotics, hallucinogens, other drugs) were obtained by interview (Structured Clinical Interview for the DSM-III-R) from 1,118 adult trauma inpatients. Prevalence of lifetime dependence among lifetime users ranged from 80.7% for opiates and 70.9% for cocaine to 33.3% for hallucinogens and 26.6% for sedative-hypnotics. The rank order of addictive risk was similar to that found in the general population. Trauma inpatients had a higher absolute addictive risk than the general population, comparable to the risk found in patients in treatment for substance use disorders, suggesting the importance of screening trauma inpatients for substance dependence. [ABSTRACT FROM AUTHOR]

Describes the French Law and system for human research subject protection, with its advantages and disadvantages, and compares it with the U.S. System. The aim is to improve the latter by identifying aspects of the French system that are potentially worth adapting and adopting. [ABSTRACT FROM AUTHOR]

Objective: To assess the prevalence of psychoactive substance use disorders (PSUDs) among a large, unselected group of seriously injured trauma center patients, using a standardized diagnostic interview and criteria.Design: Prevalence study.Setting: A level I regional trauma center.Patients: Trauma center patients fulfilling the following criteria were eligible subjects: aged 18 years or older, admission from injury scene, length of stay of 2 days or longer, and intact cognition.Outcome Measures: The PSUDs were diagnosed using the Structured Clinical Interview (SCID) for the Diagnostic and Statistical Manual of Mental Disorders, Revised Third Edition (DSM-III-R) and were categorized as abuse or dependence and past or current (within past 6 months). The SCID results were analyzed with respect to demographic factors, injury type, and blood alcohol concentration and urine toxicology results, using chi2 and logistic regression techniques.Results: Of the 1220 patients approached for study, 1118 (91.6%) consented. More than half (54.2%) had a diagnosis of a PSUD in their lifetime. Approximately 90% of alcohol and other drug use diagnoses were for dependence and more than 62% were current. Overall, 24.1% of patients were currently alcohol dependent (men, 27.7%; women, 14.7%; P<.001), and 17.7% were currently dependent on other drugs (men, 20.2%; women, 11.2%; P<.001). Current alcohol dependence rates were not associated with race; rates of dependence on other drugs were higher among nonwhites and victims classified with intentional injuries. While 54.3% of blood alcohol-positive patients were currently alcohol dependent and 38.7% of patients with positive urine screening test results for drugs other than alcohol and nicotine were currently drug dependent, 11.7% of blood alcohol-negative and 3.9% of drug-negative patients, respectively, had current diagnoses of dependence on psychoactive substances.Conclusions: A high percentage of seriously injured trauma center patients are at risk of having current PSUDs. Patients with positive toxicology screening test results and/or positive screening questionnaire responses should be referred for formal evaluation and treatment. [ABSTRACT FROM AUTHOR]

Nuclear receptors are important in normal physiology and disease. Physicians and scientists who study nuclear receptors organize and attend conferences and symposia devoted to foundational and translational nuclear receptor research, but the field lacks a platform for early-stage investigators and aspiring leaders. In 2019, Zeynep Madak-Erdogan, Rebecca Riggins, and Matthew Sikora founded Nuclear Receptor (NR) Interdisciplinary Meeting for Progress And Collaboration Together (IMPACT, https://nrimpact.com), a collaborative group designed for early- and mid-career faculty who study nuclear receptors in any context or organism [1]. NR IMPACT addresses challenges for early- and mid-career faculty. Here, we review the progress of NR IMPACT and discuss how our peer-mentoring cohort is removing hurdles for new faculty and advancing nuclear receptor biology., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Endocrine Society.)

Androgens are classically thought to act through intracellular androgen receptors (AR/NR3C4), but they can also trigger non-genomic effects via membrane proteins. Although several membrane androgen receptors have been characterized in vitro, their functions in vivo remain unclear. Using a chemical-genetic screen in zebrafish, we found that GPRC6A, a G-protein coupled receptor, mediates non-genomic androgen actions during embryonic development. Exposure to androgens (androstanedione, DHT, and testosterone) caused cardiac edema or tail curvature in wild-type embryos, as well as in ar mutants, suggesting AR-independent pathways. We then mutated putative membrane androgen receptors (gprc6a, hcar1-4, and zip9) and found that only gprc6a mutants exhibited a significant reduction in cardiac edema following testosterone exposure. Additionally, co-treatment of wild-type embryos with testosterone and GPRC6A antagonists significantly suppressed the cardiac edema phenotype. Using RNA-seq and RNA rescue approaches, we found that testosterone-GPRC6A causes cardiac phenotypes by reducing Pak1 signaling. Our results indicate that testosterone induces cardiac edema in zebrafish embryos through GPRC6A, independent of nuclear androgen receptors, highlighting a novel non-genomic androgen signaling pathway in embryonic development., (© 2024. Published by The Company of Biologists Ltd.)

Introduction: Clozapine is the most effective antipsychotic for treatment-resistant schizophrenia, but it is markedly underutilized, particularly in the US Black population, partly because of concern over clozapine-associated low absolute neutrophil count (ANC). People of African descent have a lower normative ANC range than the White population, which is associated with a specific "ACKR1-null" ("Duffy null") CC genotype (SNP rs2814778) on the ACKR1 gene, termed benign ethnic neutropenia (BEN). The range of ANC variability and safety of clozapine have not been established in people with BEN or examined prospectively in people of African descent., Methods: We completed a multisite, 6-month, prospective, open-label clinical trial of clozapine treatment in people of African descent with schizophrenia spectrum disorders for whom clozapine was clinically indicated, with or without the ACKR1-null genotype. We examined clozapine safety and weekly ANC during clozapine treatment and evaluated ANC variability by ACKR1-null genotype, sex, study site, and clozapine dosing using repeated measures analysis of covariance. Genotype was assayed using TaqMan® technology., Results: We enrolled 274 participants, of whom 227 (82.8 %) completed 6 months of clozapine treatment. There was one case of severe neutropenia (<500 cells/mm 3 ) (0.36 %) over 1467.6 person-months of clozapine exposure. This participant recovered without sequelae after discontinuation of clozapine. Of the 249 participants with known genotypes, 199 (79.9 %) had the ACKR1-null genotype. Neutropenia (<1500 cells/mm 3 ) occurred significantly more often in the ACKR1-null group (33 % [65/199]) than in those with the T allele (6 % (3/50); p < 0.001). Fourteen (5 %) patients discontinued due to adverse events. Rates of infection and fever were low and sialorrhea was the commonest side effect (N = 187, 68 %)., Conclusion: To our knowledge, this is the largest prospective clozapine trial in people of African descent. Severe neutropenia was rare, despite the high prevalence (80 %) of the ACKR1-null genotype. Our findings suggest that clozapine can be used safely in Black patients including those with BEN., Competing Interests: Declaration of competing interest Dr. Deanna Kelly serves on an advisory board for Alkermes, Teva, and Janssen. Dr. Richard Adebayo has served on the advisory boards of Janssen and a consultant for Invega product. Dr. Raymond Loves serves as a consultant for the Maryland Department of Health and the SMI Advisor, American Psychiatric Association. Dr. Sophie Lanzkron has served as an advisor for Bluebird bio, Novo Nordisk, Pfizer, Novartis and Magenta. She has research funding from Imana, Novartis, GBT, Takeda, CSL-Behring, HRSA, PCORI, MD CHRC. Dr. Robert Buchanan has served on the advisory board for Acadia, Merck and Neurocranial, is a consultant for Boehringer-Ingelheim, and is on DSMB for Merck, Negron, and Roche. All other authors have no conflicts to disclose., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The initial step in estrogen-regulated transcription is the binding of a ligand to its cognate receptors, named estrogen receptors (ERα and ERβ). Phytochemicals present in foods and environment can compete with endogenous hormones to alter physiological responses. We screened 224 flavonoids in our engineered biosensor ERα and ERβ PRL-array cell lines to characterize their activity on several steps of the estrogen signaling pathway. We identified 83 and 96 flavonoids that can activate ERα or ERβ, respectively. While most act on both receptors, many appear to be subtype-selective, including potent flavonoids that activate ER at sub-micromolar concentrations. We employed an orthogonal assay using a transgenic zebrafish in vivo model that validated the estrogenic potential of these compounds. To our knowledge, this is the largest study thus far on flavonoids and the ER pathway, facilitating the identification of a new set of potential endocrine disruptors acting on both ERα and ERβ., Competing Interests: The authors declare no competing interests., (© 2024 The Authors.)

In this study we present an inducible biosensor model for the Estrogen Receptor Beta (ERβ), GFP-ERβ:PRL-HeLa, a single-cell-based high throughput (HT) in vitro assay that allows direct visualization and measurement of GFP-tagged ERβ binding to ER-specific DNA response elements (EREs), ERβ-induced chromatin remodeling, and monitor transcriptional alterations via mRNA fluorescence in situ hybridization for a prolactin (PRL)-dsRED2 reporter gene. The model was used to accurately (Z' = 0.58-0.8) differentiate ERβ-selective ligands from ERα ligands when treated with a panel of selective agonists and antagonists. Next, we tested an Environmental Protection Agency (EPA)-provided set of 45 estrogenic reference chemicals with known ERα in vivo activity and identified several that activated ERβ as well, with varying sensitivity, including a subset that is completely novel. We then used an orthogonal ERE-containing transgenic zebrafish (ZF) model to cross validate ERβ and ERα selective activities at the organism level. Using this environmentally relevant ZF assay, some compounds were confirmed to have ERβ activity, validating the GFP-ERβ:PRL-HeLa assay as a screening tool for potential ERβ active endocrine disruptors (EDCs). These data demonstrate the value of sensitive multiplex mechanistic data gathered by the GFP-ERβ:PRL-HeLa assay coupled with an orthogonal zebrafish model to rapidly identify environmentally relevant ERβ EDCs and improve upon currently available screening tools for this understudied nuclear receptor., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Authors.)

Mechanistic toxicology seeks to identify the molecular and cellular mechanisms by which toxicants exert their deleterious effects. One powerful approach is to generate mutations in genes that respond to a particular toxicant, and then test how such mutations change the effects of the toxicant. CRISPR is a rapid and versatile approach to generate mutations in cultured cells and in animal models. Many studies use CRISPR to generate short insertions or deletions in a target gene and then assume that the resulting mutation, such as a premature termination codon, causes a loss of functional protein. However, recent studies demonstrate that this assumption is flawed. Cells can compensate for short insertion and deletion mutations, leading toxicologists to draw erroneous conclusions from mutant studies. In this review, we will discuss mechanisms by which a mutation in one gene may be rescued by compensatory activity. We will discuss how CRISPR insertion and deletion mutations are susceptible to compensation by transcriptional adaptation, alternative splicing, and rescue by maternally derived gene products. We will review evidence that measuring levels of messenger RNA transcribed from a mutated gene is an unreliable indicator of the severity of the mutation. Finally, we provide guidelines for using CRISPR to generate mutations that avoid compensation., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

During embryonic development, a subset of cells in the mesoderm germ layer are specified as hemato-vascular progenitor cells, which then differentiate into endothelial cells and hematopoietic stem and progenitor cells. In zebrafish, the transcription factor npas4l (cloche) is required for the specification of hemato-vascular progenitor cells. However, it is unclear whether npas4l is the sole factor at the top of the hemato-vascular specification cascade. Here, we show that arnt1 and arnt2 genes are required for hemato-vascular specification. We found that arnt1;arnt2 double mutant zebrafish embryos, but not arnt1 or arnt2 single mutants, lack blood cells and most endothelial cells. arnt1/2 mutants have reduced or absent expression of etsrp and tal1, the earliest known endothelial and hematopoietic transcription factor genes. We found that Npas4l binds both Arnt1 and Arnt2 proteins in vitro, consistent with the idea that PAS domain-containing bHLH transcription factors act in a multimeric complex to regulate gene expression. Our results demonstrate that npas4l, arnt1 and arnt2 act together to regulate endothelial and hematopoietic cell fate, where each gene is necessary, but not sufficient, to drive hemato-vascular specification., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2023. Published by The Company of Biologists Ltd.)

Background: In a previous study, ondansetron, a serotonin 5-HT 3 receptor antagonist, reduced drinking intensity (drinks/drinking day [DPDD]) among European-ancestry (EA) participants with moderate-to-severe alcohol use disorder (AUD) and variants in genes encoding the serotonin transporter (SLC6A4) and 5-HT 3A (HTR3A), and 5-HT 3B (HTR3B) receptors. We tested whether (1) ondansetron reduces DPDD among individuals of either European or African ancestry (AA), and (2) that reductions in DPDD are greatest among ondansetron-treated individuals with population-specific combinations of genotypes at SLC6A4, HTR3A, and HTR3B., Methods: In this 16-week, double-blind, placebo-controlled, parallel-group clinical trial, adults with AUD were randomized to receive low-dose oral ondansetron (0.33 mg twice daily) or placebo stratified by "responsive" versus "nonresponsive" genotype defined using population-specific genotypes at the three genetic loci. Generalized estimating equation regression models and a modified intent-to-treat analysis were used to compare the treatment groups on the primary outcome-DPDD-and two secondary outcomes-heavy drinking days per week [HDD] and drinks per day [DPD] across the 16 weeks of treatment., Results: Of 296 prospective participants screened, 95 (58 EA and 37 AA) were randomized and received at least one dose of study medication. In the modified intent-to-treat analysis, the ondansetron group averaged 0.40 more DPDD (p = 0.51), 1.35 times as many HDD (p = 0.16), and 1.06 times as many DPD (p = 0.59) as the placebo group. There were no significant interactions with genotype. There were no study-related serious adverse events (AEs) and similar proportions of participants in the two treatment groups experienced AEs across organ systems., Conclusions: We found no evidence that low-dose oral ondansetron is beneficial in the treatment of AUD, irrespective of genotype, thus failing to confirm prior study findings. However, the study was underpowered to identify medication by genotype interactions., (© 2022 Research Society on Alcoholism.)

Background: The serotonin transporter (SERT) mRNA was previously reported to be a quantitative and pathophysiology-based biomarker of heavy drinking in 5HTTLPR:LL genotype-carriers treated with ondansetron. Here, we validated the potential use of SERT mRNA for quantitative prediction of recent alcohol consumption (in the absence of treatment) and compared it with the known biomarkers ethyl glucuronide (EtG) and ethyl sulfate (EtS)., Methods: Binge drinking men and women of European ancestry aged 21 to 65 years were enrolled in a 12-day, in-patient, randomized, double-blind, crossover study, where they were administered three beverage doses (placebo, 0.5 g/kg [0.4 g/kg] ethanol, and 1 g/kg [0.9 g/kg] ethanol for men [women]) individually in three 4-day periods (experiments), separated by minimum 7-day washout period. Diet, sleep, and physical activity were controlled throughout the inpatient experiments. Twenty-nine participants were randomized to receive beverage doses counterbalancing the sequence of treatment and gender within subgroups stratified by SERT genotypes 5HTTLPR:LL+rs25531:AA (L A L A ) versus 5HTTLPR:LS/SS. Peripheral venous blood was collected daily for (1) quantification of SERT mRNA (the primary outcome measure) using qRT-PCR and (2) plasma EtG and EtS levels using tandem mass-spectrometry., Results: The association between administered beverage dose and SERT mRNA from completers of at least one 4-day experiment (N = 18) assessed by a linear mixed model was not statistically significant. Significant positive associations were found with beverage dose and plasma EtG, EtS and EtG/EtS ratio (β = 5.8, SE = 1.2, p < 0.0001; β = 1.3, SE = 0.6, p = 0.023; and β = 3.0, SE = 0.7, p < 0.0001, respectively; the C-statistics for discriminating outcomes were 0.97, 0.8, and 0.92, respectively). Additionally, we observed a sequence effect with a greater placebo effect on SERT mRNA when it was administered during the first experiment (p = 0.0009), but not on EtG/EtS measures., Conclusion: The findings do not validate the use of SERT as a biomarker of heavy drinking. Larger and more innovative studies addressing the effects of placebo, race, gender, and response to treatment with serotonergic agents are needed to fully assess the utility of SERT as a biomarker of heavy and binge drinking., (© 2022 Research Society on Alcoholism.)

Forty-one US jurisdictions (37 states) have legalized comprehensive medical cannabis programs since 1996. The number of qualifying conditions per jurisdiction varies from 5 to 29. Five (12%) of 42 qualifying conditions have conclusive or substantial evidence of efficacy and are listed in more than half of all jurisdictions. Half (50%) of qualifying conditions have no or insufficient scientific evidence of benefit from medical cannabis; 9% of qualifying conditions have limited evidence of harm from medical cannabis. The mean number of qualifying conditions per jurisdiction and the proportion of conditions with and without evidence of benefit have not changed since 1996., Competing Interests: Disclosure Dr G.M. Burnett has no disclosures. Dr D.A. Gorelick receives royalties from Wolters Kluwer for writing medical articles about cannabis and honoraria from Springer Nature and Colorado State University—Pueblo for serving as Editor-in-Chief of the Journal of Cannabis Research. Dr K.P. Hill is a consultant to Greenwich Biosciences and is an author with Wolters Kluwer., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Kratom is the common term for Mitragyna speciosa and its products. Its major active compounds are mitragynine and 7-hydroxymitragynine. An estimated 2.1 million US residents used kratom in 2020, as a "legal high" and self-medication for pain, opioid withdrawal, and other conditions. Up to 20% of US kratom users report symptoms consistent with kratom use disorder. Kratom use is associated with medical toxicity and death. Causality is difficult to prove as almost all cases involve other psychoactive substances. Daily, high-dose use may result in kratom use disorder and opioid-like withdrawal on cessation of use. These are best treated with buprenorphine., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Proteins that contain basic helix-loop-helix (bHLH) and Per-Arnt-Sim motifs (PAS) function as transcription factors. bHLH-PAS proteins exhibit essential and diverse functions throughout the body, from cell specification and differentiation in embryonic development to the proper function of organs like the brain and liver in adulthood. bHLH-PAS proteins are divided into two classes, which form heterodimers to regulate transcription. Class I bHLH-PAS proteins are typically activated in response to specific stimuli, while class II proteins are expressed more ubiquitously. Here, we discuss the general structure and functions of bHLH-PAS proteins throughout the animal kingdom, including family members that do not fit neatly into the class I-class II organization. We review heterodimerization between class I and class II bHLH-PAS proteins, binding partner selectivity and functional redundancy. Finally, we discuss the evolution of bHLH-PAS proteins, and why a class I protein essential for cardiovascular development in vertebrates like chicken and fish is absent from mammals., (© 2022 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.)

In vivo models to detect estrogenic compounds are very valuable for screening for endocrine disruptors. Here we describe the use of transgenic estrogen reporter zebrafish as an in vivo model for the identification of estrogenic properties of compounds. Live imaging of these transgenic fish provides knowledge of estrogen receptor specificity of different ligands as well as dynamics of estrogen signaling. Coupled to image analysis, the model can provide quantitative concentration-response information on estrogenic activity of chemical compounds., (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Steroid hormones bind receptors in the cell nucleus and in the cell membrane. The most widely studied class of steroid hormone receptors are the nuclear receptors, named for their function as ligand-dependent transcription factors in the cell nucleus. Nuclear receptors, such as estrogen receptor alpha, can also be anchored to the plasma membrane, where they respond to steroids by activating signaling pathways independent of their function as transcription factors. Steroids can also bind integral membrane proteins, such as the G protein-coupled estrogen receptor. Membrane estrogen and progestin receptors have been cloned and characterized in vitro and influence the development and function of many organ systems. Membrane androgen receptors were cloned and characterized in vitro, but their function as androgen receptors in vivo is unresolved. We review the identity and function of membrane proteins that bind estrogens, progestins, and androgens. We discuss evidence that membrane glucocorticoid and mineralocorticoid receptors exist, and whether glucocorticoid and mineralocorticoid nuclear receptors act at the cell membrane. In many cases, integral membrane steroid receptors act independently of nuclear steroid receptors, even though they may share a ligand., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Endocrine Society.)

Introduction: Psychoactive substance use disorders (SUDs) are common in trauma patients and substance use has become a leading cause of death in the United States. The purpose of this study is to examine the impact of a lifetime SUD and SUD characteristics (substance used, current SUD versus in remission from dependence, etc.) on the long-term survival of trauma patients., Methods: Cohort study of consecutive adult trauma inpatients who were discharged alive from a level-one trauma center (1994-1996). The presence of lifetime SUD was determined at the time of admission by the Structured Clinical Interview for the Diagnostic and Statistical Manual III-R. Mortality follow-up through the end of 2017 was obtained by linking patients to a national database of death certificates. Cox proportional hazards analysis was used to determine the association of lifetime SUD and death after adjusting for age and tobacco use., Results: 1,220 patients were approached, 1,118 consented to participate, and 1,099 had personal identifiers for matching. 789 (71.8%) of subjects were men, 596 (54.2%) had lifetime SUDs, and 325 (29.6%) died. Injury was the most common cause of death (24.6%, 80/325), with poisonings (40.0%, 32/80) being the most common injury-related cause of death. Compared to those without a lifetime SUD, lifetime SUD was associated with increased all-cause mortality (adjusted hazard ratio [HR adj ]=1.83; 95% CI, 1.4 to 2.4), injury death (HR adj =2.47; 95% CI: 1.4 to 4.2), and fatal opioid overdose (HR adj =12.96; 95% CI, 1.7 to 100.4)(p ≤ 0.01 for all HR adj )., Conclusions: The presence of a lifetime SUD was associated with early death, particularly from reinjury, in trauma patients. It is important to address a patient's SUD during admission to decrease their chances of dying after discharge, especially due to injury-related causes., Competing Interests: Declaration of Competing Interest This manuscript has not previously been published and is not currently under consideration by another journal. A portion of the results were presented at the International Cannabinoid Research Society meeting in June 2019. The initial data collection was funded by the National Institutes of Health/National Institute on Alcohol Abuse and Addiction (R01 AA009050 01A2 awarded to CAS). The analysis was supported by the Maryland Emergency Medicine Network and the National Foundation of Emergency Medicine (awarded to ZDWD). None of the authors have any commercial associations or sources of support that suggest or constitute a conflict of interest., (Copyright © 2020. Published by Elsevier Ltd.)

Background and Aims: During liver development, bipotent progenitor cells differentiate into hepatocytes and biliary epithelial cells to ensure a functional liver required to maintain organismal homeostasis. The developmental cues controlling the differentiation of committed progenitors into these cell types, however, are incompletely understood. Here, we discover an essential role for estrogenic regulation in vertebrate liver development to affect hepatobiliary fate decisions., Approach and Results: Exposure of zebrafish embryos to 17β-estradiol (E2) during liver development significantly decreased hepatocyte-specific gene expression, liver size, and hepatocyte number. In contrast, pharmacological blockade of estrogen synthesis or nuclear estrogen receptor (ESR) signaling enhanced liver size and hepatocyte marker expression. Transgenic reporter fish demonstrated nuclear ESR activity in the developing liver. Chemical inhibition and morpholino knockdown of nuclear estrogen receptor 2b (esr2b) increased hepatocyte gene expression and blocked the effects of E2 exposure. esr2b -/- mutant zebrafish exhibited significantly increased expression of hepatocyte markers with no impact on liver progenitors, other endodermal lineages, or vasculature. Significantly, E2-stimulated Esr2b activity promoted biliary epithelial differentiation at the expense of hepatocyte fate, whereas loss of esr2b impaired biliary lineage commitment. Chemical and genetic epistasis studies identified bone morphogenetic protein (BMP) signaling as a mediator of the estrogen effects. The divergent impact of estrogen on hepatobiliary fate was confirmed in a human hepatoblast cell line, indicating the relevance of this pathway for human liver development., Conclusions: Our studies identify E2, esr2b, and downstream BMP activity as important regulators of hepatobiliary fate decisions during vertebrate liver development. These results have significant clinical implications for liver development in infants exposed to abnormal estrogen levels or estrogenic compounds during pregnancy., (© 2020 by the American Association for the Study of Liver Diseases.)

Objective: To assess divided-attention performance when driving under the influence of cannabis with and without alcohol. Three divided-attention tasks were performed following administration of placebo, cannabis, and/or alcohol., Methods: Healthy adult cannabis users participated in 6 sessions, receiving combinations of cannabis (placebo/low-THC/high-THC) and alcohol (placebo/active) in randomized order, separated by washout periods of ≥1 week. At 0.5 hours post-dosing, participants performed simulator drives in the University of Iowa National Advanced Driving Simulator (NADS-1), a full vehicle cab simulator with a 360° horizontal field of view and motion base that provides realistic feedback. Drives contained repeated instances of three tasks: a side-mirror task (reaction to a triangle appearing in the side-mirrors), an artist-search task (select a specified artist from a navigable menu on the vehicle's console), and a message-reading task (read aloud a message displayed on the console). Blood THC and breath alcohol concentration (BrAC) were interpolated using individual power curves from samples collected approximately 0.17, 0.42, 1.4, and 2.3 hours post-dose. Driving measures during tasks were compared to equal-duration control periods occurring just prior to the task. Performance shifts, task completion, and lane departures were modeled relative to blood THC and BrAC using mixed-effects regression models., Results: Each 1 µg/L increase in blood THC concentration predicted increased odds of failing to complete the artist-search task (OR: 1.05, 95% CI: 1.01-1.11, p = 0.046), increased odds of selecting at least one incorrect response (OR: 1.05, 95% CI: 1.00-1.09, p = 0.041), declines in speed during the side-mirror task (0.005 m/s, 95% CI: 0.001-0.009, p = 0.023), and longer lane departure durations during the artist-search task (0.74% of task-period, 95% CI: 0.12-1.36 p = 0.020). BrAC (approximately 0.05%) was not associated with task performance, though each 0.01 g/210 L increase predicted longer departure durations during the side-mirror task (1.41% of task-period, 95% CI: 0.08-2.76, p = 0.040) and increased standard deviation of lane position in the message-reading task (0.61 cm, 95% CI: 0.14-1.08, p = 0.011)., Conclusions: With increasing medical and legal cannabis use, understanding the impact of acute cannabis use on driving performance, including divided-attention, is essential. These data indicate that impaired divided-attention performance is a safety concern.

Objective: Maternal folate (vitamin B9) status is the largest known modifier of neural tube defect risk, so we evaluated folate-related mechanisms of action for dolutegravir (DTG) developmental toxicity., Design: Folate receptor 1 (FOLR1) was examined as a target for DTG developmental toxicity using protein and cellular interaction studies and an animal model., Methods: FOLR1 competitive binding studies were used to test DTG for FOLR1 antagonism. Human placenta cell line studies were used to test interactions with DTG, folate, and cations. Zebrafish were selected as an animal model to examine DTG-induced developmental toxicity and rescue strategies., Results: FOLR1 binding studies indicate DTG is a noncompetitive FOLR1 antagonist at therapeutic concentrations. In-vitro testing indicates calcium (2 mmol/l) increases FOLR1-folate interactions and alters DTG-FOLR1-folate interactions and cytotoxicity. DTG does not inhibit downstream folate metabolism by dihydrofolate reductase. Early embryonic exposure to DTG is developmentally toxic in zebrafish, and supplemental folic acid can mitigate DTG developmental toxicity., Conclusion: Folates and FOLR1 are established modifiers of risk for neural tube defects, and binding data indicates DTG is a partial antagonist of FOLR1. Supplemental folate can ameliorate increased developmental toxicity due to DTG in zebrafish. The results from these studies are expected to inform and guide future animal models and clinical studies of DTG-based antiretroviral therapy in women of childbearing age.

The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that binds environmental toxicants and regulates gene expression. AHR also regulates developmental processes, like craniofacial development and hematopoiesis, in the absence of environmental exposures. Zebrafish have 3 paralogs of AHR: ahr1a, ahr1b, and ahr2. Adult zebrafish with mutations in ahr2 exhibited craniofacial and fin defects. However, the degree to which ahr1a and ahr1b influence ahr2 signaling and contribute to fin and craniofacial development are not known. We compared morphology of adult ahr2 mutants and ahr1a;ahr1b single and double mutant zebrafish. We found that ahr1a;ahr1b single and double mutants were morphologically normal whereas ahr2 mutant zebrafish demonstrated fin and craniofacial malformations. At 5 days post fertilization, both ahr1a;ahr1b and ahr2 mutant larvae were normal, suggesting that adult phenotypes are due to defects in maturation or maintenance. Next, we analyzed the function of zebrafish AHRs activated by environmental ligands. The prototypical AHR ligand, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), induces toxicity in humans and rodents via AHR and causes cardiotoxicity in zebrafish embryos. It has been shown that embryos with mutations in ahr2 are resistant to TCDD toxicity, yet it is unclear whether ahr1 receptors are required. Furthermore, though AHR was shown to interact with estrogen receptor alpha following TCDD treatment, it is not known whether this interaction is constitutive or context-dependent. To determine whether estrogen receptors are constitutive cofactors for AHR signaling, we used genetic and pharmacologic techniques to analyze TCDD-dependent toxicity in estrogen receptor and ahr mutant embryos. We found that embryos with mutations in ahr1a;ahr1b or estrogen receptor genes are susceptible to TCDD toxicity whereas ahr2 mutant embryos are TCDD-resistant. Moreover, pharmacologic blockade of nuclear estrogen receptors failed to prevent TCDD toxicity. These findings suggest that ahr1 genes do not have overlapping functions with ahr2 in fin and craniofacial development or TCDD-dependent toxicity, and that estrogen receptors are not constitutive partners of ahr2., (© The Author(s) 2019. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background & Aims: Patients with cirrhosis are at high risk for hepatocellular carcinoma (HCC) and often have increased serum levels of estrogen. It is not clear how estrogen promotes hepatic growth. We investigated the effects of estrogen on hepatocyte proliferation during zebrafish development, liver regeneration, and carcinogenesis. We also studied human hepatocytes and liver tissues., Methods: Zebrafish were exposed to selective modifiers of estrogen signaling at larval and adult stages. Liver growth was assessed by gene expression, fluorescent imaging, and histologic analyses. We monitored liver regeneration after hepatocyte ablation and HCC development after administration of chemical carcinogens (dimethylbenzanthrazene). Proliferation of human hepatocytes was measured in a coculture system. We measured levels of G-protein-coupled estrogen receptor (GPER1) in HCC and nontumor liver tissues from 68 patients by immunohistochemistry., Results: Exposure to 17β-estradiol (E2) increased proliferation of hepatocytes and liver volume and mass in larval and adult zebrafish. Chemical genetic and epistasis experiments showed that GPER1 mediates the effects of E2 via the phosphoinositide 3-kinase-protein kinase B-mechanistic target of rapamycin pathway: gper1-knockout and mtor-knockout zebrafish did not increase liver growth in response to E2. HCC samples from patients had increased levels of GPER1 compared with nontumor tissue samples; estrogen promoted proliferation of human primary hepatocytes. Estrogen accelerated hepatocarcinogenesis specifically in male zebrafish. Chemical inhibition or genetic loss of GPER1 significantly reduced tumor development in the zebrafish., Conclusions: In an analysis of zebrafish and human liver cells and tissues, we found GPER1 to be a hepatic estrogen sensor that regulates liver growth during development, regeneration, and tumorigenesis. Inhibitors of GPER1 might be developed for liver cancer prevention or treatment., Transcript Profiling: The accession number in the Gene Expression Omnibus is GSE92544., (Copyright © 2019 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Background: Previous studies showed reduction of brain cannabinoid CB 1 receptors in adults with cannabis and alcohol use disorders. Preclinical data suggest that these receptors also contribute to nicotine reward and dependence. Tobacco smoking may confound clinical studies of psychiatric disorders because many patients with such disorders smoke tobacco. Whether human subjects who smoke tobacco but are otherwise healthy have altered CB 1 receptor binding in brain is unknown., Methods: We measured CB 1 receptors in brains of 18 healthy men who smoke tobacco (frequent chronic cigarette smokers), and 28 healthy men who do not smoke tobacco, using positron emission tomography and [ 18 F]FMPEP-d 2 , a radioligand for CB 1 receptors. We collected arterial blood samples during scanning to calculate the distribution volume (V T ), which is nearly proportional to CB 1 receptor density. Repeated-measures analysis of variance compared V T between groups in various brain regions., Results: Brain CB 1 receptor V T was about 20% lower in subjects who smoke tobacco than in subjects who do not. Decreased V T was found in all brain regions, but reduction did not correlate with years of smoking, number of cigarettes smoked per day, or measures of nicotine dependence., Conclusions: Tobacco-smoking healthy men have a widespread reduction of CB 1 receptor density in brain. Reduction of CB 1 receptors appears to be a common feature of substance use disorders. Future clinical studies on the CB 1 receptor should control for tobacco smoking., (Copyright © 2018 Society of Biological Psychiatry. All rights reserved.)

Estrogens regulate vertebrate development and function through binding to nuclear estrogen receptors α and β (ERα and ERβ) and the G protein-coupled estrogen receptor (GPER). Studies in mutant animal models demonstrated that ERα and ERβ are required for normal ovary development and function. However, the degree to which GPER signaling contributes to ovary development and function is less well understood. Previous studies using cultured fish oocytes found that estradiol inhibits oocyte maturation in a GPER-dependent manner, but whether GPER regulates oocyte maturation in vivo is not known. To test the hypothesis that GPER regulates oocyte maturation in vivo, we assayed ovary development and function in gper mutant zebrafish. We found that homozygous mutant gper embryos developed into male and female adults with normal sex ratios. Adult mutant fish exhibited normal secondary sex characteristics and fertility. Additionally, mutant ovaries were histologically normal. We observed no differences in the number of immature versus mature oocytes in mutant versus wild-type ovaries from both young and aged adults. Furthermore, expression of genes associated with sex determination and ovary function was normal in gper mutant ovaries compared with wild type. Our findings suggest that GPER is not required for sex determination, ovary development, or fertility in zebrafish.

Background: Non-alcohol substance use disorders (drug use disorders [DUDs]) are common in trauma patients., Objective: To determine the test characteristics of a 4-item drug CAGE questionnaire to detect DUDs in a cohort of adult trauma inpatients., Methods: Observational cross-sectional cohort of 1,115 adult patients admitted directly to a level-one trauma center between September, 1994 and November, 1996. All participants underwent both a 4-item drug CAGE questionnaire and the substance use disorder section of a structured psychiatric diagnostic clinical interview (SCID) (DSM-IIIR criteria), administered by staff unaware of their clinical status. Sensitivity, specificity, positive (PPV) and negative predictive values (NPV), positive (LR+) and negative likelihood ratios (LR-), and the area under the receiver operating curve (AUC) were calculated for each individual question and the overall questionnaire, using SCID-generated DUD diagnoses as the standard. Performance characteristics of the screen were also compared across selected sociodemographic, injury mechanism, and diagnostic sub-groups., Results: Subjects with DUDs were common (n = 349, 31.3%), including cannabis (n = 203, 18.2%), cocaine (n = 199, 17.8%), and opioids (n = 156, 14.0%). The screen performed well overall (AUC = 0.90, 95% CI: 0.88-0.91) and across subgroups based on age, sex, race, marriage status, income, education, employment status, mechanism of injury, and current/past DUD status (AUCs 0.75-1.00). Answering any one question in the affirmative had a sensitivity = 83.4% (95% CI: 79.1-87.1), specificity = 92.3% (95% CI: 90.2-94.1), PPV = 83.1%, LR+ = 10.8., Conclusions: The 4-item drug CAGE and its individual questions had good-to-excellent ability to detect DUDs in this adult trauma inpatient population, suggesting its usefulness as a screening tool., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

To study the effects of environmental endocrine disruptor compounds (EDCs) on aquatic animals, embryos and larvae are typically incubated in water containing defined concentrations of EDCs. However, the amount of EDC uptake into the animal is often difficult to determine. Using radiolabeled estradiol ([ 3 H]E2), we previously developed a rapid, straightforward assay to measure estradiol uptake from water into zebrafish embryos and larvae. Here, we extend this approach to measure the uptake of two additional EDCs, bisphenol A (BPA) and ethinyl estradiol (EE2). As with E2, the uptake of each compound by individual larvae was low (<6%), and increased with increasing concentration, duration, and developmental stage. We found that E2 and EE2 had similar uptake under equivalent exposure conditions, while BPA had comparatively lower uptake. One application of this assay is to test factors that influence EDC uptake or efflux. It has been suggested that persistent organic pollutants (POPs) inhibit ABC transporters that may normally efflux EDCs and their metabolites, inducing toxicity in aquatic organisms. We measured [ 3 H]E2 levels in zebrafish in the presence or absence of the POP PDBE-100, and cyclosporine A, a known inhibitor of ABC transporters. Neither chemical significantly affected [ 3 H]E2 levels in zebrafish, suggesting that zebrafish maintain estradiol efflux in the presence of PDBE-100, independently of cyclosporine A-responsive transporters. These uptake results will be a valuable reference for EDC exposure studies in developing zebrafish, and provide a rapid assay to screen for chemicals that influence estrogen-like EDC levels in vivo., (Copyright © 2017 Elsevier Inc. All rights reserved.)

In 2005, two groups independently discovered that the G protein-coupled receptor GPR30 binds estradiol in cultured cells and, in response, initiates intracellular signaling cascades Revankar et al. (2005), Thomas et al. (2005). GPR30 is now referred to as GPER, the G-protein coupled estrogen receptor Prossnitz and Arterburn (2015). While studies in animal models are illuminating GPER function, there is controversy as to whether GPER acts as an autonomous estrogen receptor in vivo, or whether GPER interacts with nuclear estrogen receptor signaling pathways in response to estrogens. Here, we review the evidence that GPER acts as an autonomous estrogen receptor in vivo and discuss experimental approaches to test this hypothesis directly. We propose that the degree to which GPER influences nuclear estrogen receptor signaling likely depends on cell type, developmental stage and pathology., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Objective: Cannabis withdrawal has not been studied in adults with attention-deficit/hyperactivity disorder (ADHD) who have high rates of cannabis use. We aimed to describe cannabis withdrawal, motivations to quit, and strategies to quit cannabis use in cannabis-dependent adults with ADHD., Methods: Twenty-three adults with ADHD enrolled in a controlled clinical trial of pharmacotherapy (atomoxetine) for cannabis dependence (DSM-IV criteria) completed the Marijuana Quit Questionnaire (MJQQ) to provide information on their "most serious" quit attempt made without formal treatment. The study was conducted between November 2005 and June 2008., Results: Participants were predominantly male (82.6%, n = 19), with a mean (SD) age of 27.4 (8.5) years (range, 18-53) at the start of their index quit attempt. The most common motive for quitting cannabis was "to save money" (87%, n = 20); the most common strategy to maintain abstinence was "stopped associating with people who smoke marijuana" (43%, n = 10). Almost all (96%, n = 22) subjects reported ≥ 1 cannabis withdrawal symptom; 7 (30%) met DSM-5 diagnostic criteria for cannabis withdrawal syndrome., Conclusions: Participants with comorbid ADHD and cannabis dependence reported withdrawal symptoms similar to other samples of non-treatment-seeking cannabis-dependent adults with no psychiatric comorbidity. These findings suggest that ADHD does not influence cannabis withdrawal in the way that it does tobacco (nicotine) withdrawal., Trial Registration: Data used in this secondary analysis came from ClinicalTrials.gov identifier: NCT00360269., (© Copyright 2018 Physicians Postgraduate Press, Inc.)