Characterizing the post-traumatic growth trajectory in gastric cancer survivors: a population-based longitudinal study.

OBJECTIVES: Post-traumatic growth can improve the quality of life of cancer survivors. The objective of this study was to investigate post-traumatic growth heterogeneity trajectory in perioperative gastric cancer survivors, and to identify characteristics that predict membership for each trajectory. METHODS: Gastric cancer survivors (n = 403) were recruited before surgery, their baseline assessment (including post-traumatic growth and related characteristics) was completed, and post-traumatic growth levels were followed up on the day they left the intensive care unit, at discharge, and 1 month after discharge. Latent growth mixture mode was used to identify the heterogeneous trajectory of post-traumatic growth, and the core predictors of trajectory subtypes were explored using a decision tree model. RESULTS: Three post-traumatic growth development trajectories were identified among gastric cancer survivors: stable high of PTG group (20.6%), fluctuation of PTG group (44.4%), persistent low of PTG group (35.0%). The decision tree model showed anxiety, coping style, and psychological resilience-which was the primary predictor-might be used to predict the PTG trajectory subtypes of gastric cancer survivors. CONCLUSIONS: There was considerable variability in the experience of post-traumatic growth among gastric cancer survivors. Recognition of high-risk gastric cancer survivors who fall into the fluctuation or persistent low of PTG group and provision of psychological resilience-centered support might allow medical professionals to improve patients' post-traumatic growth and mitigate the impact of negative outcomes.

Cell Membrane Perforation: Patterns, Mechanisms and Functions.

Cell membrane is crucial for the cellular activities, and any disruption to it may affect the cells. It is demonstrated that cell membrane perforation is associated with some biological processes like programmed cell death (PCD) and infection of pathogens. Specific developments make it a promising technique to perforate the cell membrane controllably and precisely. The pores on the cell membrane provide direct pathways for the entry and exit of substances, and can also cause cell death, which means reasonable utilization of cell membrane perforation is able to assist intracellular delivery, eliminate diseased or cancerous cells, and bring about other benefits. This review classifies the patterns of cell membrane perforation based on the mechanisms into 1) physical patterns, 2) biological patterns, and 3) chemical patterns, introduces the characterization methods and then summarizes the functions according to the characteristics of reversible and irreversible pores, with the aim of providing a comprehensive summary of the knowledge related to cell membrane perforation and enlightening broad applications in biomedical science.

Covalent organic frameworks: linkage types, synthetic methods and bio-related applications.

Covalent organic frameworks (COFs) are composed of small organic molecules linked via covalent bonds, which have tunable mesoporous structure, good biocompatibility and functional diversities. These excellent properties make COFs a promising candidate for constructing biomedical nanoplatforms and provide ample opportunities for nanomedicine development. A systematic review of the linkage types and synthesis methods of COFs is of indispensable value for their biomedical applications. In this review, we first summarize the types of various linkages of COFs and their corresponding properties. Then, we highlight the reaction temperature, solvent and reaction time required by different synthesis methods and show the most suitable synthesis method by comparing the merits and demerits of various methods. To appreciate the cutting-edge research on COFs in bioscience technology, we also summarize the bio-related applications of COFs, including drug delivery, tumor therapy, bioimaging, biosensing and antimicrobial applications. We hope to provide insight into the interdisciplinary research on COFs and promote the development of COF nanomaterials for biomedical applications and their future clinical translations.

Identification of circulating T-cell immunoglobulin and mucin domain 4 as a potential biomarker for coronary heart disease.

Efferocytosis, the process of engulfing and removing apoptotic cells, is attenuated in vulnerable plaques of advanced atherosclerosis. T-cell immunoglobulin and mucin domain 4 (TIMD4) is a recognition receptor protein for efferocytosis that has been implicated in atherosclerosis mouse models. However, the role of serum-soluble TIMD4 (sTIMD4) in coronary heart disease (CHD) remains unknown. In this study, we analyzed serum samples collected from two groups: Group 1 (36 healthy controls and 70 CHD patients) and Group 2 (44 chronic coronary syndrome [CCS]) and 81 acute coronary syndrome [ACS] patients). We found that sTIMD4 levels in patients with CHD were significantly higher than those in healthy controls and were also higher in ACS than in CCS patients. The area under the receiver operating characteristic curve was 0.787. Furthermore, our in vitro results showed that low-density lipoprotein/lipopolysaccharide activated p38 mitogen-activated protein kinase, which in turn enhanced a disintegrin and metalloproteinase 17, resulting in increased secretion of sTIMD4. This impairment of macrophage efferocytosis promoted inflammation. Thus, this study is not only the first identification of a potential novel biomarker of CHD, sTIMD4, but also demonstrated its pathogenesis mechanism, providing a new direction for the diagnosis and treatment of CHD.

Glabridin Ameliorates Alcohol-Caused Liver Damage by Reducing Oxidative Stress and Inflammation via p38 MAPK/Nrf2/NF-κB Pathway.

Licorice is a traditional and versatile herbal medicine and food. Glabridin (Gla) is a kind of isoflavone extracted from the licorice root, which has anti-obesity, anti-atherosclerotic, and antioxidative effects. Alcoholic liver disease (ALD) is a widespread liver disease induced by chronic alcohol consumption. However, studies demonstrating the effect of Gla on ALD are rare. The research explored the positive effect of Gla in C57BL/6J mice fed by the Lieber-DeCarli ethanol mice diet and HepG2 cells treated with ethanol. Gla alleviated ethanol-induced liver injury, including reducing liver vacuolation and lipid accumulation. The serum levels of inflammatory cytokines were decreased in the Gla-treated mice. The reactive oxygen species and apoptosis levels were attenuated and antioxidant enzyme activity levels were restored in ethanol-induced mice by Gla treatment. In vitro, Gla reduced ethanol-induced cytotoxicity, nuclear factor kappa B (NF-κB) nuclear translocation, and enhanced nuclear factor (erythroid-derived 2)-like 2 (Nrf2) nuclear translocation. Anisomycin (an agonist of p38 MAPK) eliminated the positive role of Gla on ethanol-caused oxidative stress and inflammation. On the whole, Gla can alleviate alcoholic liver damage via the p38 MAPK/Nrf2/NF-κB pathway and may be used as a novel health product or drug to potentially alleviate ALD.

6-Methyl flavone inhibits Nogo-B expression and improves high fructose diet-induced liver injury in mice.

Excessive fructose consumption increases hepatic de novo lipogenesis, resulting in cellular stress, inflammation and liver injury. Nogo-B is a resident protein of the endoplasmic reticulum that regulates its structure and function. Hepatic Nogo-B is a key protein in glycolipid metabolism, and inhibition of Nogo-B has protective effects against metabolic syndrome, thus small molecules that inhibit Nogo-B have therapeutic benefits for glycolipid metabolism disorders. In this study we tested 14 flavones/isoflavones in hepatocytes using dual luciferase reporter system based on the Nogo-B transcriptional response system, and found that 6-methyl flavone (6-MF) exerted the strongest inhibition on Nogo-B expression in hepatocytes with an IC50 value of 15.85 µM. Administration of 6-MF (50 mg· kg-1 ·d-1, i.g. for 3 weeks) significantly improved insulin resistance along with ameliorated liver injury and hypertriglyceridemia in high fructose diet-fed mice. In HepG2 cells cultured in a media containing an FA-fructose mixture, 6-MF (15 µM) significantly inhibited lipid synthesis, oxidative stress and inflammatory responses. Furthermore, we revealed that 6-MF inhibited Nogo-B/ChREBP-mediated fatty acid synthesis and reduced lipid accumulation in hepatocytes by restoring cellular autophagy and promoting fatty acid oxidation via the AMPKα-mTOR pathway. Thus, 6-MF may serve as a potential Nogo-B inhibitor to treat metabolic syndrome caused by glycolipid metabolism dysregulation.

Application of the Knowledge, Attitude, and Practice model combined with motivational interviewing for health education in female patients with systemic lupus erythematosus.

To explore the application effect of the Knowledge, Attitude, and Practice (KAP) model combined with motivational interviewing for health education in the chronic disease management of female patients with systemic lupus erythematosus (SLE). In this study, 84 women with SLE who were admitted to a tertiary hospital in Tianjin from July 2021 to April 2022 were enrolled in this study and divided into observation (n = 42) and control groups (n = 42). The control group received routine health education and treatment for chronic diseases. Based on the control group, the KAP method and questionnaire survey were adopted. Health literacy and compliance in the 2 groups were compared in the first and third months after the intervention. The observation group had a higher total score of health literacy in the third month than the control group. From before the intervention to the first and third months, improvement in the observation group was compared with that in the control group (F = 36.543, P 6..001; F = 4.884, P = .03; F = 23.881, P 3..001). The observation group had a higher total compliance score in the third month than the control group (t = 5.101, P = .007). From before the intervention to the first and third months of the intervention, the improvement in the observation group demonstrated an interaction with the time group compared with that in the control group (F = 68.116, P 8..001; F = 4.884, P = .032; F = 24.789, P < .001). Motivational interviewing based on the KAP model is effective in the short-term overall health literacy of female patients with SLE, especially in terms of communication, health improvement, and information acquisition; after 3 months, it can influence and maintain high patient compliance.

Overexpression of NgBR inhibits high-fat diet-induced atherosclerosis in ApoE-deficiency mice.

BACKGROUND: Hyperlipidemia (hypercholesterolemia and/or hypertriglyceridemia) is a risk factor for atherosclerosis. Nogo-B receptor (NgBR) plays important roles in hepatic steatosis and cholesterol transport. However, the effect of NgBR overexpression on atherosclerosis remains unknown. MATERIALS AND METHODS: Apolipoprotein E deficient (ApoE-/-) mice infected with adeno-associated virus (AAV)-NgBR expression vector were fed a high-fat diet for 12 weeks, followed by determination of atherosclerosis and the involved mechanisms. RESULTS: We determined that high expression of NgBR by AAV injection mainly occurs in the liver and it can substantially inhibit en face and aortic root sinus lesions. NgBR overexpression also reduced levels of inflammatory factors in the aortic root and serum, and levels of cholesterol, triglyceride, and free fatty acids in the liver and serum. Mechanistically, NgBR overexpression increased the expression of scavenger receptor type BI and the genes for bile acid synthesis, and decreased the expression of cholesterol synthesis genes by reducing sterol regulatory element-binding protein 2 maturation in the liver, thereby reducing hypercholesterolemia. In addition, NgBR overexpression activated AMP-activated protein kinase α via the Ca2+ signaling pathway, which inhibited fat synthesis and improved hypertriglyceridemia. CONCLUSIONS: Taken together, our study demonstrates that overexpression of NgBR enhanced cholesterol metabolism and inhibited cholesterol/fatty acid synthesis to reduce hyperlipidemia, and reduced vascular inflammation, thereby inhibiting atherosclerosis in ApoE-/- mice. Our study indicates that NgBR might be a potential target for atherosclerosis treatment.

Loss of parkin causes endoplasmic reticulum calcium dyshomeostasis by upregulation of reticulocalbin 1.

Increasing evidence suggests that astrocytes play an important role in the progression of Parkinson's disease (PD). Previous studies on our parkin knockout mouse demonstrated a higher accumulation of damaged mitochondria in astrocytes than in surrounding dopaminergic (DA) neurons, suggesting that Parkin plays a crucial role regarding their interaction during PD pathogenesis. In the current study, we examined primary mesencephalic astrocytes and neurons in a direct co-culture system and discovered that the parkin deletion causes an impaired differentiation of mesencephalic neurons. This effect required the parkin mutation in astrocytes as well as in neurons. In Valinomycin-treated parkin-deficient astrocytes, ubiquitination of Mitofusin 2 was abolished, whereas there was no significant degradation of the outer mitochondrial membrane protein Tom70. This result may explain the accumulation of damaged mitochondria in parkin-deficient astrocytes. We examined differential gene expression in the substantia nigra region of our parkin-KO mouse by RNA sequencing and identified an upregulation of the endoplasmic reticulum (ER) Ca2+ -binding protein reticulocalbin 1 (RCN1) expression, which was validated using qPCR. Immunostaining of the SN brain region revealed RCN1 expression mainly in astrocytes. Our subcellular fractionation of brain extract has shown that RCN1 is located in the ER and in mitochondria-associated membranes (MAM). Moreover, a loss of Parkin function reduced ATP-stimulated calcium-release in ER mesencephalic astrocytes that could be attenuated by siRNA-mediated RCN1 knockdown. Our results indicate that RCN1 plays an important role in ER-associated calcium dyshomeostasis caused by the loss of Parkin function in mesencephalic astrocytes, thereby highlighting the relevance of astrocyte function in PD pathomechanisms.

N-Oxide Zwitterion Functionalized Positively Charged Polyamide Composite Membranes for Nanofiltration.

N-Oxide zwitterionic polyethyleneimine (ZPEI), a new kind of aqueous phase monomer synthesized by commercially branched polyethyleneimine (PEI) via oxidation reaction, was prepared for fabrication of thin-film composite (TFC) polyamide membranes via interfacial polymerization. The main factors, including the monomer concentration and immersion time of the aqueous phase and organic phase, were investigated. Compared with PEI-TFC membranes, the obtained optimal defect-free ZPEI-TFC membranes exhibited a lower roughness (3.3 ± 0.3 nm), a better surface hydrophilicity, and a smaller pore size (238 Da of MWCO). The positively charged ZPEI-TFC membranes (isoelectric point at pH 8.05) showed higher rejections toward both divalent cationic (MgCl2, 93.0%) and anionic (Na2SO4, 96.1%) salts with a water permeation flux of up to 81.0 L·m-2·h-1 at 6 bar, which surpassed currently reported membranes. More importantly, mainly owing to N-oxide zwitterion with strong hydration capability, ZPEI-TFC membranes displayed a high flux recovery ratio (97.0%) toward a model protein contaminant (bovine serum albumin), indicating good anti-fouling properties. Therefore, the novel N-oxide zwitterion functionalized positively charged nanofiltration membranes provide an alternative for water desalination and sewage reclamation.

GLO1 Contributes to the Drug Resistance of Escherichia coli Through Inducing PER Type of Extended-Spectrum ß-Lactamases.

Background: Escherichia coli-associated antimicrobial resistance (AMR) issue so far needs urgent considerations. This study aims to screen the potent genes associated with extended-spectrum ß-lactamases (ESBLs) in drug-resistant Escherichia coli and elucidate the specific drug-resistant mechanism. Methods: Clinical ESBLs-EC samples were obtained based on the microbial identification, and the whole genome was sequenced. In combination with the significantly enriched pathways, several differently expressed genes were screened and verified by RT-PCR. Furthermore, through knocking out glyoxalase 1 (GLO1) gene and transfecting overexpressed plasmids, the potential relationship between GLO1 and ESBLs was then investigated. Lastly, the concentrations of ß-lactamases in bacteria and supernatant from different groups were examined by enzyme-linked immunosorbent assay (ELISA). Results: After successful isolation and identification of ESBLs-EC, the whole genome and eighteen differential metabolic pathways were analyzed to select differently expressed genes, including add, deoD, guaD, speG, GLO1, VNN1, etc. RT-PCR results showed that there were no differences in these genes between the standard bacteria and susceptible Escherichia coli. Remarkably, the relative levels of four genes including speG, Hdac10, GLO1 and Ppcdc were significantly increased in ESBLs-EC in comparison with susceptible strains, whereas other gene expression was decreased. Further experiments utilizing gene knockout and overexpression strains confirmed the role of GLO1. At last, a total of 10 subtypes of ß-lactamases were studied using ELISA, including BES-, CTX-M1-, CTX-M2-, OXA1-, OXA2-, OXA10-, PER-, SHV-, TEM-, and VEB-ESBLs, and results demonstrated that GLO1 gene expression only affected PER-ß-lactamases but had no effects on other ß-lactamases. Conclusion: SpeG, Hdac10, GLO1 and Ppcdc might be associated with the drug-resistant mechanism of Escherichia coli. Of note, this study firstly addressed the role of GLO1 in the drug resistance of ESBLs-EC, and this effect may be mediated by increasing PER-ß-lactamases.

ITRAQ-based proteomic analysis reveals possible target-related proteins in human adrenocortical adenomas.

BACKGROUND: Adrenocortical adenomas (ACAs) can lead to the autonomous secretion of aldosterone responsible for primary aldosteronism (PA), which is the most common form of secondary arterial hypertension. However, the authentic fundamental mechanisms underlying ACAs remain unclear. OBJECTIVE: Isobaric tags for relative and absolute quantitation (iTRAQ)-based proteomics and bioinformatics analyses from etiological studies of ACAs were performed to screen the differentially expressed proteins (DEPs) and investigate the relevant mechanisms of their occurrence and development. Results could help determine therapeutic targets of clinical significance. METHODS: In the present study, iTRAQ-based proteomics was applied to analyze ACA tissue samples from normal adrenal cortex tissues adjacent to the tumor. Using proteins extracted from a panel of four pairs of ACA samples, we identified some upregulated proteins and other downregulated proteins in all four pairs of ACA samples compared with adjacent normal tissue. Subsequently, we predicted protein-protein interaction networks of three DEPs to determine the authentic functional factors in ACA. RESULTS: A total of 753 DEPs were identified, including 347 upregulated and 406 downregulated proteins. The expression of three upregulated proteins (E2F3, KRT6A, and ALDH1A2) was validated by Western blot in 24 ACA samples. Our data suggested that some DEPs might be important hallmarks during the development of ACA. CONCLUSIONS: This study is the first proteomic research to investigate alterations in protein levels and affected pathways in ACA using the iTRAQ technique. Thus, this study not only provides a comprehensive dataset on overall protein changes but also sheds light on its potential molecular mechanism in human ACAs.

Hypoxia facilitates neurogenic dural plasma protein extravasation in mice: a novel animal model for migraine pathophysiology.

Migraine animal models generally mimic the onset of attacks and acute treatment processes. A guinea pig model used the application of meta-chlorophenylpiperazine (mCPP) to trigger immediate dural plasma protein extravasation (PPE) mediated by 5-HT2B receptors. This model has predictive value for antimigraine drugs but cannot explain the delayed onset of efficacy of 5-HT2B receptor antagonists when clinically used for migraine prophylaxis. We found that mCPP failed to induce dural PPE in mice. Considering the role 5-HT2B receptors play in hypoxia-induced pulmonary vessel muscularization, we were encouraged to keep mice under hypoxic conditions and tested whether this treatment will render them susceptible to mCPP-induced dural PPE. Following four-week of hypoxia, PPE, associated with increased transendothelial transport, was induced by mCPP. The effect was blocked by sumatriptan. Chronic application of 5-HT2B receptor or nitric oxide synthase blockers during hypoxia prevented the development of susceptibility. Here we present a migraine model that distinguishes between a migraine-like state (hypoxic mice) and normal, normoxic mice and mimics processes that are related to chronic activation of 5-HT2B receptors under hypoxia. It seems striking, that chronic endogenous activation of 5-HT2B receptors is crucial for the sensitization since 5-HT2B receptor antagonists have strong, albeit delayed migraine prophylactic efficacy.

BF-1--a novel selective 5-HT2B receptor antagonist blocking neurogenic dural plasma protein extravasation in guinea pigs.

Serotonin 5-HT2B receptor antagonists have been proposed as migraine prophylactic drugs, but previously available 5-HT2B receptor antagonists displayed multiple monoaminergic side effects and had to be withdrawn from the market. Here, we set out to identify a novel antagonist with high affinity and selectivity towards 5-HT2B receptors. To test the affinity of new compounds towards various receptors, we generated a broad series of cells functionally coupling human monoaminergic receptors to luciferase. Using the cell lines we revealed pimethixene (1-methyl-4-(9H-thioxanthen-9-ylidene)piperidine) as highly potent, albeit non-selective 5-HT2B receptor antagonist and optimized its chemical structure to create highly potent and selective 5-HT2B receptor antagonists. We selected the methoxythioxanthene BF-1 for further analysis. In comparison to pimethixene, it lacked high affinities to 5-HT1A, 5-HT2A, 5-HT2C, histamine H1, dopamine D1 and D2 as well as muscarinic M1 and M2 receptors. BF-1 was tested as potential migraine prophylactic drug by blocking meta-chlorophenylpiperazine, (mCPP) or BW723C86 (5-((thiophen-2-yl)methoxy)-α-methyltryptamine) induced neurogenic dural plasma protein extravasation in a guinea pig model that may resemble a migraine attack. BF-1 was significantly more potent in this assay compared to the well know non-selective 5-HT2B antagonists, methysergide ((6aR,9R)-N-[(2S)-1-Hydroxybutan-2-yl]-4,7-dimethyl-6,6a,8,9-tetrahydroindolo[4,3-fg]quinoline-9-carboxamide) or pizotifen (4-(1-methyl-4-piperidylidine)-9,10-dihydro-4H-benzo-[4,5]cyclohepta[1,2]-thiophene). Therefore, we propose BF-1 as a new compound that may be developed for prophylactic migraine treatment without the typical monoaminergic side effects.

Synaptic adhesion molecules in Cadm family at the neuromuscular junction.

RA175/SynCAM1/Cadm1 (Cadm1), a member of the immunoglobulin superfamily, is a synaptic cell adhesion molecule that has a PDZ-binding motif at the C-terminal region. It promotes the formation of presynaptic terminals and induces functional synapses in the central nervous system. Cadm1-deficient (knockout [KO]) mice show behavioral abnormalities, including excessive aggression and anxiety, but do not show any symptoms of neuromuscular disorder, although neuromuscular junctions (NMJs) have structures similar to synapses. We have examined the expression of members of the Cadm family in the mouse muscle tissues. Cadm4 and Cadm1 were major components of the Cadm family, and Cadm3 was faintly detected, but Cadm2 was not detected by RT-PCR. Cadm4 as well as Cadm1 colocalized with alpha-bungarotoxin at the NMJs and interacted with the multiple PDZ domain protein Mupp1. Cadm4 was expressed in Cadm1-KO mice and might compensate for Cadm1 loss through interactions with Mupp1.

GABAA receptors involved in sleep and anaesthesia: ß1- versus ß3-containing assemblies.

The histaminergic neurons of the posterior hypothalamus (tuberomamillary nucleus-TMN) control wakefulness, and their silencing through activation of GABA(A) receptors (GABA(A)R) induces sleep and is thought to mediate sedation under propofol anaesthesia. We have previously shown that the ß1 subunit preferring fragrant dioxane derivatives (FDD) are highly potent modulators of GABA(A)R in TMN neurons. In recombinant receptors containing the ß3N265M subunit, FDD action is abolished and GABA potency is reduced. Using rat, wild-type and ß3N265M mice, FDD and propofol, we explored the relative contributions of ß1- and ß3-containing GABA(A)R to synaptic transmission from the GABAergic sleep-on ventrolateral preoptic area neurons to TMN. In ß3N265M mice, GABA potency remained unchanged in TMN neurons, but it was decreased in cultured posterior hypothalamic neurons with impaired modulation of GABA(A)R by propofol. Spontaneous and evoked GABAergic synaptic currents (IPSC) showed ß1-type pharmacology, with the same effects achieved by 3 µM propofol and 10 µM PI24513. Propofol and the FDD PI24513 suppressed neuronal firing in the majority of neurons at 5 and 100 µM, and in all cells at 10 and 250 µM, respectively. FDD given systemically in mice induced sedation but not anaesthesia. Propofol-induced currents were abolished (1-6 µM) or significantly reduced (12 µM) in ß3N265M mice, whereas gating and modulation of GABA(A)R by PI24513 as well as modulation by propofol were unchanged. In conclusion, ß1-containing (FDD-sensitive) GABA(A)R represent the major receptor pool in TMN neurons responding to GABA, while ß3-containing (FDD-insensitive) receptors are gated by low micromolar doses of propofol. Thus, sleep and anaesthesia depend on different GABA(A)R types.

Spinal cord pathology in alpha-synuclein transgenic mice.

Accumulation of α-synuclein is observed in neurodegenerative diseases like Parkinson's disease and Multiple System Atrophy. In previous studies with transgenic C57BL/6 mice overexpressing α-synuclein carrying the mutations A53T and A30P found in Parkinson's disease or with a parkin-null background, we reported severe mitochondrial impairments in neurons and to a larger extent in glial cells of the mesencephalon. Neuron death was not observed in the brain. Here we show that the mice show severe motor impairments in behavioral tests. In addition, these mice exhibit astrocytic cell death in the spinal cord, accompanied by extensive gliosis and microglial activation. This is shown by cell death staining and immunohistochemistry. Ultrastructural analyses revealed severe mitochondrial impairments not only in astrocytes, but also in oligodendrocytes and, to a small extent, in neurons. Thus, the transgenic mice show a profound pathology in glial cells of the spinal cord.

Fragrant dioxane derivatives identify beta1-subunit-containing GABAA receptors.

Nineteen GABA(A) receptor (GABA(A)R) subunits are known in mammals with only a restricted number of functionally identified native combinations. The physiological role of beta1-subunit-containing GABA(A)Rs is unknown. Here we report the discovery of a new structural class of GABA(A)R positive modulators with unique beta1-subunit selectivity: fragrant dioxane derivatives (FDD). At heterologously expressed alpha1betaxgamma2L (x-for 1,2,3) GABA(A)R FDD were 6 times more potent at beta1- versus beta2- and beta3-containing receptors. Serine at position 265 was essential for the high sensitivity of the beta1-subunit to FDD and the beta1N286W mutation nearly abolished modulation; vice versa the mutation beta3N265S shifted FDD sensitivity toward the beta1-type. In posterior hypothalamic neurons controlling wakefulness GABA-mediated whole-cell responses and GABAergic synaptic currents were highly sensitive to FDD, in contrast to beta1-negative cerebellar Purkinje neurons. Immunostaining for the beta1-subunit and the potency of FDD to modulate GABA responses in cultured hypothalamic neurons was drastically diminished by beta1-siRNA treatment. In conclusion, with the help of FDDs we reveal a functional expression of beta1-containing GABA(A)Rs in the hypothalamus, offering a new tool for studies on the functional diversity of native GABA(A)Rs.

The neuronal RhoA GEF, Tech, interacts with the synaptic multi-PDZ-domain-containing protein, MUPP1.

Tech is a RhoA guanine nucleotide exchange factor (GEF) that is highly enriched in hippocampal and cortical neurons. To help define its function, we have conducted studies aimed at identifying partner proteins that bind to its C-terminal PDZ ligand motif. Yeast two hybrid studies using the Tech C-terminal segment as bait identified MUPP1, a protein that contains 13 PDZ domains and has been localized to the post-synaptic compartment, as a candidate partner protein for Tech. Co-transfection of Tech and MUPP1 in human embryonic kidney 293 cells confirmed that these full-length proteins interact in a PDZ-dependent fashion. Furthermore, we confirmed that endogenous Tech co-precipitates with MUPP1, but not PSD-95, from hippocampal and cortical extracts prepared from rat brain. In addition, immunostaining of primary cortical cultures revealed co-localization of MUPP1 and Tech puncta in the vicinity of synapses. In assessing which PDZ domains of MUPP1 mediate binding to Tech, we found that Tech can bind to either PDZ domain 10 or 13 of MUPP1 as mutation of both these domains is needed to disrupt their interaction. Taken together, these findings demonstrate that Tech binds to MUPP1 and suggest that it regulates RhoA signaling pathways in the vicinity of synapses.

Non-motor behavioural impairments in parkin-deficient mice.

Mutations in the parkin gene are the major cause of early-onset familial Parkinson's disease (PD). We previously reported the generation and analysis of a knockout mouse carrying a deletion of exon 3 in the parkin gene. F1 hybrid pa+/- mice were backcrossed to wild-type C57Bl/6 for three more generations to establish a pa-/-(F4) mouse line. The appearance of tyrosine hydroxylase-positive neurons was normal in young and aged pa-/- (F4) animals. Loss of parkin function in mice did not enhance vulnerability of dopaminergic neurons to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) toxicity. However, the pa-/- (F4) mice displayed impaired exploration and habituation to a new environment and exhibited thigmotaxis behaviour in the open field and Morris water maze. Abnormal anxiety-related behaviour of pa-/- (F4) mice was also observed in the light/dark exploration test paradigm. Dopamine metabolism was enhanced in the striatum of pa-/- (F4) mice, as revealed by increased homovanillic acid (HVA) content and a reduced ratio of dihydroxyphenylacetic acid (DOPAC)/HVA. The alterations found in the dopaminergic system could be responsible for the behavioural impairments of pa-/- (F4) mice. Consistent with a recent observation of cognitive dysfunction in parkin-linked patients with PD, our findings provide evidence of a physiological role of parkin in non-motor behaviour, possibly representing a disease stage that precedes dopaminergic neuron loss.