Fermented foods: Harnessing their potential to modulate the microbiota-gut-brain axis for mental health.

Over the past two decades, whole food supplementation strategies have been leveraged to target mental health. In addition, there has been increasing attention on the ability of gut microbes, so called psychobiotics, to positively impact behaviour though the microbiota-gut-brain axis. Fermented foods offer themselves as a combined whole food microbiota modulating intervention. Indeed, they contain potentially beneficial microbes, microbial metabolites and other bioactives, which are being harnessed to target the microbiota-gut-brain axis for positive benefits. This review highlights the diverse nature of fermented foods in terms of the raw materials used and type of fermentation employed, and summarises their potential to shape composition of the gut microbiota, the gut to brain communication pathways including the immune system and, ultimately, modulate the microbiota-gut-brain axis. Throughout, we identify knowledge gaps and challenges faced in designing human studies for investigating the mental health-promoting potential of individual fermented foods or components thereof. Importantly, we also suggest solutions that can advance understanding of the therapeutic merit of fermented foods to modulate the microbiota-gut-brain axis.

Telehealth Utilization in High-Risk Pregnancies During COVID-19.

Purpose: To determine how telehealth has influenced outcomes in high-risk obstetrics patients during the Coronavirus disease 2019 (COVID-19) pandemic. Methods: A retrospective chart review was conducted to identify patterns in both telehealth and in-person clinic visits among patients of a Maternal Fetal Medicine (MFM) department from the onset of the COVID-19 pandemic from March 2020 until October 2021. For the descriptive analysis, p-values were calculated using Wilcoxon rank sum for continuous variables and chi-square or Fisher exact (where cell n < 5) for categorical variables. Variables of interest were then tested for their univariate association with telehealth utilization using logistic regression. Variables found to meet the criterion of p < 0.2 in the univariate case were introduced into a multivariable logistic model with a backward elimination for determining variable retention. We aimed to analyze whether telehealth visits significantly impacted pregnancy outcomes. Results: Four hundred nineteen high-risk patients visited the clinic via in-person and/or telehealth appointments during the study period: 320 patients without telehealth visits and 99 patients with telehealth visits. Care provided by telehealth visits was not found to be related to self-reported race (p = 0.81), maternal body mass index (p = 1.0), or maternal age (p = 0.53). Patients with private insurance were more likely to have telehealth visits than patients with public insurance (79.9% vs. 65.5%, p < 0.01). In univariate logistic analyses, patients with diagnoses of anxiety (p < 0.01), asthma (p = 0.03), and depression (p < 0.01), at the time care was established, were more likely to have telehealth visits. Those patients with telehealth visits did not have any statistical differences in mode of delivery (p = 0.2) or pregnancy outcomes (p = 0.12), including fetal demise, preterm delivery, or delivery at term as compared with patients with all in-office visits. In multivariable analysis, patient conditions of anxiety (p < 0.01), maternal obesity (p < 0.01), and twin pregnancy (p = 0.04) were associated with higher rates of telehealth visits. Conclusion: Patients with certain pregnancy complications elected to have more telehealth visits. Patients with private insurance were more likely to have telehealth visits than patients with public insurance. There are benefits for patients with certain pregnancy complications to incorporate telehealth visits in addition to regularly scheduled in-person clinic visits and may be suitable in a post-pandemic setting as well. Further research in this field is needed to better understand the impact of implementing telehealth in high-risk obstetrics patients.

Cholesterol and PIP2 Modulation of BKCa Channels.

Ca2+/voltage-gated, large conductance K+ channels (BKCa) are formed by homotetrameric association of α (slo1) subunits. Their activity, however, is suited to tissue-specific physiology largely due to their association with regulatory subunits (ß and γ types), chaperone proteins, localized signaling, and the channel's lipid microenvironment. PIP2 and cholesterol can modulate BKCa activity independently of downstream signaling, yet activating Ca2+i levels and regulatory subunits control ligand action. At physiological Ca2+i and voltages, cholesterol and PIP2 reduce and increase slo1 channel activity, respectively. Moreover, slo1 proteins provide sites that seem to recognize cholesterol and PIP2: seven CRAC motifs in the slo1 cytosolic tail and a string of positively charged residues (Arg329, Lys330, Lys331) immediately after S6, respectively. A model that could explain the modulation of BKCa activity by cholesterol and/or PIP2 is hypothesized. The roles of additional sites, whether in slo1 or BKCa regulatory subunits, for PIP2 and/or cholesterol to modulate BKCa function are also discussed.

Phylodynamics of deer tick virus in North America.

The burden of ticks and the pathogens they carry is increasing worldwide. Powassan virus (POWV; Flaviviridae: Flavivirus), the only known North American tick-borne flavivirus, is of particular concern due to rising cases and the severe morbidity of POWV encephalitis. Here, we use a multifaceted approach to evaluate the emergence of the II POWV lineage, known as deer tick virus (DTV), in parts of North America where human cases occur. We detected DTV-positive ticks from eight of twenty locations in the Northeast USA with an average infection rate of 1.4 per cent. High-depth, whole-genome sequencing of eighty-four POWV and DTV samples allowed us to assess geographic and temporal phylodynamics. We observed both stable infection in the Northeast USA and patterns of geographic dispersal within and between regions. A Bayesian skyline analysis demonstrated DTV population expansion over the last 50 years. This is concordant with the documented expansion of Ixodes scapularis tick populations and suggests an increasing risk of human exposure as the vector spreads. Finally, we isolated sixteen novel viruses in cell culture and demonstrated limited genetic change after passage, a valuable resource for future studies investigating this emerging virus.

Jamestown Canyon Virus in Collected Mosquitoes, Maine, United States, 2017-2019.

Jamestown Canyon virus (JCV) is a mosquito-borne arbovirus that circulates in North America. We detected JCV in 4 pools of mosquitoes collected from midcoastal Maine, USA, during 2017-2019. Phylogenetic analysis of a JCV sequence obtained from Aedes cantator mosquitoes clustered within clade A, which also circulates in Connecticut, USA.

Differential distribution of cholesterol pools across arteries under high-cholesterol diet.

Excessive cholesterol constitutes a major risk factor for vascular disease. Within cells, cholesterol is distributed in detergent-sensitive and detergent-resistant fractions, with the largest amount of cholesterol residing in cellular membranes. We set out to determine whether various arteries differ in their ability to accumulate esterified and non-esterified cholesterol in detergent-sensitive versus detergent-resistant fractions throughout the course of a high-cholesterol diet. Male Sprague-Dawley rats were placed on 2 % cholesterol diet while a control group was receiving iso-caloric standard chow. Liver, aorta, and pulmonary, mesenteric, and cerebral arteries were collected at 2-6, 8-12, 14-18, and 20-24 weeks from the start of high-cholesterol diet. After fraction separation, esterified and free non-esterified cholesterol levels were measured. In all arteries, largest cholesterol amounts were present in detergent-sensitive fractions in the non-esterified form. Overall, cholesterol in aorta and cerebral arteries was elevated during 14-18 weeks of high-cholesterol diet. Cerebral arteries also exhibited increase in esterified cholesterol within detergent-sensitive domains, as well as increase in cholesterol level in the detergent-resistant fraction at earlier time-points of diet. Pulmonary artery and mesenteric artery were largely resistant to cholesterol accumulation. Quantitative polymerase chain reaction (qPCR) analysis revealed up-regulation of low-density lipoprotein receptor (Ldlr) and low-density lipoprotein receptor-related protein 1 (Lrp1) gene expression in cerebral arteries when compared to mesenteric and pulmonary arteries, respectively. In summary, we unveiled the differential ability of arteries to accumulate cholesterol over the course of a high-cholesterol diet. The differential accumulation of cholesterol seems to correlate with the up-regulated gene expression of proteins responsible for cholesterol uptake.

Utility of an experimental medicine model to evaluate efficacy, side-effects and mechanism of action of novel treatments for obesity and binge-eating disorder.

Obesity and Binge-Eating Disorder (BED) are prevalent conditions that are associated with increased risk of morbidity and mortality. There is evidence that the use of pharmacotherapy alongside behavioural treatments can improve quality of life and reduce disease risk for patients with these disorders. However, there are few approved drug therapies for obesity, and these are limited by poor efficacy and/or side effects and only one drug has been approved for the treatment of BED. There is considerable potential to use experimental medicine models to identify new drug treatments for obesity and BED, with greater efficacy and an improved side effect profile, at an early stage of development. Here, we present a model developed in our laboratory that incorporates both behavioural and neuroimaging measures which can be used to facilitate drug development for obesity and BED. The results from validation studies conducted to date using our model suggest that it is sensitive to the effects of agents with behavioural, neurophysiological and neuropharmacological mechanisms of action known to be associated with weight loss and reductions in binge eating. Future studies using the model will be valuable to evaluate the potential efficacy and side-effects of new candidate drugs at an early stage in the development pipeline for both obesity and BED.

The effect of intranasal insulin on appetite and mood in women with and without obesity: an experimental medicine study.

BACKGROUND/OBJECTIVES: Intranasal (IN) administration of insulin decreases appetite in humans, but the underlying mechanisms are unclear, and it is unknown whether IN insulin affects the food intake of women with obesity. SUBJECTS/METHODS: In a double-blind, placebo-controlled, crossover design, participants (35 lean women and 17 women with obesity) were randomized to receive 160 IU/1.6 mL of IN insulin or placebo in a counterbalanced order in the post prandial state. The effects of IN insulin on cookie intake, appetite, mood, food reward, cognition and neural activity were assessed. RESULTS: IN insulin in the post prandial state reduced cookie intake, appetite and food reward relative to placebo and these effects were more pronounced for women with obesity compared with lean women. IN insulin also improved mood in women with obesity. In both BMI groups, IN insulin increased neural activity in the insula when viewing food pictures. IN insulin did not affect cognitive function. CONCLUSIONS: These results suggest that IN insulin decreases palatable food intake when satiated by reducing food reward and that women with obesity may be more sensitive to this effect than lean women. Further investigation of the therapeutic potential of IN insulin for weight management in women with obesity is warranted.

The association of hospital research publications and clinical quality.

OBJECTIVE: To assess the quantity and impact of research publications among US acute care hospitals; to identify hospital characteristics associated with publication volumes; and to estimate the independent association of bibliometric indicators with Hospital Compare quality measures. DATA SOURCES: Hospital Compare; American Hospital Association Survey; Magnet Recognition Program; Science Citation Index Expanded. STUDY DESIGN: In cross-sectional studies using a 40% random sample of US Medicare-participating hospitals, we estimated associations of hospital characteristics with publication volumes and associations of hospital-linked bibliometric indicators with 19 Hospital Compare quality metrics. DATA COLLECTION/EXTRACTION METHODS: Using standardized search strategies, we identified all publications attributed to authors from these institutions from January 1, 2015 to December 31, 2016 and their subsequent citations through July 2020. PRINCIPAL FINDINGS: Only 647 of 1604 study hospitals (40.3%) had ≥1 publication. Council of Teaching Hospitals and Health Systems (COTH) hospitals had significantly more publications (average 599 vs. 11 for non-COTH teaching and 0.6 for nonteaching hospitals), and their publications were cited more frequently (average 22.6/publication) than those from non-COTH teaching (18.2 citations) or nonteaching hospitals (12.8 citations). In multivariable regression, teaching intensity, hospital beds, New England or Pacific region, and not-for-profit or government ownership were significant predictors of higher publication volumes; the percentage of Medicaid admissions was inversely associated. In multivariable linear regression, hospital publications were associated with significantly lower risk-adjusted mortality rates for acute myocardial infarction (coefficient -0.52, p = 0.01), heart failure (coefficient -0.74, p = 0.004), pneumonia (coefficient -1.02, p = 0.001), chronic obstructive pulmonary disease (coefficient -0.48, p = 0.005), and coronary artery bypass surgery (coefficient -0.73, p < 0.0001); higher overall Hospital Consumer Assessment of Healthcare Providers and Systems (HCAHPS) ratings (coefficient 2.37, p = 0.04); and greater patient willingness to recommend (coefficient 3.38, p = 0.01). CONCLUSIONS: A minority of US hospitals published in the biomedical literature. Publication quantity and impact indicators are independently associated with lower risk-adjusted mortality and higher HCAHPS scores.

The effects of lisdexamfetamine dimesylate on eating behaviour and homeostatic, reward and cognitive processes in women with binge-eating symptoms: an experimental medicine study.

Lisdexamfetamine dimesylate (LDX) is the only drug currently approved by the FDA for the treatment of Binge-Eating Disorder (BED), but little is known about the behavioural mechanisms that underpin the efficacy of LDX in treating BED. We examined the behavioural and neural effects of an acute dose of LDX (50 mg) in 22 women with binge-eating symptomatology using a randomised, crossover, double-blind, placebo-controlled experimental medicine design. LDX reduced self-reported appetite ratings and intake of both a pasta meal and a palatable cookie snack. LDX also decreased the eating rate of pasta but not of cookies and reduced self-reported liking ratings for pasta at the end of the meal. When viewing food pictures during an fMRI scan, LDX reduced activity bilaterally in the thalamus. LDX enhanced sustained attention and reduced impulsive responding in a continuous performance task but had no effect on emotional bias or working memory. These results suggest the observed effects of LDX on food intake (and by implication the efficacy of LDX in treating BED) may be related to the actions of the drug to enhance satiety, reduce food-related reward responding when full and/or increase cognitive control. Novel pharmacotherapies for BED might be most effective if they have a broad spectrum of effects on appetite, reward and cognition.

Comparing two seized drug workflows for the analysis of synthetic cannabinoids, cathinones, and opioids.

As the challenges faced by drug chemists persist, due to the presence of emerging drugs, laboratories continue to look for new solutions, ranging from existing methods to implementation of entirely new technology. A common barrier for making workflow changes is a lack of pre-existing data demonstrating the potential impact of these changes. In this study, we compare, qualitatively and quantitatively, an existing workflow for seized drug analysis to an experimental workflow. Four chemists were asked to analyze a total of 50 mock case samples across the two workflows. The existing workflow employed color tests for screening alongside general purpose GC-FID and GC-MS analyses for confirmation. The experimental workflow combined DART-MS screening with class-specific (targeted) GC-MS analysis for confirmation. Comparison of the workflows showed that screening by DART-MS required the same amount of time as color tests but yielded more accurate and specific information. Confirmation using the existing workflow required more than twice the amount of instrument time and data interpretation time while also presenting other analytical challenges that prevented compound confirmation in select samples. Targeted GC-MS methods simplified data interpretation, reduced consumption of reference materials, and addressed almost all limitations of general-purpose methods. While the experimental workflow requires modifications and answering of additional research questions, this study shows how rethinking analytical workflows for seized drug analysis could reduce turnaround times, backlogs, and standards consumption. It also demonstrates the potential impact of being able to investigate workflow changes prior to implementation.

Emergence of Ixodes scapularis (Acari: Ixodidae) in a Small Mammal Population in a Coastal Oak-Pine Forest, Maine, USA.

In the United States, surveillance has been key to tracking spatiotemporal emergence of blacklegged ticks [Ixodes scapularis Say (Ixodida:Ixodidae)] and their pathogens such as Borrelia burgdorferi Johnson, Schmid, Hyde, Steigerwalt & Brenner (Spirochaetales: Spirochaetaceae), the agent of Lyme disease. On the Holt Research Forest in midcoastal Maine, collection of feeding ticks from live-trapped small mammal hosts allowed us to track the emergence and establishment of I. scapularis, 1989-2019. From 1989-1995, we collected only I. angustus Neumann (Ixodida: Ixodidae)(vole tick), Dermacentor variabilis Say (Ixodida: Ixodidae) (American dog tick), and I. marxi Banks (Ixodida: Ixodidae) (squirrel tick) from seven species of small mammals. The most abundant tick host was the white-footed mouse [Peromyscus leucopus Rafinesque (Rodentia:Cricetidae)] followed by the red-backed vole (Myodes gapperi Vigors (Rodentia: Cricetidae)). Emergence of I. scapularis was signaled via the appearance of subadult I. scapularis in 1996. Emergence of B. burgdorferi was signaled through its appearance in I. scapularis feeding on mice in 2005. There was a substantial increase in I. scapularis prevalence (proportion of hosts parasitized) and burdens (ticks/host) on white-footed mice and red-backed voles in 2007. The ~11-yr time-to-establishment for I. scapularis was consistent with that seen in other studies. White-footed mice comprised 65.9% of all captures and hosted 94.1% of the total I. scapularis burden. The white-footed mouse population fluctuated interannually, but did not trend up as did I. scapularis prevalence and burdens. There were concurrent declines in I. angustus and D. variabilis. We discuss these results in the broader context of regional I. scapularis range expansion.

Lisdexamfetamine and binge-eating disorder: A systematic review and meta-analysis of the preclinical and clinical data with a focus on mechanism of drug action in treating the disorder.

Binge-Eating Disorder (BED) is the most common eating disorder in the United States. Lisdexamfetamine (LDX) was approved in 2015 by the FDA for treatment of BED and is the only drug approved for treating the disorder. There has been no systematic evaluation of the published clinical and preclinical evidence for efficacy of LDX in treating BED and the mechanisms responsible for the therapeutic action of the drug. To address this gap, we conducted a systematic review and meta-analysis using PRISMA guidelines. Fourteen clinical and seven preclinical articles were included. There is consistent evidence from clinical studies that LDX is an effective treatment for BED and that the drug reduces the BED symptoms and body weight of patients with the disorder. There is also consistent evidence from preclinical studies that LDX reduces food intake but no consistent evidence for a preferential reduction of palatable food consumption by the drug in rodents. The evidence on mechanism of action is more limited and suggests LDX may reduce binge eating by a combination of effects on appetite/satiety, reward, and cognitive processes, including attention and impulsivity/inhibition, that are mediated by catecholamine and serotonin mechanisms in the brain. There is an urgent need for adequately powered, placebo-controlled, behavioural and neuroimaging studies with LDX (recruiting patients and/or individuals with subclinical BED symptoms) to further investigate the mechanism of action of the drug in treating BED. An improved understanding of the behavioural and neurochemical mechanisms of action of LDX could lead to the development of improved drug therapies to treat BED.

Association of Surgeon Representation on NIH Study Sections With Receipt of Funding by Surgeon-scientists.

OBJECTIVE: Our goal was to evaluate the relationship between surgeon representation on NIH study sections and success in grant funding. SUMMARY OF BACKGROUND DATA: NIH funding for surgeon-scientists is declining. Prior work has called for increased surgeon participation in the grant review process as a strategy to increase receipt of funding by surgeon-scientists. METHODS: A retrospective review of surgeon (primary department: General, Urology, Orthopedic, Ophthalmology, Otolaryngology, Neurosurgery) representation on NIH study sections and receipt of funding was performed using NIH Research Portfolio Online Reporting Tools Expenditures and Results (RePORTER) and 2019 Blue Ridge Institute for Medical Research data. NIH chartered study section panels and ad hoc reviewers for each 2019 review date were also obtained. RESULTS: In 2019, 9239 individuals reviewed in at least 1 of the 168 study sections [190 (2.1%) surgeons, 64 (0.7%) standing members, 126 (1.4%) ad-hoc]. Most surgeons on study sections were male (65%) professors (63%). Surgeons most commonly served on bioengineering, technology, and surgical sciences (29.6% surgeons), diseases and pathophysiology of the visual system (28.3%), and surgery, anesthesiology and trauma (21%). In 2019, 773 surgeons received 1235 NIH grants (>$580 M) out of a total of 55,012 awards (2.2%). Funded surgeons were predominantly male (79%), White (68%), non-Hispanic (97%), full professors (50%), and 43% had additional advanced degrees (MPH/PhD/MBA). surgery, anesthesiology and trauma, diseases and pathophysiology of the visual system, and bioengineering, technology, and surgical sciences were the most common study sections that reviewed funded grants to surgeon-scientists. Ninety-two surgeons both received grant funding and served on study section. Study sections with higher surgeon representation were more likely to fund surgeon-scientists (P < 0.001). CONCLUSIONS: Surgeon representation on NIH study sections is strongly associated with receipt of funding by surgeon-scientists. Increasing NIH study section representation by surgeons may help to preserve the surgeon-scientist phenotype.

IL-1ß-MyD88-mTOR Axis Promotes Immune-Protective IL-17A+Foxp3+ Cells During Mucosal Infection and Is Dysregulated With Aging.

CD4+Foxp3+Tregs maintain immune homeostasis, but distinct mechanisms underlying their functional heterogeneity during infections are driven by specific cytokine milieu. Here we show that MyD88 deletion in Foxp3+ cells altered their function and resulted in increased fungal burden and immunopathology during oral Candida albicans (CA) challenge. Excessive inflammation due to the absence of MyD88 in Tregs coincided with a reduction of the unique population of IL-17A expressing Foxp3+ cells (Treg17) and an increase in dysfunctional IFN-γ+/Foxp3+ cells (TregIFN-γ) in infected mice. Failure of MyD88-/- Tregs to regulate effector CD4+ T cell functions correlated with heightened levels of IFN-γ in CD4+ T cells, as well as increased infiltration of inflammatory monocytes and neutrophils in oral mucosa in vivo. Mechanistically, IL-1ß/MyD88 signaling was required for the activation of IRAK-4, Akt, and mTOR, which led to the induction and proliferation of Treg17 cells. In the absence of IL-1 receptor signaling, Treg17 cells were reduced, but IL-6-driven expansion of TregIFN-γ cells was increased. This mechanism was physiologically relevant during Candida infection in aged mice, as they exhibited IL-1 receptor/MyD88 defect in Foxp3+ cells, loss of p-mTORhighTreg17 cells and reduced levels of IL-1ß in oral mucosa, which coincided with persistent tongue inflammation. Concurrent with Treg dysfunction, aging was associated with increased CD4+ T cell hyperactivation and heightened levels of IL-6 in mice and humans in oral mucosa in vivo. Taken together, our data identify IL-1ß/MyD88/Treg axis as a new component that modulates inflammatory responses in oral mucosa. Also, dysregulation of this axis in an aging immune system may skew host defense towards an immunopathological response in mucosal compartments.

Microbiome Dependent Regulation of Tregs and Th17 Cells in Mucosa.

Mammals co-exist with resident microbial ecosystem that is composed of an incredible number and diversity of bacteria, viruses and fungi. Owing to direct contact between resident microbes and mucosal surfaces, both parties are in continuous and complex interactions resulting in important functional consequences. These interactions govern immune homeostasis, host response to infection, vaccination and cancer, as well as predisposition to metabolic, inflammatory and neurological disorders. Here, we discuss recent studies on direct and indirect effects of resident microbiota on regulatory T cells (Tregs) and Th17 cells at the cellular and molecular level. We review mechanisms by which commensal microbes influence mucosa in the context of bioactive molecules derived from resident bacteria, immune senescence, chronic inflammation and cancer. Lastly, we discuss potential therapeutic applications of microbiota alterations and microbial derivatives, for improving resilience of mucosal immunity and combating immunopathology.

Role of Short Chain Fatty Acids in Controlling Tregs and Immunopathology During Mucosal Infection.

Interactions between mucosal tissues and commensal microbes control appropriate host immune responses and inflammation, but very little is known about these interactions. Here we show that the depletion of resident bacteria using antibiotics (Abx) causes oral and gut immunopathology during oropharyngeal candidiasis (OPC) infection. Antibiotic treatment causes reduction in the frequency of Foxp3+ regulatory cells (Tregs) and IL-17A producers, with a concomitant increase in oral tissue pathology. While C. albicans (CA) is usually controlled in the oral cavity, antibiotic treatment led to CA dependent oral and gut inflammation. A combination of short chain fatty acids (SCFA) controlled the pathology in Abx treated mice, correlating to an increase in the frequency of Foxp3+, IL-17A+, and Foxp3+IL-17A+ double positive (Treg17) cells in tongue and oral draining lymph nodes. However, SCFA treatment did not fully reverse the gut inflammation suggesting that resident microbiota have SCFA independent homeostatic mechanisms in gut mucosa. We also found that SCFA potently induce Foxp3 and IL-17A expression in CD4+ T cells, depending on the cytokine milieu in vitro. Depletion of Tregs alone in FDTR mice recapitulated oral inflammation in CA infected mice, showing that Abx mediated reduction of Tregs was involved in infection induced pathology. SCFA did not control inflammation in Treg depleted mice in CA infected FDTR mice, showing that Foxp3+ T cell induction was required for the protective effect mediated by SCFA. Taken together, our data reveal that SCFA derived from resident bacteria play a critical role in controlling immunopathology by regulating T cell cytokines during mucosal infections. This study has broader implications on protective effects of resident microbiota in regulating pathological infections.

Using c-Jun to identify fear extinction learning-specific patterns of neural activity that are affected by single prolonged stress.

Neural circuits via which stress leads to disruptions in fear extinction is often explored in animal stress models. Using the single prolonged stress (SPS) model of post traumatic stress disorder and the immediate early gene (IEG) c-Fos as a measure of neural activity, we previously identified patterns of neural activity through which SPS disrupts extinction retention. However, none of these stress effects were specific to fear or extinction learning and memory. C-Jun is another IEG that is sometimes regulated in a different manner to c-Fos and could be used to identify emotional learning/memory specific patterns of neural activity that are sensitive to SPS. Animals were either fear conditioned (CS-fear) or presented with CSs only (CS-only) then subjected to extinction training and testing. C-Jun was then assayed within neural substrates critical for extinction memory. Inhibited c-Jun levels in the hippocampus (Hipp) and enhanced functional connectivity between the ventromedial prefrontal cortex (vmPFC) and basolateral amygdala (BLA) during extinction training was disrupted by SPS in the CS-fear group only. As a result, these effects were specific to emotional learning/memory. SPS also disrupted inhibited Hipp c-Jun levels, enhanced BLA c-Jun levels, and altered functional connectivity among the vmPFC, BLA, and Hipp during extinction testing in SPS rats in the CS-fear and CS-only groups. As a result, these effects were not specific to emotional learning/memory. Our findings suggest that SPS disrupts neural activity specific to extinction memory, but may also disrupt the retention of fear extinction by mechanisms that do not involve emotional learning/memory.

Self-Association of Rafoxanide in Aqueous Media and Its Application in Preparing Amorphous Solid Dispersions.

Our primary objective is to characterize the self-association of rafoxanide in alkaline media. The second objective is to illustrate the feasibility of using rafoxanide micellar solution as the feed solution to prepare amorphous solid dispersion via spray drying. Rafoxanide is a poorly water-soluble drug. It is a weak acid, and its poor aqueous solubility is due to its hydrophobicity. The surface-active property of rafoxanide has not been previously reported. It was discovered that the addition of a small percentage of organic solvents is required to elevate the solubility of rafoxanide above the critical micelle concentration to allow for the formation of micelles. Our fluorescence decay study confirms the self-association of rafoxanide in a cosolvent consisting of 70%, v/v, NaOH solution and 30%, v/v, acetone. The position of each functional group in the micellar structures using the 1H NMR technique was identified. The critical micelle concentration of rafoxanide in the cosolvent is determined to be 302 µg/mL using a surface tension method. The solubility of rafoxanide in 0.1 N NaOH solution is less than 11 µg/mL. Interestingly, the apparent solubility increased to 38,400 µg/mL in the presence of 30% acetone as the result of micelle formation. This unique solubility characteristic makes it feasible to prepare rafoxanide amorphous solid dispersions by spray drying a predominantly aqueous (70% 0.1 N NaOH solution and 30% acetone) based feed solution. Povidone and copovidone were both used as polymeric carriers. Based on solid-state characterization, including differential scanning calorimetry, X-ray powder diffraction, and hot-stage polarized light microscopy, our results indicate that rafoxanide solid dispersions prepared using this novel process are amorphous. Approximately 750-fold increase in the concentration of rafoxanide in aqueous media at pH 6.8 was achieved with the amorphous solid dispersions.