Anhedonia is the inability to experience pleasure from rewarding or enjoyable activities and is a core symptom of depression in humans. Here, we describe a protocol for the measurement of anhedonia ...in mice, in which anhedonia is measured by a sucrose preference test (SPT) based on a two-bottle choice paradigm. A reduction in the sucrose preference ratio in experimental relative to control mice is indicative of anhedonia. To date, inconsistent and variable results have been reported following the use of the SPT by different groups, probably due to the use of different protocols and equipment. In this protocol, we describe how to set up a clearly defined apparatus for SPT and provide a detailed protocol to ensure greater consistency when carrying out SPT. This optimized protocol is highly sensitive, reliable, and adaptable for evaluation of chronic stress-related anhedonia, as well as morphine-induced dependence. The whole SPT, including adaptation, baseline measurement, and testing, takes 8 d.
Nitric oxide (NO), a free gaseous signaling molecule, is involved in the regulation of the cardiovascular, nervous and immune system. The neurotransmitter function of nitric oxide is dependent on ...dynamic regulation of its biosynthetic enzyme, nitric oxide synthase (NOS). There are three types of NOS, neuronal nitric oxide synthase (nNOS), endothelial nitric oxide synthase (eNOS) and inducible nitric oxide synthase (iNOS). Of the three NOS, we focus on nNOS in the present review. Brain nNOS exists in particulate and soluble forms and the differential subcellular localization of nNOS may contribute to its diverse functions. Proteins bearing PDZ domains can interact directly with the PDZ domain of nNOS, influencing the subcellular distribution and/or activity of the enzyme. During the past several years, an increasing number of reports have demonstrated the importance of nNOS in a variety of synaptic signaling events. nNOS has been implicated in modulating physiological functions such as learning, memory, and neurogenesis, as well as being involved in a number of human diseases. In this review we concentrate on recent findings regarding the structural features, subcellular localization and factors regulating nNOS function. In particular, we conclude with a section discussing the role of nNOS in a wide range of physiological and pathological conditions.
Background and Purpose
Anxiety disorder is a common mental health disorder. However, there are few safe and fast‐acting anxiolytic drugs available that can treat anxiety disorder. We previously ...demonstrated that the interaction of neuronal NOS (nNOS) with its carboxy‐terminal PDZ ligand (CAPON) is involved in regulating anxiety‐related behaviours. Here, we further investigated the anxiolytic effects of nNOS–CAPON disruptors in chronic stress‐induced anxiety in animals.
Experimental Approach
Mice were intravenously treated with nNOS–CAPON disruptors, ZLc‐002 or Tat‐CAPON12C, at the last week of chronic mild stress (CMS) exposure. We also infused corticosterone (CORT) into the hippocampus of mice to model anxiety behaviours and also delivered ZLc‐002 or Tat‐CAPON12C on the last week of chronic CORT treatment via pre‐implanted cannula. Anxiety‐related behaviours were examined using elevated plus maze, open field, novelty‐suppressed feeding and light–dark (LD) tests. The level of nNOS–CAPON interaction was determined by co‐immunoprecipitation (CO‐IP) and proximity ligation assay (PLA). The neural mechanisms underlying the behavioural effects of nNOS–CAPON uncoupling in anxiety animal models were assessed by western blot, immunofluorescence and Golgi‐Cox staining.
Key Results
ZLc‐002 and Tat‐CAPON12C reversed CMS‐ or CORT‐induced anxiety‐related behaviours. ZLc‐002 and Tat‐CAPON12C increased synaptogenesis along with improved dendritic remodelling in CMS mice or CORT‐treated cultured neurons. Meanwhile, blocking nNOS–CAPON interaction significantly activated the cAMP response element‐binding protein (CREB)–brain‐derived neurotrophic factor (BDNF) pathway, which is associated with synaptic plasticity.
Conclusion and Implications
Collectively, these results provide evidence for the anxiolytic effects of nNOS–CAPON uncouplers and their underlying mechanisms in anxiety disorders.
Exosomes derived from bone marrow mesenchymal stem cells can inhibit neuroinflammation through regulating microglial phenotypes and promoting nerve injury repair. However, the underlying molecular ...mechanism remains unclear. In this study, we investigated the mechanism by which exosomes derived from bone marrow mesenchymal stem cells inhibit neuroinflammation. Our in vitro co-culture experiments showed that bone marrow mesenchymal stem cells and their exosomes promoted the polarization of activated BV2 microglia to their anti-inflammatory phenotype, inhibited the expression of proinflammatory cytokines, and increased the expression of anti-inflammatory cytokines. Our in vivo experiments showed that tail vein injection of exosomes reduced cell apoptosis in cortical tissue of mouse models of traumatic brain injury, inhibited neuroinflammation, and promoted the transformation of microglia to the anti-inflammatory phenotype. We screened some microRNAs related to neuroinflammation using microRNA sequencing and found that microRNA-181b seemed to be actively involved in the process. Finally, we regulated the expression of miR181b in the brain tissue of mouse models of traumatic brain injury using lentiviral transfection. We found that miR181b overexpression effectively reduced apoptosis and neuroinflamatory response after traumatic brain injury and promoted the transformation of microglia to the anti-inflammatory phenotype. The interleukin 10/STAT3 pathway was activated during this process. These findings suggest that the inhibitory effects of exosomes derived from bone marrow mesenchymal stem cells on neuroinflamation after traumatic brain injury may be realized by the action of miR181b on the interleukin 10/STAT3 pathway.
Anxiety is common in patients suffering from chronic pain. Here, we report anxiety-like behaviors in mouse models of chronic pain and reveal that nNOS-expressing neurons in ventromedial prefrontal ...cortex (vmPFC) are essential for pain-induced anxiety but not algesia, using optogenetic and chemogenetic strategies. Additionally, we determined that excitatory projections from the posterior subregion of paraventricular thalamic nucleus (pPVT) provide a neuronal input that drives the activation of vmPFC nNOS-expressing neurons in our chronic pain models. Our results suggest that the pain signal becomes an anxiety signal after activation of vmPFC nNOS-expressing neurons, which causes subsequent release of nitric oxide (NO). Finally, we show that the downstream molecular mechanisms of NO likely involve enhanced glutamate transmission in vmPFC CaMKIIα-expressing neurons through S-nitrosylation-induced AMPAR trafficking. Overall, our data suggest that pPVT excitatory neurons drive chronic pain-induced anxiety through activation of vmPFC nNOS-expressing neurons, resulting in NO-mediated AMPAR trafficking in vmPFC pyramidal neurons.
Background
Tyrophagus putresecentiae is an important mite species in rural and urban environments, causing sensitization and allergic disease. While evidence suggests that microRNAs (miRNAs) may ...regulate the expression of allergen‐encoding genes, no study has directly investigated this possibility. Here, this gap was addressed by profiling miRNAs and elucidating their target allergen messenger RNAs (mRNAs) in this mite species.
Methods
Small RNA and transcriptome libraries were constructed for eggs, larvae, nymphs, and adults. After deep miRNA and whole‐transcriptome sequencing were performed, the miRNA and allergen‐encoding mRNA regulatory networks were explored.
Results
A total of 540 miRNAs were identified, including 155 with expression levels differing significantly across the four mite developmental stages (p < .01), 59 of which were novel. The mRNA expression for allergens was higher for Tyr p 1 in adults than in other developmental stages; Tyr p 2–5, 7, 10, 13, 33, and 34 in immature stages; and Tyr p 28, 35, and 36 in eggs and adults. A combined miRNA and transcriptome bioinformatics analysis showed that allergen Tyr p 3 was regulated by miRNA PC‐5p‐5698441_1, Tyr p 4 was regulated by PC‐5p‐7050653_1, and Tyr p 34 was regulated by PC‐5p‐5534223_1 and PC‐5p‐5698441_1. These three allergen mRNA and three miRNAs were identified using qRT‐PCR, and their regulatory roles were confirmed by double‐fluorescent reporter gene system and site‐directed mutagenesis technology.
Conclusions
For the first time, allergen mRNA expression and miRNAs were profiled throughout the life cycle for an allergen‐producing mite, and the results showed that miRNAs bind to target allergen mRNAs to regulate their expression.
We collected the eggs, larvae, nymphs, and adults of Tyrophagus putresecentiae. We verified mRNA (Tyr p 3, Tyr p 4, and Tyr p 34) and microRNA (PC‐5p‐5698441_1, PC‐5p‐7050653_1, PC‐5p‐5534223_1, and PC‐5p‐5698441_1) expression by RT‐PCR. The luciferase reporter assay showed that Tyr p 3 was regulated by the miRNA PC‐5p‐5698441_1, Tyr p 4 was regulated by PC‐5p‐7050653_1, and Tyr p 34 was regulated by both PC‐5p‐5534223_1 and PC‐5p‐5698441_1.
Major depressive disorder (MDD) is one of the most common mental disorders. We designed a fast-onset antidepressant that works by disrupting the interaction between the serotonin transporter (SERT) ...and neuronal nitric oxide synthase (nNOS) in the dorsal raphe nucleus (DRN). Chronic unpredictable mild stress (CMS) selectively increased the SERT-nNOS complex in the DRN in mice. Augmentation of SERT-nNOS interactions in the DRN caused a depression-like phenotype and accounted for the CMS-induced depressive behaviors. Disrupting the SERT-nNOS interaction produced a fast-onset antidepressant effect by enhancing serotonin signaling in forebrain circuits. We discovered a small-molecule compound, ZZL-7, that elicited an antidepressant effect 2 hours after treatment without undesirable side effects. This compound, or analogous reagents, may serve as a new, rapidly acting treatment for MDD.
Anxiety disorders are associated with a high social burden worldwide. Recently, increasing evidence suggests that nuclear factor kappa B (NF‐κB) has significant implications for psychiatric diseases, ...including anxiety and depressive disorders. However, the molecular mechanisms underlying the role of NF‐κB in stress‐induced anxiety behaviors are poorly understood. In this study, we show that chronic mild stress (CMS) and glucocorticoids dramatically increased the expression of NF‐κB subunits p50 and p65, phosphorylation and acetylation of p65, and the level of nuclear p65 in vivo and in vitro, implicating activation of NF‐κB signaling in chronic stress‐induced pathological processes. Using the novelty‐suppressed feeding (NSF) and elevated‐plus maze (EPM) tests, we found that treatment with pyrrolidine dithiocarbamate (PDTC; intra‐hippocampal infusion), an inhibitor of NF‐κB, rescued the CMS‐ or glucocorticoid‐induced anxiogenic behaviors in mice. Microinjection of PDTC into the hippocampus reversed CMS‐induced up‐regulation of neuronal nitric oxide synthase (nNOS), carboxy‐terminal PDZ ligand of nNOS (CAPON), and dexamethasone‐induced ras protein 1 (Dexras1) and dendritic spine loss of dentate gyrus (DG) granule cells. Moreover, over‐expression of CAPON by infusing LV‐CAPON‐L‐GFP into the hippocampus induced nNOS‐Dexras1 interaction and anxiety‐like behaviors, and inhibition of NF‐κB by PDTC reduced the LV‐CAPON‐L‐GFP‐induced increases in nNOS‐Dexras1 complex and anxiogenic‐like effects in mice. These findings indicate that hippocampal NF‐κB mediates anxiogenic behaviors, probably via regulating the association of nNOS‐CAPON‐Dexras1, and uncover a novel approach to the treatment of anxiety disorders.
The molecular mechanisms underlying anxiety behaviors are poorly understood. Here, we propose a model of a signaling cascade whereby hippocampal nuclear factor kappa B (NF‐κB) contributes to chronic stress‐induced anxiety behaviors. Activation of NF‐κB signaling by chronic stress‐stimulated glucocorticoid overproduction leads to up‐regulation of neuronal nitric oxide synthase (nNOS), carboxy‐terminal PDZ ligand of nNOS (CAPON), and dexamethasone‐induced ras protein 1 (Dexras1) expression and nNOS‐CAPON‐Dexras1 association, thus inducing anxiety‐related behaviors. Our findings suggest a new target for the intervention of anxiety‐related disorders.
Deep neural networks are a powerful tool for characterizing quantum states. Existing networks are typically trained with experimental data gathered from the quantum state that needs to be ...characterized. But is it possible to train a neural network offline, on a different set of states? Here we introduce a network that can be trained with classically simulated data from a fiducial set of states and measurements, and can later be used to characterize quantum states that share structural similarities with the fiducial states. With little guidance of quantum physics, the network builds its own data-driven representation of a quantum state, and then uses it to predict the outcome statistics of quantum measurements that have not been performed yet. The state representations produced by the network can also be used for tasks beyond the prediction of outcome statistics, including clustering of quantum states and identification of different phases of matter.
Summary
In neurons, increased protein–protein interactions between neuronal nitric oxide synthase (nNOS) and its carboxy‐terminal PDZ ligand (CAPON) contribute to excitotoxicity and abnormal ...dendritic spine development, both of which are involved in the development of Alzheimer's disease. In models of Alzheimer's disease, increased nNOS–CAPON interaction was detected after treatment with amyloid‐β in vitro, and a similar change was found in the hippocampus of APP/PS1 mice (a transgenic mouse model of Alzheimer's disease), compared with age‐matched background mice in vivo. After blocking the nNOS–CAPON interaction, memory was rescued in 4‐month‐old APP/PS1 mice, and dendritic impairments were ameliorated both in vivo and in vitro. Furthermore, we demonstrated that S‐nitrosylation of Dexras1 and inhibition of the ERK–CREB–BDNF pathway might be downstream of the nNOS–CAPON interaction.
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