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Chinese Medical Journal Dec 2023Posttraumatic stress disorder (PTSD) and depression are highly comorbid. Psilocybin exerts substantial therapeutic effects on depression by promoting neuroplasticity....
BACKGROUND
Posttraumatic stress disorder (PTSD) and depression are highly comorbid. Psilocybin exerts substantial therapeutic effects on depression by promoting neuroplasticity. Fear extinction is a key process in the mechanism of first-line exposure-based therapies for PTSD. We hypothesized that psilocybin would facilitate fear extinction by promoting hippocampal neuroplasticity.
METHODS
First, we assessed the effects of psilocybin on percentage of freezing time in an auditory cued fear conditioning (FC) and fear extinction paradigm in mice. Psilocybin was administered 30 min before extinction training. Fear extinction testing was performed on the first day; fear extinction retrieval and fear renewal were tested on the sixth and seventh days, respectively. Furthermore, we verified the effect of psilocybin on hippocampal neuroplasticity using Golgi staining for the dendritic complexity and spine density, Western blotting for the protein levels of brain derived neurotrophic factor (BDNF) and mechanistic target of rapamycin (mTOR), and immunofluorescence staining for the numbers of doublecortin (DCX)- and bromodeoxyuridine (BrdU)-positive cells.
RESULTS
A single dose of psilocybin (2.5 mg/kg, i.p.) reduced the increase in the percentage of freezing time induced by FC at 24 h, 6th day and 7th day after administration. In terms of structural neuroplasticity, psilocybin rescued the decrease in hippocampal dendritic complexity and spine density induced by FC; in terms of neuroplasticity related proteins, psilocybin rescued the decrease in the protein levels of hippocampal BDNF and mTOR induced by FC; in terms of neurogenesis, psilocybin rescued the decrease in the numbers of DCX- and BrdU-positive cells in the hippocampal dentate gyrus induced by FC.
CONCLUSIONS
A single dose of psilocybin facilitated rapid and sustained fear extinction; this effect might be partially mediated by the promotion of hippocampal neuroplasticity. This study indicates that psilocybin may be a useful adjunct to exposure-based therapies for PTSD and other mental disorders characterized by failure of fear extinction.
Topics: Humans; Mice; Animals; Psilocybin; Fear; Extinction, Psychological; Brain-Derived Neurotrophic Factor; Bromodeoxyuridine; Hippocampus; Neuronal Plasticity; TOR Serine-Threonine Kinases
PubMed: 37000971
DOI: 10.1097/CM9.0000000000002647 -
Antiviral Chemistry & Chemotherapy 2023Brivudin, (()-5-(2-bromovinyl)-2'-deoxyuridine (BVDU) can be considered the gold standard for the treatment of varicella-zoster virus (VZV) infections, such as herpes... (Review)
Review
Brivudin, (()-5-(2-bromovinyl)-2'-deoxyuridine (BVDU) can be considered the gold standard for the treatment of varicella-zoster virus (VZV) infections, such as herpes zoster (shingles). It is available for clinical use in most European countries (except for the UK) and over the whole world (except for the US and Canada). Besides VZV its activity spectrum also includes various other herpesviruses, such as herpes simplex virus type 1 (HSV-1). Its activity against VZV and HSV-1 depends on phosphorylation by the virus-encoded thymidine kinase (TK). In its active form (BVDU TP or BVDU 5'-triphosphate), it can act as both substrate and inhibitor of the viral (i.e., HSV-1) DNA polymerase. It has proven to be effective against herpes zoster, including post-herpetic neuralgia (PHN). It is contra-indicated in patients concomitantly treated by 5-fluorouracil (FU), since its degradation product, ()-5-(2-bromovinyl)uracil, is inhibitory to the catabolism of FU, which may enhance the toxicity of the latter. A new compound, the bicyclic nucleoside analogue (BCNA) Cf-1743, has been described, which is a more potent inhibitor of VZV replication than BVDU and which does not interfere with the catabolism of FU. It is applicable orally, as its 5'-valine ester FV-100 (Fermavir), but has not (yet) been marketed for clinical use.
Topics: Humans; Antiviral Agents; Bromodeoxyuridine; Herpesvirus 3, Human; Herpes Zoster; Herpesvirus 1, Human; Fluorouracil; Thymidine Kinase
PubMed: 36710501
DOI: 10.1177/20402066231152971 -
International Journal of Molecular... Feb 2023Cellular growth and the preparation of cells for division between two successive cell divisions is called the cell cycle. The cell cycle is divided into several phases;... (Review)
Review
Cellular growth and the preparation of cells for division between two successive cell divisions is called the cell cycle. The cell cycle is divided into several phases; the length of these particular cell cycle phases is an important characteristic of cell life. The progression of cells through these phases is a highly orchestrated process governed by endogenous and exogenous factors. For the elucidation of the role of these factors, including pathological aspects, various methods have been developed. Among these methods, those focused on the analysis of the duration of distinct cell cycle phases play important role. The main aim of this review is to guide the readers through the basic methods of the determination of cell cycle phases and estimation of their length, with a focus on the effectiveness and reproducibility of the described methods.
Topics: Bromodeoxyuridine; Reproducibility of Results; Cell Cycle; Cell Division; Cell Proliferation
PubMed: 36835083
DOI: 10.3390/ijms24043674 -
Cells Jun 2022Tagging proliferating cells with thymidine analogs is an indispensable research tool; however, the issue of the potential in vivo cytotoxicity of these compounds remains...
Tagging proliferating cells with thymidine analogs is an indispensable research tool; however, the issue of the potential in vivo cytotoxicity of these compounds remains unresolved. Here, we address these concerns by examining the effects of BrdU and EdU on adult hippocampal neurogenesis and EdU on the perinatal somatic development of mice. We show that, in a wide range of doses, EdU and BrdU label similar numbers of cells in the dentate gyrus shortly after administration. Furthermore, whereas the administration of EdU does not affect the division and survival of neural progenitor within 48 h after injection, it does affect cell survival, as evaluated 6 weeks later. We also show that a single injection of various doses of EdU on the first postnatal day does not lead to noticeable changes in a panel of morphometric criteria within the first week; however, higher doses of EdU adversely affect the subsequent somatic maturation and brain growth of the mouse pups. Our results indicate the potential caveats in labeling the replicating DNA using thymidine analogs and suggest guidelines for applying this approach.
Topics: Animals; Bromodeoxyuridine; Cell Count; Cell Proliferation; Mice; Neurogenesis; Thymidine
PubMed: 35741018
DOI: 10.3390/cells11121888 -
Current Protocols Sep 2022Three-dimensional (3D) human organotypic skin cultures provide a physiologically relevant model that recapitulates in vivo skin features. Most commonly, organotypic skin...
Three-dimensional (3D) human organotypic skin cultures provide a physiologically relevant model that recapitulates in vivo skin features. Most commonly, organotypic skin cultures are created by seeding isolated epidermal keratinocytes onto a collagen/fibroblast plug and lifting to an air liquid interface. These conditions are sufficient to drive stratification and differentiation of the keratinocytes to form an epidermal-like sheet with remarkable similarities to human epidermis. Coupled with genetic or pharmacological treatments, these cultures provide a powerful tool for elucidating keratinocyte biology. Recent focus has been placed on increasing the utility of organotypic skin cultures by incorporating other cell types that are present in the skin, such as melanocytes, immune cells, and other cells. Here we describe a step-by-step protocol for the isolation of neonatal human epidermal keratinocytes and melanocytes from foreskins, and the creation of organotypic skin cultures that include both cell types. We also describe methods that can be used to assess melanocyte behavior in these organotypic cultures, including methods for whole mount staining, measurement of melanocyte dendricity, staining for pigment, and 5-bromo-2'-deoxyuridine (BrdU) labeling for identification of proliferating cells. © 2022 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Isolation of primary cells Alternate Protocol: Isolation of primary cells using differential trypsinization Basic Protocol 2: Organotypic culture protocol Support Protocol 1: Culture and maintenance of NHEKs and melanocytes Support Protocol 2: Lentiviral transduction of melanocytes Support Protocol 3: Retroviral transduction of NHEKs Support Protocol 4: Whole mount immunostaining protocol Support Protocol 5: Measuring melanocyte dendricity Support Protocol 6: Fontana-Masson staining protocol Support Protocol 7: BrdU labeling and staining.
Topics: Bromodeoxyuridine; Collagen; Humans; Infant, Newborn; Keratinocytes; Melanocytes; Skin
PubMed: 36165649
DOI: 10.1002/cpz1.536 -
Nature Communications Jun 2022Little is known about replication fork velocity variations along eukaryotic genomes, since reference techniques to determine fork speed either provide no sequence...
Little is known about replication fork velocity variations along eukaryotic genomes, since reference techniques to determine fork speed either provide no sequence information or suffer from low throughput. Here we present NanoForkSpeed, a nanopore sequencing-based method to map and extract the velocity of individual forks detected as tracks of the thymidine analogue bromodeoxyuridine incorporated during a brief pulse-labelling of asynchronously growing cells. NanoForkSpeed retrieves previous Saccharomyces cerevisiae mean fork speed estimates (≈2 kb/min) in the BT1 strain exhibiting highly efficient bromodeoxyuridine incorporation and wild-type growth, and precisely quantifies speed changes in cells with altered replisome progression or exposed to hydroxyurea. The positioning of >125,000 fork velocities provides a genome-wide map of fork progression based on individual fork rates, showing a uniform fork speed across yeast chromosomes except for a marked slowdown at known pausing sites.
Topics: Bromodeoxyuridine; Chromosomes; DNA Replication; Nanopore Sequencing; Saccharomyces cerevisiae
PubMed: 35676270
DOI: 10.1038/s41467-022-31012-0 -
Neuroscience Letters Jun 20195-bromo-2'-dexoyuridine (BrdU) is often used in neuroscience research as a marker of newly-divided cells. However, several studies suggest that BrdU can produce unwanted...
5-bromo-2'-dexoyuridine (BrdU) is often used in neuroscience research as a marker of newly-divided cells. However, several studies suggest that BrdU can produce unwanted side effects, including changes in animal behavior and cellular function. In this study, we investigated the effect of BrdU injections on locomotor behavior in a rodent model of ischemic stroke. Ischemic strokes were induced in adult rats, and 50 mg/kg BrdU was intraperitoneally injected over 5 days beginning 2 weeks post-stroke, while control animals received vehicle. Locomotor activity was evaluated by videotaping the rats in their home cages for 30 min, beginning one hour after BrdU injection. BrdU-injected rats showed a nearly three-fold increase in locomotor activity compared to control animals. These findings suggest that BrdU induces a hyperlocomotor effect in rats following brain injury, pointing to the need for caution when interpreting behavioral results in such studies.
Topics: Animals; Bromodeoxyuridine; Male; Motor Activity; Rats, Long-Evans; Stroke
PubMed: 30853407
DOI: 10.1016/j.neulet.2019.03.007 -
Cells Jun 2021The synthetic halogenated pyrimidine analog, 5-bromo-2'-deoxyuridine (BrdU), is a marker of DNA synthesis. This exogenous nucleoside has generated important insights... (Review)
Review
The synthetic halogenated pyrimidine analog, 5-bromo-2'-deoxyuridine (BrdU), is a marker of DNA synthesis. This exogenous nucleoside has generated important insights into the cellular mechanisms of the central nervous system development in a variety of animals including insects, birds, and mammals. Despite this, the detrimental effects of the incorporation of BrdU into DNA on proliferation and viability of different types of cells has been frequently neglected. This review will summarize and present the effects of a pulse of BrdU, at doses ranging from 25 to 300 µg/g, or repeated injections. The latter, following the method of the progressively delayed labeling comprehensive procedure. The prenatal and perinatal development of the cerebellum are studied. These current data have implications for the interpretation of the results obtained by this marker as an index of the generation, migration, and settled pattern of neurons in the developing central nervous system. Caution should be exercised when interpreting the results obtained using BrdU. This is particularly important when high or repeated doses of this agent are injected. I hope that this review sheds light on the effects of this toxic maker. It may be used as a reference for toxicologists and neurobiologists given the broad use of 5-bromo-2'-deoxyuridine to label dividing cells.
Topics: Animals; Bromodeoxyuridine; Cerebellum; DNA; Humans; Neural Stem Cells; Neurogenesis; Staining and Labeling
PubMed: 34200598
DOI: 10.3390/cells10061453 -
Science (New York, N.Y.) Feb 2019The gut microbiota is implicated in the metabolism of many medical drugs, with consequences for interpersonal variation in drug efficacy and toxicity. However,...
The gut microbiota is implicated in the metabolism of many medical drugs, with consequences for interpersonal variation in drug efficacy and toxicity. However, quantifying microbial contributions to drug metabolism is challenging, particularly in cases where host and microbiome perform the same metabolic transformation. We combined gut commensal genetics with gnotobiotics to measure brivudine drug metabolism across tissues in mice that vary in a single microbiome-encoded enzyme. Informed by these measurements, we built a pharmacokinetic model that quantitatively predicts microbiome contributions to systemic drug and metabolite exposure, as a function of bioavailability, host and microbial drug-metabolizing activity, drug and metabolite absorption, and intestinal transit kinetics. Clonazepam studies illustrate how this approach disentangles microbiome contributions to metabolism of drugs subject to multiple metabolic routes and transformations.
Topics: Animals; Bacteroides thetaiotaomicron; Biological Availability; Biotransformation; Bromodeoxyuridine; Clonazepam; Gastrointestinal Microbiome; Germ-Free Life; Mice
PubMed: 30733391
DOI: 10.1126/science.aat9931 -
Genes Feb 2022Eukaryotes duplicate their chromosomes during the cell cycle S phase using thousands of initiation sites, tunable fork speed and megabase-long spatio-temporal...
Eukaryotes duplicate their chromosomes during the cell cycle S phase using thousands of initiation sites, tunable fork speed and megabase-long spatio-temporal replication programs. The duration of S phase is fairly constant within a given cell type, but remarkably plastic during development, cell differentiation or various stresses. Characterizing the dynamics of S phase is important as replication defects are associated with genome instability, cancer and ageing. Methods to measure S-phase duration are so far indirect, and rely on mathematical modelling or require cell synchronization. We describe here a simple and robust method to measure S-phase duration in cell cultures using a dual EdU-BrdU pulse-labeling regimen with incremental thymidine chases, and quantification by flow cytometry of cells entering and exiting S phase. Importantly, the method requires neither cell synchronization nor genome engineering, thus avoiding possible artifacts. It measures the duration of unperturbed S phases, but also the effect of drugs or mutations on it. We show that this method can be used for both adherent and suspension cells, cell lines and primary cells of different types from human, mouse and . Interestingly, the method revealed that several commonly-used cancer cell lines have a longer S phase compared to untransformed cells.
Topics: Animals; Bromodeoxyuridine; Cell Division; Chromosomes; Flow Cytometry; Mice; S Phase
PubMed: 35327961
DOI: 10.3390/genes13030408