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Michelle L Witkop,1 Angela Lambing,2 Cynthia D Nichols,3 James E Munn,4 Terry L Anderson,5 Bartholomew J Tortella5 1Nursing Department, Northern Regional Bleeding Disorders Center, Munson Medical Center, Traverse City, MI, USA; 2Nursing Department, Henry Ford Hemophilia and Thrombosis Treatment Center, Detroit, MI, USA; 3Nursing Department, Munson Medical Center, Traverse City, MI, USA; 4Hemophilia & Coagulation Disorders Program, University of Michigan Hemophilia Treatment Center, Ann Arbor, MI, USA; 5Medical Affairs, Pfizer Inc, Collegeville, PA, USACorrespondence: Michelle L WitkopNational Hemophilia Foundation, 7 Penn Plaza, Suite 1204, New York, NY 10001, USATel +1 347 541 2963Fax +1 212 328 3777Email mwitkop@hemophilia.orgPurpose: Depression, anxiety, pain, and treatment adherence have reciprocal effects not characterized extensively in hemophilia. This study explored the relationships between depression, anxiety, chronic pain, and treatment adherence in adults with hemophilia.Patients and methods: Adults with self-reported hemophilia A or B completed the cross-sectional IMPACT QoL II survey. Depression (9-item Patient Health Questionnaire [PHQ-9]), anxiety (7-item Generalized Anxiety Disorder scale [GAD-7]), chronic pain (Faces Pain Scale–Revised [FPS-R]), social support (Duke UNC Functional Social Support questionnaire), level of pain control, clotting factor treatment adherence (VERITAS-Pro or -PRN), and previous depression/anxiety were analyzed.Results: Among 200 participants (male, 77.3%; female, 22.8%), 54% had PHQ-9 and 52% had GAD-7 scores indicating moderate to severe depression or anxiety without diagnosis of either disorder. Participants with PHQ-9 scores ≥10 (moderate to severe depression) were more likely to have lower treatment adherence than those with PHQ-9 scores

Functional selectivity in the context of serotonin 2A (5-HT 2A ) receptor agonists is often described as differences psychedelic compounds have in the activation of Gq vs β-arrestin signaling in the brain and how that may relate to inducing psychoactive and hallucinatory properties with respect to each other. However, the presence of 5-HT 2A receptors throughout the body in several cell types, including endothelial, endocrine, and immune-related tissues, suggests that functional selectivity may exist in the periphery as well. Here, we examine functional selectivity between two 5-HT 2A receptor agonists of the phenylalkylamine class: ( R )-2,5-dimethoxy-4-iodoamphetamine [( R )-DOI] and ( R )-2,5-dimethoxy-4-trifluoromethylamphetamine [( R )-DOTFM]. Despite comparable in vitro activity at the 5-HT 2A receptor as well as similar behavioral potency, ( R )-DOTFM does not exhibit an ability to prevent inflammation or elevated airway hyperresponsiveness (AHR) in an acute murine ovalbumin-induced asthma model as does ( R )-DOI. Furthermore, there are distinct differences between protein expression and inflammatory-related gene expression in pulmonary tissues between the two compounds. Using ( R )-DOI and ( R )-DOTFM as tools, we further elucidated the anti-inflammatory mechanisms underlying the powerful anti-inflammatory effects of certain psychedelics and identified key mechanistic components of the anti-inflammatory effects of psychedelics, including suppression of arginase 1 expression., Competing Interests: The authors declare the following competing financial interest(s): CDN, TPF and ALH had a sponsored research contract with Eleusis Therapeutics and CDN and TPF were members of its scientific advisory board when portions of this study were performed. CDN and TPF are founders of 2A Biosciences and receive sponsored research funding from 2A Biosciences., (© 2024 American Chemical Society.)

5-MeO-DMT is a tryptamine being developed as a potential antidepressant that may display a distinct therapeutic mechanism due to its unique pharmacology and subjective effects compared to typical psychedelics. In this article, we parallel the relatively distinct phenomenology and behavioral effects of the acute and post-acute effects of 5-MeO-DMT to those induced by epileptiform activity, particularly in instances within epileptogenic zones of the temporal lobes. This is done by reviewing aberrant 5-HT 1A receptor functioning in epilepsy, noting that 5-MeO-DMT has notable 5-HT 1A receptor agonist properties-and then comparing the acute behavioral and subjective effects induced by 5-MeO-DMT to those that occur in seizures. It might be that 5-MeO-DMT's therapeutic mechanism is partly mediated by evoking temporary epileptiform activity, suggesting a similarity to electroconvulsive therapy. It is also noted that "reactivations," the sudden re-experiencing of drug effects common after 5-MeO-DMT but not after typical psychedelics, may suggest that 5-MeO-DMT produces recurrent epileptiform activity. Overall, this review indicates that further evaluation of 5-MeO-DMT's unique mechanisms in research settings and among naturalistic users are warranted., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Background: Objectives were to 1) assess COVID-19-associated medical and psychological challenges facing persons with inherited bleeding disorders (PIBD) and their parents/guardians (PG) in Germany, the US, and the UK; 2) describe similarities and differences among these countries; 3) identify needs and opportunities for intervention by patient advocacy organizations (PAGs)., Research Design & Methods: A cross-sectional, international survey was conducted in three countries using validated psychometric instruments and investigator-developed items., Results: Five hundred and four surveys were included. Significant differences between countries were found including experiences with medical care, specific thoughts, and concerns about COVID-19, anxiety, and other mental health measures, as well as resources used to cope with stress. Age, education, income, race, IBD diagnosis, PIBD vs. BD group, and gender had moderating effects on resources used. Communication with friends/relatives and use of PAG and HTCs as resources for information/coping decreased in all countries during the pandemic., Conclusions: There were similarities and differences between respondents across the country in the perceived impact of the pandemic, mental health scores, and strategies used to cope with stress.  Recommendations: strategies to increase PAG access for PIBD and their PG during pandemics and natural disasters, ongoing assessment and adaptation to provide supportive resources to specific patient subgroups.

Psychedelic compounds have shown extraordinary potential in treating a wide range of neuropsychiatric disorders. Psilocybin, for example, has now been shown in several clinical trials to induce a rapid (within days) and persistent (3-12 months) improvement in human treatment-resistant depression and other neuropsychiatric conditions. Here we review the preclinical models and experimental approaches that have been used to study the neurobiological actions of psychedelic drugs. We further summarize the insights these studies have provided into the possible mechanisms underlying the induction of their therapeutic actions, including the receptors to which psychedelics bind and the second messenger signaling cascades that they activate. We also discuss potential biological processes that psychedelics may alter to produce the lasting amelioration of symptoms, including improvements in synaptic structure and function and suppression of inflammation. Improved mechanistic understanding of psychedelic drug actions will aid in the advancement of these promising new medicines. This article is part of the Special Issue on "National Institutes of Health Psilocybin Research Speaker Series"., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

G protein-coupled receptors (GPCR) and receptor tyrosine kinases (RTK) modulate vascular tone and contraction via rapid and long-term processes. Sustained activation of these receptor types can change vascular structure, and the ability of vasculature to adapt to high pressure. In this study, the interaction between serotonin (5-HT) receptors and epidermal growth factor receptors (EGFR) on vasoconstriction and the mechanisms of EGFR transactivation and its downstream mediators were investigated. We measured 5-HT-induced vasoconstriction in the aorta and the mesenteric artery; and the effects of EGFR, Src and PI3K, and their downstream mediators Erk1/2 and Akt phosphorylation on 5-HT-mediated vasoconstriction in the presence or absence of pharmacological inhibitors of Ca 2+ /CaM, EGFR, Src, and PI3K. Furthermore, we determined the contribution of 5-HT receptor subtypes to 5-HT-induced vasoconstriction and EGFR transactivation using selective 5-HT 2A and 5-HT 1B receptors ligands. Our results show that EGFR, Src, and PI3K are involved in 5-HT-induced vasoconstriction both in the aorta and the mesenteric artery, and that these kinases have a more prominent role in the mesenteric artery than the aorta. With regard to EGFR transactivation by 5-HT, Ca 2+ /CaM, Src and PI3K are upstream mediators, and transactivation is partly mediated by Erk1/2 and Akt activation. Furthermore, Ca 2+ /CaM, Src, and PI3K are the main regulators for Akt activation, however Src only has a prominent role for Erk1/2 activation. 5-HT 2A and 5-HT 1B receptors have different EGFR transactivation profiles through Src and/or PI3K, with 5-HT 2A having a greater role than 5-HT 1B receptors., Competing Interests: Declaration of Competing Interest 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., (Copyright © 2022 Elsevier Inc. All rights reserved.)

New medicines containing classic hallucinogenic and entactogenic psychedelic substance are under development for various psychiatric and neurological disorders. Many of these, including psilocybin, lysergic acid diethylamide (LSD), and 3,4-methylenedioxymethamphetamine (MDMA) are Schedule I controlled substances of the United States Controlled Substances Act (US CSA), and similarly controlled globally. The implications of the CSA for research and medicines development, the path to approval of medicines, and their subsequent removal from Schedule I in the US are discussed. This entire process occurs within the framework of the CSA in the US and its counterparts internationally in accordance with international drug control treaties. Abuse potential related research in the US informs the eight factors of the CSA which provide the basis for rescheduling actions that must occur upon approval of a drug that contains a Schedule I substance. Abuse-related research also informs drug product labeling and the risk evaluation and mitigation strategies (REMS) will likely be required for approved medicines. Human abuse potential studies typically employed in CNS drug development may be problematic for substances with strong hallucinogenic effects such as psilocybin, and alternative strategies are discussed. Implications for research, medicinal development, and controlled substance scheduling are presented in the context of the US CSA and FDA requirements with implications for global regulation. We also discuss how abuse-related research can contribute to understanding mechanisms of action and therapeutic effects as well as the totality of the effects of the drugs on the brain, behavior, mood, and the constructs of spirituality and consciousness., (Copyright © 2022. Published by Elsevier Ltd.)

Psychedelic drugs are gaining attention from the scientific community as potential new compounds for the treatment of psychiatric diseases such as mood and substance use disorders. The 5-HT 2A receptor has been identified as the main molecular target, and early studies pointed to an effect on the expression of neuroplasticity genes. Analysing RNA-seq data from the prefrontal cortex of rats chronically treated with lysergic acid diethylamide (LSD), we describe the psychedelic-induced rewiring of gene co-expression networks, which become less centralised but more complex, with an overall increase in signalling entropy typical of highly plastic systems. Intriguingly, signalling entropy mirrors, at the molecular level, the increased brain entropy reported through neuroimaging studies in human, suggesting the underlying mechanisms of higher-order phenomena. Moreover, from the analysis of network topology, we identify potential transcriptional regulators and propose the involvement of different cell types in psychedelics' activity., (© 2021 The Authors. Journal of Neurochemistry published by John Wiley & Sons Ltd on behalf of International Society for Neurochemistry.)

Psilocybin has been shown to be a powerful, long-lasting antidepressant in human clinical trials and in rodent models. Although rodents have commonly been used to model psychiatric disorders, Drosophila have neurotransmitter systems similar to mammals and many comparable brain structures involved in similar behaviors. The forced swim test (FST), which has been used extensively to evaluate compounds for antidepressant efficacy, has recently been adapted for Drosophila. The fly FST has potential to be a cost-effective, high-throughput assay for evaluating potential antidepressants. For this study we pharmacologically validated the fly FST using methamphetamine, DL-α-methyltyrosine, and the antidepressant citalopram. While methamphetamine and DL-α-methyltyrosine altered overall locomotor activity in the Drosophila Activity Monitor System (DAMS), they had no significant impact on measures of immobility in the FST. Conversely, chronic citalopram decreased measures of immobility in the FST in both sexes without increasing DAMS activity. We used the validated FST to evaluate the antidepressant-like effects of high (3.5 mM) and low (0.03 mM) doses of psilocybin. Both doses of psilocybin significantly reduced measures of immobility in male flies, but not females. 0.03 mM had an effect size comparable to chronic citalopram, and 3.5 mM had an effect size approximately twice that of chronic citalopram., (© 2022. The Author(s).)

Herpes simplex virus-associated diseases are a complex interaction between cytolytic viral replication and inflammation. Within the normally avascular and immunoprivileged cornea, HSV ocular infection can result in vision-threatening immune-mediated herpetic keratitis, the leading infectious cause of corneal blindness in the industrialized world. Viral replicative processes are entirely dependent upon numerous cellular biosynthetic and metabolic pathways. Consistent with this premise, HSV infection was shown to profoundly alter gene expression associated with cellular amino acid biosynthetic pathways, including key tryptophan metabolism genes. The essential amino acid tryptophan is crucial for pathogen replication, the generation of host immune responses, and the synthesis of neurotransmitters, such as serotonin. Intriguingly, Tryptophan hydroxylase 2 (TPH2), the neuronal specific rate-limiting enzyme for serotonin synthesis, was the most significantly upregulated gene by HSV in an amino acid metabolism PCR array. Despite the well-defined effects of serotonin in the nervous system, the association of peripheral serotonin in disease-promoting inflammation has only recently begun to be elucidated. Likewise, the impact of serotonin on viral replication and ocular disease is also largely unknown. We therefore examined the effect of HSV-induced serotonin-associated synthesis and transport pathways on HSV-1 replication, as well as the correlation between HSV-induced ocular serotonin levels and disease severity. HSV infection induced expression of the critical serotonin synthesis enzymes TPH-1, TPH-2, and DOPA decarboxylase (DDC), as well as the serotonin transporter, SERT. Concordantly, HSV-infected cells upregulated serotonin synthesis and its intracellular uptake. Increased serotonin synthesis and uptake was shown to influence HSV replication. Exogenous addition of serotonin increased HSV-1 yield, while both TPH-1/2 and SERT pharmacological inhibition reduced viral yield. Congruent with these in vitro findings, rabbits intraocularly infected with HSV-1 exhibited significantly higher aqueous humor serotonin concentrations that positively and strongly correlated with viral load and ocular disease severity. Collectively, our findings indicate that HSV-1 promotes serotonin synthesis and cellular uptake to facilitate viral replication and consequently, serotonin's proinflammatory effects may enhance the development of ocular disease., Competing Interests: The authors declare that TF and CN received funding for this and related studies from Eleusis Corporation. The funder was not involved in the study design, collection, analysis, interpretation of data, the writing of this article, or the decision to submit it for publication. TF and CN have pending intellectual property through the LSU Health Sciences Center that is currently licensed by Eleusis Corporation and serve as members of one of Eleusis’ scientific advisory boards. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Battaglia, Sanchez-Pino, Nichols and Foster.)

Introduction: Practice patterns and utilization of physical therapists (PTs) affiliated with Hemophilia Treatment Centers (HTCs) in the United States (US) are not well known., Aims: Describe utilization, role responsibilities and practice patterns of US HTC PTs. Identify practice patterns specifically focusing on assessment and treatment of pain. Recognize gaps in utilization and role responsibilities of PTs as part of the multidisciplinary team and suggest guidelines for PT involvement within the HTC., Methods: Respondents were a subset of a convenience sample of healthcare providers who responded to a non-validated survey developed by a multi-disciplinary panel of haemophilia experts., Results: A 33.0% response-rate (n = 59) representing all regions of US HTCs was achieved. Those working ≥10 hours per week were more likely to provide nutrition education (P = .026) and surgical options education (P < .001). Those who billed insurance for their services during comprehensive visits were more likely to provide education regarding surgical options (P = .046). The majority of PTs (95.0%) evaluated pain regardless of time spent in clinic and felt comfortable treating pain. Fifty-eight percent used a formal pain measurement tool and more likely to use a formal pain measurement tool if billing insurance (P = .004). Top five non-pharmacologic treatments recommended for pain management included splints/braces (84.8%), aquatic therapy (74.6%), orthotics (71.2%), surgical options (47.5%) and yoga (32.2%)., Conclusions: This study demonstrated PT utilization across HTC centres varies widely. Gaps in care may be addressed through salary support, funded education, greater regional/national collaboration of PTs specializing in bleeding disorders and advocacy for insurance coverage for appropriate services., (© 2022 John Wiley & Sons Ltd.)

Objectives: Pain is a known complication in persons with hemophilia (PWH) as a result of muscle and joint bleeding. Little is known regarding national Hemophilia Treatment Center (HTC) practice patterns related to pain management. The aim of this study was to: 1) Describe pain management practice patterns of HTC providers, 2) Identify gaps and areas of alignment with the CDC pain guidelines, and 3) Address educational opportunities for pain management. This survey is the first extensive description of multidisciplinary practice patterns of pain management for PWH., Methods: This descriptive study involved physicians, nurse practitioners, nurses, physical therapists, and social workers from federally funded Hemophilia Treatment Centers (HTC) eligible to complete an online survey exploring pain management practice patterns within the CDC pain guidelines., Results: Results of this survey shed light on areas of strength and cohesiveness between HTC providers, including the following: dedication to effective pain management, utilization of non-pharmacological pain options, trial of non-opioid medications first before opioids, maintaining follow-up with patients after opioid prescription initiation, recognizing and utilizing clinically important findings before prescribing opioids, and counseling their patients regarding potential risk factors., Conclusions: There remain opportunities to incorporate into clinical practice consistent use of tools such as formal screening questionnaires, opioid use agreements, written measurable goals, ongoing prescription monitoring, and written plans for discontinuation of opioid therapy. These results provide opportunities for improvement in education of HTC team members thus optimizing pain management in persons with bleeding disorders., (© The Author(s) 2021. Published by Oxford University Press on behalf of the American Academy of Pain Medicine.)

The serotonin (5-hydroxytryptamine, 5-HT) 2A receptor is most well known as the common target for classic psychedelic compounds. Interestingly, the 5-HT 2A receptor is the most widely expressed mammalian serotonin receptor and is found in nearly every examined tissue type including neural, endocrine, endothelial, immune, and muscle, suggesting it could be a novel and pharmacological target for several types of disorders. Despite this, the bulk of research on the 5-HT 2A receptor is focused on its role in the central nervous system (CNS). Recently, activation of 5-HT 2A receptors has emerged as a new anti-inflammatory strategy. This review will describe recent findings regarding psychedelics as anti-inflammatory compounds, as well as parse out differences in functional selectivity and immune regulation that exist between a number of well-known hallucinogenic compounds., (© 2022. The Author(s), under exclusive license to Springer Nature Switzerland AG.)

Serotonin 5-HT 2 receptors are expressed in many tissues and play important roles in biological processes. Although the 5-HT 2A receptor is primarily known for its role in central nervous system, it is also expressed in peripheral tissues. We have found that 5-HT 2A receptor antagonists inhibit human subcutaneous primary adipocyte differentiation. We also show that siRNA knockdown of the 5-HT 2A receptor blocks differentiation. Using gene expression analysis in combination with receptor antagonists we found that activity of 5-HT 2A receptors is necessary very early in the differentiation process to mediate expression of adipogenic genes, including peroxisome proliferator-activated receptor gamma (ppar-γ), adipocyte protein 2 (aP2), adiponectin, and serine/threonine-protein kinase 1 (sgk1). We show here for the first time that 5-HT 2A receptor activity is necessary for differentiation of human primary subcutaneous preadipocytes to adipocytes, and that 5-HT 2A receptor activity mediates key genes related to adipogenesis during this process. Importantly, this work contributes to a greater understanding of the adipocyte differentiation process, as well as to the role of 5-HT 2A receptors in peripheral tissues, and may be relevant to the development of novel therapeutic strategies targeting this receptor for the treatment of obesity related diseases., (© 2021. The Author(s).)

Psychedelic drugs can exert potent anti-inflammatory effects. However, anti-inflammatory effects do not appear to correlate with behavioral activity, suggesting different underlying mechanisms. We hypothesized that the distinct structural features of psychedelics underlie functionally selective mechanisms at the target 5-HT 2A receptor to elicit maximal anti-inflammatory effects. In order to test this hypothesis, we developed a new rat-based screening platform for allergic asthma. Next, we investigated 21 agonists at the 5-HT 2A receptor from the three primary chemotypes (phenylalkylamine, ergoline, and tryptamine) for their ability to prevent airways hyperresponsiveness as a measure of pulmonary inflammation. Furthermore, we assessed each drug for in vitro activation of the canonical signaling pathway, calcium mobilization, from the 5-HT 2A receptor. We find that the drug 2,5-dimethoxyphenethylamine (2C-H) represents the pharmacophore for anti-inflammatory activity and identify structural modifications that are either permissive or detrimental to anti-inflammatory activity. Additionally, there is no correlation between the ability of a particular psychedelic to activate intracellular calcium mobilization and to prevent the symptoms of asthma or with behavioral potencies. Our results support the notions that specific structural features mediate functional selectivity underlying anti-inflammatory activity and that relevant receptor activated pathways necessary for anti-inflammatory activity are different from canonical signaling pathways. Our results inform on the nature of interactions between ligands at the 5-HT 2A receptor as they relate to anti-inflammatory activity and are crucial for the development of new 5-HT 2A receptor agonists for anti-inflammatory therapeutics in the clinic that may be devoid of behavioral activity., Competing Interests: The authors declare the following competing financial interest(s): C.D.N. is supported by a sponsored research contract from Eleusis Therapeutics and is a member of its Scientific Advisory Board.

Psilocybin shows efficacy to alleviate depression in human clinical trials for six or more months after only one or two treatments. Another hallucinogenic drug, esketamine, has recently been U.S. Food and Drug Administration (FDA)-approved as a rapid-acting antidepressant. The mechanistic basis for the antidepressant effects of psilocybin and ketamine appear to be conserved. The efficacy of these two medications has not, however, been directly compared either clinically or preclinically. Further, whether or not a profound subjective existential experience is necessary for psilocybin to have antidepressant effects is unknown. To address these questions, we tested psilocybin, lysergic acid diethylamide (LSD), and ketamine in a rat model for depression. As in humans, a single administration of psilocybin or LSD produced persistent antidepressant-like effects in our model. In contrast, ketamine produced only a transient antidepressant-like effect. Our results indicate that classic psychedelics may have therapeutic efficacy that is more persistent than that of ketamine, and also suggest that a subjective existential experience may not be necessary for therapeutic effects.

Research has shown that psychedelics, such as lysergic acid diethylamide (LSD), have profound anti-inflammatory properties mediated by 5-HT 2A receptor signaling, supporting their evaluation as a therapeutic for neuroinflammation associated with neurodegenerative disease., Objective: This study evaluated the safety, tolerability, pharmacokinetics, and pharmacodynamics of orally repeated administration of 5 μg, 10 μg, and 20 μg LSD in older healthy individuals. In the current paper, we present safety, tolerability, pharmacokinetics, and pharmacodynamic measures that relate to safety, tolerability, and dose response., Methods: This was a phase 1 double-blind, placebo-controlled, randomized study. Volunteers were randomly assigned to 1 of 4 dose groups (5 μg, 10 μg, 20 μg LSD, and placebo), and received their assigned dose on six occasions (i.e., every 4 days)., Results: Forty-eight older healthy volunteers (mean age = 62.9 years) received placebo (n = 12), 5 μg (n = 12), 10 μg (n = 12), or 20 μg (n = 12) LSD. LSD plasma levels were undetectable for the 5 μg group and peak blood plasma levels for the 10 μg and 20 μg groups occurred at 30 min. LSD was well tolerated, and the frequency of adverse events was no higher than for placebo. Assessments of cognition, balance, and proprioception revealed no impairment., Conclusions: Our results suggest safety and tolerability of orally administered 5 μg, 10 μg, and 20 μg LSD every fourth day over a 21-day period and support further clinical development of LSD for the treatment and prevention of Alzheimer's disease (AD).

Introduction: The majority of contemporary psychedelic research has focused on ayahuasca, lysergic acid diethylamide, and psilocybin, though there are hundreds of novel psychedelic compounds that may have clinical utility. The purpose of the present study was to evaluate the therapeutic potential of classic and novel phenethylamine, tryptamine, and lysergamide psychedelics via a large, nationally representative population-based survey., Methods: We tested the unique associations of lifetime classic and novel phenethylamine, tryptamine, and lysergamide psychedelics with past month psychological distress and past year suicidality among respondents pooled from years 2008-2017 of the National Survey on Drug Use and Health (weighted N = 260,964,827)., Results: Lifetime classic tryptamine use was associated with a decreased odds of past month psychological distress [aOR = 0.76; (0.69-0.83)] and past year suicidal thinking [aOR = 0.79; (0.72-0.87)]. Lifetime novel phenethylamine use, on the other hand, was associated with an increased odds of past year suicidal thinking [aOR = 1.44; (1.06-1.95)] and past year suicidal planning [aOR = 1.60; (1.06-2.41)]. No other significant associations were found., Discussion and Conclusions: These findings, which may be driven by differences in pharmacodynamics, suggest that classic tryptamines may hold the greatest therapeutic potential of the psychedelics, whereas novel phenethylamines may pose risk for harm. The present findings thus support continued research on the clinical application of classic tryptamines. Though the current results caution against the clinical utility of novel phenethylamines, further study of these and other novel psychedelic substances is nonetheless warranted to better understand their potential application., (Copyright © 2020 Sexton, Nichols and Hendricks.)

Aims: Although the bulk of research into the biology of serotonin 5-HT 2A receptors has focused on its role in the CNS, selective activation of these receptors in peripheral tissues can produce profound anti-inflammatory effects. We previously demonstrated that the small molecule 5-HT 2 receptor agonist (R)-2,5-dimethoxy-4-iodoamphetamine [(R)-DOI] inhibits TNF-α-mediated proinflammatory signaling cascades and inflammation via 5-HT 2A receptor activation and prevents the development of, and inflammation associated with, acute allergic asthma in a mouse ovalbumin (OVA) model. Here, we investigated the ability of (R)-DOI to reverse inflammation and symptoms associated with established asthma in a newly developed model of chronic asthma., Methods: An 18-week ovalbumin challenge period was performed to generate persistent, chronic asthma in BALB/c mice. Four once daily intranasal treatments of (R)-DOI were administered one week after allergen cessation, with respiratory parameters being measured by whole-body plethysmography (WBP). Cytokine and chemokine levels were measured by quantitative real-time polymerase chain reaction (qRT-PCR) in homogenized lung tissue, bronchoalveolar (BALF) fluid was analyzed for chemokine modulation by multiplex assays, and Periodic Acid-Schiff and Masson's Trichrome staining was performed to determine goblet cell infiltration and overall changes to lung morphology., Key Findings: 5-HT 2 activation via (R)-DOI attenuates elevated airway hyperresponsiveness to methacholine, reduces pulmonary inflammation and mucus production, and reduces airway structural remodeling and collagen deposition by nearly 70%., Significance: Overall, these data provide support for the therapeutic potential of (R)-DOI and 5-HT 2 receptor activation for the treatment of asthma, and identifies (R)-DOI as a novel therapeutic compound against pulmonary fibrosis., (Copyright © 2019. Published by Elsevier Inc.)

Coronary artery disease (CAD) is a progressive cardiovascular syndrome characterized by cholesterol-induced focal arterial lesions that impair oxygen delivery to the heart. As both innate and adaptive immune cells play critical roles in the formation and progression of arterial plaques and endothelial cell dysfunction, CAD is commonly viewed as a chronic inflammatory disorder. Our lab has previously discovered that 5-HT 2A receptor activation with the 5-HT 2 receptor selective agonist (R)-2,5-dimethoxy-4-iodoamphetamine [(R)-DOI] has potent anti-inflammatory activity in both cell culture and whole animal models. Here we have examined the putative therapeutic effects of (R)-DOI in the ApoE -/- high fat model of cardiovascular disease. Subcutaneously implanted osmotic minipumps were used to infuse sustained low rates (0.15 μg / hr) of (R)-DOI∙HCl to mice fed a high-fat "Western" diet. (R)-DOI treated mice had significant reductions in expression levels of mRNA for inflammatory markers like Il6 in vascular tissue, normalized glucose homeostasis, and reduced circulating cholesterol levels. As cardiovascular disease is a leading cause of death both globally and in the Western world, activation of 5-HT 2A receptors at sub-behavioral levels may represent a new strategy to treat inflammation-based cardiovascular disease.

Background: In the past few years, the issue of 'microdosing' psychedelics has been openly discussed in the public arena where claims have been made about their positive effect on mood state and cognitive processes such as concentration. However, there are very few scientific studies that have specifically addressed this issue, and there is no agreed scientific consensus on what microdosing is., Aim: This critique paper is designed to address questions that need to be answered by future scientific studies and to offer guidelines for these studies., Approach: Owing to its proximity for a possible approval in clinical use and short-lasting pharmacokinetics, our focus is predominantly on psilocybin. Psilocybin is allegedly, next to lysergic acid diethylamide (LSD), one of the two most frequently used psychedelics to microdose. Where relevant and available, data for other psychedelic drugs are also mentioned., Conclusion: It is concluded that while most anecdotal reports focus on the positive experiences with microdosing, future research should also focus on potential risks of (multiple) administrations of a psychedelic in low doses. To that end, (pre)clinical studies including biological (e.g. heart rate, receptor turnover and occupancy) as well as cognitive (e.g. memory, attention) parameters have to be conducted and will shed light on the potential negative consequences microdosing could have.

Serotonin (5-hydroxytryptamine, 5-HT) 2A receptor agonists have recently emerged as promising new treatment options for a variety of disorders. The recent success of these agonists, also known as psychedelics, like psilocybin for the treatment of anxiety, depression, obsessive-compulsive disorder (OCD), and addiction, has ushered in a renaissance in the way these compounds are perceived in the medical community and populace at large. One emerging therapeutic area that holds significant promise is their use as anti-inflammatory agents. Activation of 5-HT 2A receptors produces potent anti-inflammatory effects in animal models of human inflammatory disorders at sub-behavioural levels. This review discusses the role of the 5-HT 2A receptor in the inflammatory response, as well as highlight studies using the 5-HT 2A agonist ( R )-2,5-dimethoxy-4-iodoamphetamine [( R )-DOI] to treat inflammation in cellular and animal models. It also examines potential mechanisms by which 5-HT 2A agonists produce their therapeutic effects. Overall, psychedelics regulate inflammatory pathways via novel mechanisms, and may represent a new and exciting treatment strategy for several inflammatory disorders.

The classic serotonergic hallucinogens, or psychedelics, have the ability to profoundly alter perception and behavior. These can include visual distortions, hallucinations, detachment from reality, and mystical experiences. Some psychedelics, like LSD, are able to produce these effects with remarkably low doses of drug. Others, like psilocybin, have recently been demonstrated to have significant clinical efficacy in the treatment of depression, anxiety, and addiction that persist for at least several months after only a single therapeutic session. How does this occur? Much work has recently been published from imaging studies showing that psychedelics alter brain network connectivity. They facilitate a disintegration of the default mode network, producing a hyperconnectivity between brain regions that allow centers that do not normally communicate with each other to do so. The immediate and acute effects on both behaviors and network connectivity are likely mediated by effector pathways downstream of serotonin 5-HT2A receptor activation. These acute molecular processes also influence gene expression changes, which likely influence synaptic plasticity and facilitate more long-term changes in brain neurochemistry ultimately underlying the therapeutic efficacy of a single administration to achieve long-lasting effects. In this review, we summarize what is currently known about the molecular genetic responses to psychedelics within the brain and discuss how gene expression changes may contribute to altered cellular physiology and behaviors.

Introduction: Recurrent bleeding and associated pain are critical components in the management of bleeding disorders, yet scant data describe perceptions of pain in this patient population., Objective: This study assessed perceptions of pain and pain management in adolescents and young adults (AYAs) with haemophilia or von Willebrand disease (VWD) to determine agreement/disagreement between patients, caregivers and health care providers., Methods: Using an online questionnaire, AYA patients (N=89), their caregivers (N=77), and providers (N=54) reported on pain perception, pain treatment and pain control. Acute and chronic pain was measured in patients via the Faces Pain Scale-Revised (FPS-R). Questionnaires queried about pharmacologic and non-pharmacologic pain management methods and how well providers and caregivers helped to manage pain., Results: Poor agreement existed between patients and caregivers across all pain levels, perception of pain control and effectiveness of pain management. Specifically for chronic pain, poor agreement was noted between patients and caregivers (kappa=0.04; 29% agreement) and patients and providers (kappa=-0.07; 21.4% agreement). Among patients reporting acute or chronic pain, only 67% and 43%, respectively, utilized medication for their specific pain. Patients used more opioid medications than expected by their providers. On average, AYAs reported initial use of pain medications for chronic pain at 11.5 years., Conclusions: Ongoing research is needed in haemophilia and VWD pain management, and on the differences in pain perception between patients, caregivers and providers. As chronic pain often begins at an early age, optimal pain management should include acknowledging patient complaints, exploring pharmacologic and non-pharmacologic options, and optimizing prophylaxis., (© 2017 John Wiley & Sons Ltd.)

Psychedelic drugs, such as lysergic acid diethylamide (LSD), mescaline, and psilocybin, exert profound effects on brain and behavior. After decades of difficulties in studying these compounds, psychedelics are again being tested as potential treatments for intractable biomedical disorders. Preclinical research of psychedelics complements human neuroimaging studies and pilot clinical trials, suggesting these compounds as promising treatments for addiction, depression, anxiety, and other conditions. However, many questions regarding the mechanisms of action, safety, and efficacy of psychedelics remain. Here, we summarize recent preclinical and clinical data in this field, discuss their pharmacological mechanisms of action, and outline critical areas for future studies of psychedelic drugs, with the goal of maximizing the potential benefits of translational psychedelic biomedicine to patients., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Flow cytometry has the potential to facilitate understanding of the heterogeneous responses of diverse brain cell populations to a variety of stimuli. However, existing methods of applying flow cytometry to brain tissues are each limited in certain ways. They either require genetically labeled cells to achieve separation of specific populations, are not applicable to previously fixed tissue, or are not compatible with downstream mRNA analysis. Here, we describe a group of related methods that overcome many previous limitations and allow robust sorting and downstream molecular analysis of highly enriched populations of specific neuronal and non-neuronal cells from any mammalian brain. We illustrate these techniques, which are compatible with antibodies for both nuclear and non-nuclear epitopes and do not require transgenic animals, with three examples. First, we describe the separation and downstream mRNA analysis of four types of cortical interneurons (somatostatin, parvalbumin, calretinin, and calbindin) from paraformaldehyde-fixed rat brain sections. Second, we demonstrate separation of neurons and non-neurons from zinc-fixed mouse brain cortical sections followed by analysis of enzymatic activity (ACE2 activity) and mRNA expression. Third, we show that routinely fixed post-mortem human brain can be analyzed by isolating parvalbumin-containing neurons from cortical samples that were fixed for periods of up to 8 weeks in formalin. In each case, sorted cell identity was confirmed with mRNA analysis. The neurocytometry methodology described here has the potential to significantly expand studies to analyze the effects of drugs, environmental manipulations, and disease states on the nucleic acid and protein content of specific brain cell populations.

There has recently been a resurgence of interest in psychedelics, substances that profoundly alter perception and cognition and have recently demonstrated therapeutic efficacy to treat anxiety, depression, and addiction in the clinic. The receptor mechanisms that drive their molecular and behavioral effects involve activation of cortical serotonin 5-HT 2A receptors, but the responses of specific cellular populations remain unknown. Here, we provide evidence that a small subset of 5-HT 2A -expressing excitatory neurons is directly activated by psychedelics and subsequently recruits other select cell types including subpopulations of inhibitory somatostatin and parvalbumin GABAergic interneurons, as well as astrocytes, to produce distinct and regional responses. To gather data regarding the response of specific neuronal populations, we developed methodology for fluorescence-activated cell sorting (FACS) to segregate and enrich specific cellular subtypes in the brain. These methods allow for robust neuronal sorting based on cytoplasmic epitopes followed by downstream nucleic acid analysis, expanding the utility of FACS in neuroscience research., (Copyright © 2016 Forschungsgesellschaft für Arbeitsphysiologie und Arbeitschutz e.V. Published by Elsevier B.V. All rights reserved.)

Asthma is an inflammatory disease of the lung characterized by airways hyper-responsiveness (AHR), inflammation, and mucus hyperproduction. Current mainstream therapies include bronchodilators that relieve bronchoconstriction and inhaled glucocorticoids to reduce inflammation. The small molecule hormone and neurotransmitter serotonin has long been known to be involved in inflammatory processes; however, its precise role in asthma is unknown. We have previously established that activation of serotonin 5-hydroxytryptamine (5-HT)(2A) receptors has potent anti-inflammatory activity in primary cultures of vascular tissues and in the whole animal in vasculature and gut tissues. The 5-HT(2A) receptor agonist, (R)-2,5-dimethoxy-4-iodoamphetamine [(R)-DOI] is especially potent. In this work, we have examined the effect of (R)-DOI in an established mouse model of allergic asthma. In the ovalbumin mouse model of allergic inflammation, we demonstrate that inhalation of (R)-DOI prevents the development of many key features of allergic asthma, including AHR, mucus hyperproduction, airways inflammation, and pulmonary eosinophil recruitment. Our results highlight a likely role of the 5-HT2 receptors in allergic airways disease and suggest that 5-HT2 receptor agonists may represent an effective and novel small molecule-based therapy for asthma.

Chronic administration of lysergic acid diethylamide (LSD) every other day to rats results in a variety of abnormal behaviors. These build over the 90 day course of treatment and can persist at full strength for at least several months after cessation of treatment. The behaviors are consistent with those observed in animal models of schizophrenia and include hyperactivity, reduced sucrose-preference, and decreased social interaction. In order to elucidate molecular changes that underlie these aberrant behaviors, we chronically treated rats with LSD and performed RNA-sequencing on the medial prefrontal cortex (mPFC), an area highly associated with both the actions of LSD and the pathophysiology of schizophrenia and other psychiatric illnesses. We observed widespread changes in the neurogenetic state of treated animals four weeks after cessation of LSD treatment. QPCR was used to validate a subset of gene expression changes observed with RNA-Seq, and confirmed a significant correlation between the two methods. Functional clustering analysis indicates differentially expressed genes are enriched in pathways involving neurotransmission (Drd2, Gabrb1), synaptic plasticity (Nr2a, Krox20), energy metabolism (Atp5d, Ndufa1) and neuropeptide signaling (Npy, Bdnf), among others. Many processes identified as altered by chronic LSD are also implicated in the pathogenesis of schizophrenia, and genes affected by LSD are enriched with putative schizophrenia genes. Our results provide a relatively comprehensive analysis of mPFC transcriptional regulation in response to chronic LSD, and indicate that the long-term effects of LSD may bear relevance to psychiatric illnesses, including schizophrenia., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Background: Echocardiography is available in the most basic healthcare environments. Mitral repair is potentially curative and, when possible, recommended over replacement. The efficacy of echo-guided repair has not been established., Methods and Results: We developed a succinct set of precisely defined images observed to be highly concordant with intraoperative findings. These images guided intervention on 237 consecutive patients. None were lost to follow-up, and serial echocardiography was obtained on all repairs. This analysis includes 2037 echocardiograms. The intent to repair or replace was documented preoperatively in 98.7%. Concordance was associated with successful repair (97.8% versus 57.1%; P=0.001). Three-dimensional concordance was higher than 2-dimensional (100% versus 94.4%; P=0.05). Echocardiography guided a graduated surgical approach for degenerative and myopathic repairs by quantifying segmental prolapse, anterior leaflet closing angles, and tenting for integration of secondary chord lysis (P<0.001) and commissural width (P<0.01). Repair rates increased from 46.5% to 77.6% (P<0.001). Concomitant Society of Thoracic Surgeons rates were 46.6% (versus unguided 46.5%; P=0.99) and 54.9% (versus echo guided 77.6%; P<0.001). Repair was successful in 91.5% of isolated echo-guided mitral operations (versus concomitant Society of Thoracic Surgeons 70.0%; P<0.001). Echo-guided repair rates for degenerative, myopathic, and inflammatory diseases were 99.0%, 97.1%, and 84.2% with linearized annual recurrent regurgitation of 0.63%, 2.19%, and 4.37%, respectively., Conclusions: Echocardiography can reliably identify repairable mitral disease and guide intervention. Echo-guided repair is associated with a higher rate of initial success than unguided historical and concomitant national controls. Three-dimensional echo improves concordance. Secondary chord lysis is associated with durable repair and may prevent ventricular remodeling.

The fruit fly, Drosophila melanogaster, is a good experimental organism to study the underlying mechanism of heart rate (HR) regulation. It is already known that many neuromodulators (serotonin, dopamine, octopamine, acetylcholine) change the HR in Drosophila melanogaster larvae. In this study, we investigated the role of cAMP-PKA signaling pathway in HR regulation and 5-HT positive chronotropic action. In order to obtain insight into the 5-HT mechanism of action in larvae cardiomyocytes, genetic and pharmacological approaches were used. We used transgenic flies that expressed the hM4Di receptor [designer receptors exclusively activated by designer drugs (DREADDs)] as one tool. Our previous results showed that activation of hM4Di receptors (modified muscarinic acetylcholine receptors) decreases or arrests the heart from beating. In this study, it was hypothesized that the positive chronotropic effect of serotonin [5-hydroxytryptamine (5-HT)] are mediated by serotonin receptors coupled to the adenylyl cyclase pathway and downstream cAMP and PKA activity. Activation of hM4Di by clozapine-N-oxide (CNO) was predicted to block the effects of serotonin by inhibiting adenylyl cyclase activity through Gαi pathway activation. Interestingly, we found here that manipulation of adenylyl cyclase activity and cAMP levels had no significant effect on HR. The ability of hM4Di receptor activation to slow or stop the heart is therefore likely mediated by activation of GIRK channels to produce hyperpolarization of cardiomyocytes, and not through inhibition of adenylyl cyclase.

Tumor necrosis factor alpha (TNF-α) plays a key role in inflammation, and its production and signaling contribute to many inflammatory related diseases. Recently, we discovered that selective activation of serotonin 5-HT2A receptors with the agonist (R)-DOI produces a super-potent blockade of proinflammatory markers in primary rat aortic smooth muscle cells. Here, we demonstrate that systemic administration of (R)-DOI can block the systemic effects of TNF-α in whole animal, with potent anti-inflammatory effects in the aortic arch and small intestine. This includes blockade of TNF-α-induced expression of pro-inflammatory cell adhesion (Icam-1, Vcam-1), cytokine (Il-6, IL-1b), and chemokine (Mcp-1, Cx3cl1) genes, and expression of VCAM-1 protein in the intestine. Further, systemic (R)-DOI also prevents the TNF-α-induced increase of circulating IL-6. Importantly, utilizing receptor selective antagonists, we have demonstrated that the mechanism underlying the systemic anti-inflammatory effects of (R)-DOI is activation of serotonin 5-HT2A receptors. Our results highlight a powerful new role for the serotonin 5-HT2A receptor in inflammatory processes, and indicate that agonism of serotonin receptors may represent an effective and novel approach to develop powerful small molecule therapeutics for inflammatory diseases and conditions such as atherosclerosis and inflammatory bowel disease.

We have translated a powerful genetic tool, designer receptors exclusively activated by designer drugs (DREADDs), from mammalian systems to Drosophila melanogaster to selectively, rapidly, reversibly, and dose-dependently control behaviors and physiological processes in the fly. DREADDs are muscarinic acetylcholine G protein-coupled receptors evolved for loss of affinity to acetylcholine and for the ability to be fully activated by an otherwise biologically inert chemical, clozapine-N-oxide. We demonstrate its ability to control a variety of behaviors and processes in larvae and adults, including heart rate, sensory processing, diurnal behavior, learning and memory, and courtship. The advantages of this particular technology include the dose-responsive control of behaviors, the lack of a need for specialized equipment, and the capacity to remotely control signaling in essentially all neuronal and nonneuronal fly tissues., (Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.)

Amyotrophic lateral sclerosis (ALS) is an uncommon neurodegenerative disease caused by degeneration of upper and lower motor neurons. Several genes, including SOD1, TDP-43, FUS, Ubiquilin 2, C9orf72 and Profilin 1, have been linked with the sporadic and familiar forms of ALS. FUS is a DNA/RNA-binding protein (RBP) that forms cytoplasmic inclusions in ALS and frontotemporal lobular degeneration (FTLD) patients' brains and spinal cords. However, it is unknown whether the RNA-binding ability of FUS is required for causing ALS pathogenesis. Here, we exploited a Drosophila model of ALS and neuronal cell lines to elucidate the role of the RNA-binding ability of FUS in regulating FUS-mediated toxicity, cytoplasmic mislocalization and incorporation into stress granules (SGs). To determine the role of the RNA-binding ability of FUS in ALS, we mutated FUS RNA-binding sites (F305L, F341L, F359L, F368L) and generated RNA-binding-incompetent FUS mutants with and without ALS-causing mutations (R518K or R521C). We found that mutating the aforementioned four phenylalanine (F) amino acids to leucines (L) (4F-L) eliminates FUS RNA binding. We observed that these RNA-binding mutations block neurodegenerative phenotypes seen in the fly brains, eyes and motor neurons compared with the expression of RNA-binding-competent FUS carrying ALS-causing mutations. Interestingly, RNA-binding-deficient FUS strongly localized to the nucleus of Drosophila motor neurons and mammalian neuronal cells, whereas FUS carrying ALS-linked mutations was distributed to the nucleus and cytoplasm. Importantly, we determined that incorporation of mutant FUS into the SG compartment is dependent on the RNA-binding ability of FUS. In summary, we demonstrate that the RNA-binding ability of FUS is essential for the neurodegenerative phenotype in vivo of mutant FUS (either through direct contact with RNA or through interactions with other RBPs).

Objective: This evidence-based review provides a systematic and comprehensive review of the literature regarding the utility of polysomnography for the evaluation of non-respiratory sleep disorders in children including hypersomnias, parasomnias, sleep-related movement disorders, and sleep in other special populations., Methods: A task force of pediatric sleep medicine experts performed a systematic review of the literature regarding the use of polysomnography for non-respiratory sleep disorders in children. They identified and graded 76 papers as evidence., Results: The main results include (1) polysomnography combined with the multiple sleep latency test is useful for evaluating disorders of excessive somnolence to objectively quantify sleepiness. The results have to be interpreted with consideration of the pubertal stage and regularity of the sleep patterns of the child; (2) polysomnography is indicated in children with parasomnias or sleep related movement disorders who have a high likelihood of having obstructive sleep apnea (OSA); (3) polysomnography is not routinely indicated in children with enuresis unless there is a high likelihood of OSA; (4) polysomnography can be helpful in evaluating children with restless legs syndrome (RLS) and when periodic limb movement disorder (PLMD) is suspected., Conclusions: These findings suggest that, in children with non-respiratory sleep disorders, polysomnography should be a part of a comprehensive sleep evaluation in selected circumstances to determine the nature of the events in more detail or when the suspicion of OSA is relatively high.

Recent studies have demonstrated that cargo exit from the endoplasmic reticulum (ER) may be directed by ER export motifs recognized by components of the coat protein II (COPII) vesicles. However, little is known about ER export motifs and vesicle targeting of the G protein-coupled receptor (GPCR) superfamily. Here, we have demonstrated that a triple Arg (3R) motif in the third intracellular loop functions as a novel ER export signal for α(2B)-adrenergic receptor (α(2B)-AR). The 3R motif mediates α(2B)-AR interaction with Sec24C/D and modulates ER exit, cell surface transport and function of α(2B)-AR. Furthermore, export function of the 3R motif is independent of its position within α(2B)-AR and can be conferred to CD8 glycoprotein. These data provide the first evidence implicating that export of GPCRs is controlled by code-directed interactions with selective components of the COPII transport machinery., (© 2012 John Wiley & Sons A/S.)

Drosophila melanogaster, the fruit fly, has been used to study molecular mechanisms of a wide range of human diseases such as cancer, cardiovascular disease and various neurological diseases(1). We have optimized simple and robust behavioral assays for determining larval locomotion, adult climbing ability (RING assay), and courtship behaviors of Drosophila. These behavioral assays are widely applicable for studying the role of genetic and environmental factors on fly behavior. Larval crawling ability can be reliably used for determining early stage changes in the crawling abilities of Drosophila larvae and also for examining effect of drugs or human disease genes (in transgenic flies) on their locomotion. The larval crawling assay becomes more applicable if expression or abolition of a gene causes lethality in pupal or adult stages, as these flies do not survive to adulthood where they otherwise could be assessed. This basic assay can also be used in conjunction with bright light or stress to examine additional behavioral responses in Drosophila larvae. Courtship behavior has been widely used to investigate genetic basis of sexual behavior, and can also be used to examine activity and coordination, as well as learning and memory. Drosophila courtship behavior involves the exchange of various sensory stimuli including visual, auditory, and chemosensory signals between males and females that lead to a complex series of well characterized motor behaviors culminating in successful copulation. Traditional adult climbing assays (negative geotaxis) are tedious, labor intensive, and time consuming, with significant variation between different trials(2-4). The rapid iterative negative geotaxis (RING) assay(5) has many advantages over more widely employed protocols, providing a reproducible, sensitive, and high throughput approach to quantify adult locomotor and negative geotaxis behaviors. In the RING assay, several genotypes or drug treatments can be tested simultaneously using large number of animals, with the high-throughput approach making it more amenable for screening experiments.

Insulin signaling regulates lifespan, reproduction, metabolic homeostasis, and resistance to stress in the adult organism. In Drosophila, there are seven insulin-like peptides (DILP1-7). Three of these (DILP2, 3 and 5) are produced in median neurosecretory cells of the brain, designated IPCs. Previous work has suggested that production or release of DILPs in IPCs can be regulated by a factor secreted from the fat body as well as by neuronal GABA or short neuropeptide F. There is also evidence that serotonergic neurons may regulate IPCs. Here, we investigated mechanisms by which serotonin may regulate the IPCs. We show that the IPCs in adult flies express the 5-HT(1A), but not the 5-HT(1B) or 5-HT(7) receptors, and that processes of serotonergic neurons impinge on the IPC branches. Knockdown of 5-HT(1A) in IPCs by targeted RNA interference (RNAi) leads to increased sensitivity to heat, prolonged recovery after cold knockdown and decreased resistance to starvation. Lipid metabolism is also affected, but no effect on growth was seen. Furthermore, we show that DILP2-immunolevels in IPCs increase after 5-HT(1A) knockdown; this is accentuated by starvation. Heterozygous 5-HT(1A) mutant flies display the same phenotype in all assays, as seen after targeted 5-HT(1A) RNAi, and flies fed the 5-HT(1A) antagonist WAY100635 display reduced lifespan at starvation. Our findings suggest that serotonin acts on brain IPCs via the 5-HT(1A) receptor, thereby affecting their activity and probably insulin signaling. Thus, we have identified a second inhibitory pathway regulating IPC activity in the Drosophila brain.

Learning and memory in the fruit fly, Drosophila melanogaster, is a complex behavior with many parallels to mammalian learning and memory. Although many neurotransmitters including acetylcholine, dopamine, glutamate, and GABA have previously been demonstrated to be involved in aversive olfactory learning and memory, the role of serotonin has not been well defined. Here, we present the first evidence of the involvement of individual serotonin receptors in olfactory learning and memory in the fly. We initially followed a pharmacological approach, utilizing serotonin receptor agonists and antagonists to demonstrate that all serotonin receptor families present in the fly are necessary for short-term learning and memory. Isobolographic analysis utilizing combinations of drugs revealed functional interactions are occurring between 5-HT(1A)-like and 5-HT(2), and 5-HT(2) and 5-HT(7) receptor circuits in mediating short-term learning and memory. Examination of long-term memory suggests that 5-HT(1A)-like receptors are necessary for consolidation and important for recall, 5-HT(2) receptors are important for consolidation and recall, and 5-HT(7) receptors are involved in all three phases. Importantly, we have validated our pharmacological results with genetic experiments and showed that hypomorph strains for 5-HT(2)Dro and 5-HT(1B)Dro receptors, as well as knockdown of 5-HT(7)Dro mRNA, significantly impair performance in short-term memory. Our data highlight the importance of the serotonin system and individual serotonin receptors to influence olfactory learning and memory in the fly, and position the fly as a model system to study the role of serotonin in cognitive processes relevant to mammalian CNS function., (Copyright © 2011 IBRO. Published by Elsevier Ltd. All rights reserved.)

Many people who take LSD experience a second temporal phase of LSD intoxication that is qualitatively different, and was described by Daniel Freedman as "clearly a paranoid state." We have previously shown that the discriminative stimulus effects of LSD in rats also occur in two temporal phases, with initial effects mediated by activation of 5-HT(2A) receptors (LSD30), and the later temporal phase mediated by dopamine D2-like receptors (LSD90). Surprisingly, we have now found that non-competitive NMDA antagonists produced full substitution in LSD90 rats, but only in older animals, whereas in LSD30, or in younger animals, these drugs did not mimic LSD. Chronic administration of low doses of LSD (>3 months, 0.16 mg/kg every other day) induces a behavioral state characterized by hyperactivity and hyperirritability, increased locomotor activity, anhedonia, and impairment in social interaction that persists at the same magnitude for at least three months after cessation of LSD treatment. These behaviors, which closely resemble those associated with psychosis in humans, are not induced by withdrawal from LSD; rather, they are the result of neuroadaptive changes occurring in the brain during the chronic administration of LSD. These persistent behaviors are transiently reversed by haloperidol and olanzapine, but are insensitive to MDL-100907. Gene expression analysis data show that chronic LSD treatment produced significant changes in multiple neurotransmitter system-related genes, including those for serotonin and dopamine. Thus, we propose that chronic treatment of rats with low doses of LSD can serve as a new animal model of psychosis that may mimic the development and progression of schizophrenia, as well as model the established disease better than current acute drug administration models utilizing amphetamine or NMDA antagonists such as PCP., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

The common fruit fly, Drosophila melanogaster, is a well studied and highly tractable genetic model organism for understanding molecular mechanisms of human diseases. Many basic biological, physiological, and neurological properties are conserved between mammals and D. melanogaster, and nearly 75% of human disease-causing genes are believed to have a functional homolog in the fly. In the discovery process for therapeutics, traditional approaches employ high-throughput screening for small molecules that is based primarily on in vitro cell culture, enzymatic assays, or receptor binding assays. The majority of positive hits identified through these types of in vitro screens, unfortunately, are found to be ineffective and/or toxic in subsequent validation experiments in whole-animal models. New tools and platforms are needed in the discovery arena to overcome these limitations. The incorporation of D. melanogaster into the therapeutic discovery process holds tremendous promise for an enhanced rate of discovery of higher quality leads. D. melanogaster models of human diseases provide several unique features such as powerful genetics, highly conserved disease pathways, and very low comparative costs. The fly can effectively be used for low- to high-throughput drug screens as well as in target discovery. Here, we review the basic biology of the fly and discuss models of human diseases and opportunities for therapeutic discovery for central nervous system disorders, inflammatory disorders, cardiovascular disease, cancer, and diabetes. We also provide information and resources for those interested in pursuing fly models of human disease, as well as those interested in using D. melanogaster in the drug discovery process.

Objective: This comprehensive, evidence-based review provides a systematic analysis of the literature regarding the validity, reliability, and clinical utility of polysomnography for characterizing breathing during sleep in children. Findings serve as the foundation of practice parameters regarding respiratory indications for polysomnography in children., Methods: A task force of content experts performed a systematic review of the relevant literature and graded the evidence using a standardized grading system. Two hundred forty-three evidentiary papers were reviewed, summarized, and graded. The analysis addressed the operating characteristics of polysomnography as a diagnostic procedure in children and identified strengths and limitations of polysomnography for evaluation of respiratory function during sleep., Results: The analysis documents strong face validity and content validity, moderately strong convergent validity when comparing respiratory findings with a variety of relevant independent measures, moderate-to-strong test-retest validity, and limited data supporting discriminant validity for characterizing breathing during sleep in children. The analysis documents moderate-to-strong test-retest reliability and interscorer reliability based on limited data. The data indicate particularly strong clinical utility in children with suspected sleep related breathing disorders and obesity, evolving metabolic syndrome, neurological, neurodevelopmental, or genetic disorders, and children with craniofacial syndromes. Specific consideration was given to clinical utility of polysomnography prior to adenotonsillectomy (AT) for confirmation of obstructive sleep apnea syndrome. The most relevant findings include: (1) recognition that clinical history and examination are often poor predictors of respiratory polygraphic findings, (2) preoperative polysomnography is helpful in predicting risk for perioperative complications, and (3) preoperative polysomnography is often helpful in predicting persistence of obstructive sleep apnea syndrome in patients after AT. No prospective studies were identified that address whether clinical outcome following AT for treatment of obstructive sleep apnea is improved in association with routine performance of polysomnography before surgery in otherwise healthy children. A small group of papers confirm the clinical utility of polysomnography for initiation and titration of positive airway pressure support., Conclusions: Pediatric polysomnography shows validity, reliability, and clinical utility that is commensurate with most other routinely employed diagnostic clinical tools or procedures. Findings indicate that the "gold standard" for diagnosis of sleep related breathing disorders in children is not polysomnography alone, but rather the skillful integration of clinical and polygraphic findings by a knowledgeable sleep specialist. Future developments will provide more sophisticated methods for data collection and analysis, but integration of polysomnographic findings with the clinical evaluation will represent the fundamental diagnostic challenge for the sleep specialist.

The 5-HT(7) receptor remains one of the less well characterized serotonin receptors. Although it has been demonstrated to be involved in the regulation of mood, sleep, and circadian rhythms, as well as relaxation of vascular smooth muscles in mammals, the precise mechanisms underlying these functions remain largely unknown. The fruit fly, Drosophila melanogaster, is an attractive model organism to study neuropharmacological, molecular, and behavioral processes that are largely conserved with mammals. Drosophila express a homolog of the mammalian 5-HT(7) receptor, as well as homologs for the mammalian 5-HT(1A), and 5-HT(2), receptors. Each fly receptor couples to the same effector pathway as their mammalian counterpart and have been demonstrated to mediate similar behavioral responses. Here, we report on the expression and function of the 5-HT(7)Dro receptor in Drosophila. In the larval central nervous system, expression is detected postsynaptically in discreet cells and neuronal circuits. In the adult brain there is strong expression in all large-field R neurons that innervate the ellipsoid body, as well as in a small group of cells that cluster with the PDF-positive LNvs neurons that mediate circadian activity. Following both pharmacological and genetic approaches, we have found that 5-HT(7)Dro activity is essential for normal courtship and mating behaviors in the fly, where it appears to mediate levels of interest in both males and females. This is the first reported evidence of direct involvement of a particular serotonin receptor subtype in courtship and mating in the fly.

Salmonella attachment to the intestinal epithelium triggers delivery of bacterial effector proteins into the host cytosol through a type III secretion system (T3SS), leading to pronounced membrane ruffling and macropinocytic uptake of attached bacteria. The tip of the T3SS is made up of two proteins, SipB and SipC, which insert into the host plasma membrane, forming a translocation pore. Both the N and C termini of SipC are exposed in the host cytosol and have been shown to directly modulate actin cytoskeleton assembly. We have identified a direct interaction between SipC and Exo70, a component of the exocyst complex, which mediates docking and fusion of exocytic vesicles with the plasma membrane. Here, we show that exocyst components coprecipitate with SipC and accumulate at sites of invasion by Salmonella typhimurium. Exocyst assembly requires activation of the small GTPase RalA, which we show is triggered during Salmonella infection by the translocated effector, SopE. Knockdown of RalA or Sec5 results in reduced membrane ruffling at sites of attachment and impairs bacterial entry into host cells. These findings suggest that S. typhimurium enhances invasion efficiency by promoting localized membrane expansion, directly through SipC-dependent recruitment of the exocyst and indirectly via SopE-dependent activation of RalA., (2010 Elsevier Ltd. All rights reserved.)