Phosphorylation of neuroligin-2 by PKA regulates its cell surface abundance and synaptic stabilization

The trans-synaptic adhesion molecule neuroligin-2 (NL2) is essential for the development and function of inhibitory synapses. NL2 recruits the postsynaptic scaffold protein gephyrin, which, in turn, stabilizes γ-aminobutyric acid type A receptors (GABAARs) in the postsynaptic domain. Thus, the amount of NL2 at the synapse can control synaptic GABAAR concentration to tune inhibitory neurotransmission efficacy. Here, using biochemistry, imaging, single-particle tracking, and electrophysiology, we uncovered a key role for cAMP-dependent protein kinase (PKA) in the synaptic stabilization of NL2. We found that PKA-mediated phosphorylation of NL2 at Ser714 caused its dispersal from the synapse and reduced NL2 surface amounts, leading to a loss of synaptic GABAARs. Conversely, enhancing the stability of NL2 at synapses by abolishing PKA-mediated phosphorylation led to increased inhibitory signaling. Thus, PKA plays a key role in regulating NL2 function and GABA-mediated synaptic inhibition

Selective inhibition of GluN2D-containing N-methyl-D-aspartate receptors prevents tissue plasminogen activator-promoted neurotoxicity both in vitro and in vivo

BACKGROUND: Tissue plasminogen activator (tPA) exerts multiple functions in the central nervous system, depending on the partner with which it interacts. In particular, tPA acts as a positive neuromodulator of N-methyl-D-aspartate glutamatergic receptors (NMDAR). At the molecular level, it has been proposed that the pro-neurotoxicity mediated by tPA might occur through extrasynaptic NMDAR containing the GluN2D subunit. Thus, selective antagonists targeting tPA/GluN2D-containing NMDAR signaling would be of interest to prevent noxious effects of tPA. RESULTS: Here, we compared three putative antagonists of GluN2D-containing NMDAR and we showed that the new compound UBP145 ((2R*,3S*)-1-(9-bromophenan-threne-3-carbonyl)piperazine-2,3-dicarboxylic acid) is far more selective for GluN2D subunits than memantine and PPDA (phenanthrene derivative (2S*, 3R*)-1-(phenanthrene-2-carbonyl)piperazine-2,3-dicarboxylic acid). Indeed, in vitro, in contrast to the two other compounds, UBP145 prevented NMDA toxicity only in neurons expressing GluN2D (ie, in cortical but not hippocampal neurons). Furthermore, in cultured cortical neurons, UBP145 fully prevented the pro-excitotoxic effect of tPA. In vivo, we showed that UBP145 potently prevented the noxious action of exogenous tPA on excitotoxic damages. Moreover, in a thrombotic stroke model in mice, administration of UBP145 prevented the deleterious effect of late thrombolysis by tPA. CONCLUSIONS: In conclusion, tPA exerts noxious effects on neurons by acting on GluN2D-containing NMDAR and pharmacological antagonists of GluN2D-containing NMDAR could be used to prevent the ability of tPA to promote neurotoxicity

Association between the PPP3CC gene, coding for the calcineurin gamma catalytic subunit, and bipolar disorder

<p>Abstract</p> <p>Background</p> <p>Calcineurin is a neuron-enriched phosphatase that regulates synaptic plasticity and neuronal adaptation. Activation of calcineurin, overall, antagonizes the effects of the cyclic AMP activated protein/kinase A. Thus, kinase/phosphatase dynamic balance seems to be critical for transition to long-term cellular responses in neurons, and disruption of this equilibrium should induce behavioral impairments in animal models. Genetic animal models, as well as post-mortem studies in humans have implicated calcineurin dependent calcium and cyclic AMP regulated phosphorylation/dephosphorylation in both affective responses and psychosis. Recently, genetic association between schizophrenia and genetic variation of the human calcineurin A gamma subunit gene (PPP3CC) has been reported.</p> <p>Methods</p> <p>Based on the assumption of the common underlying genetic factor in schizophrenia and bipolar affective disorder (BPAD), we performed association analysis of CC33 and CCS3 polymorphisms of the PPP3CC gene reported to be associated with schizophrenia in a French sample of 115 BPAD patients and 97 healthy controls.</p> <p>Results</p> <p>Carrying 'CT' or 'TT' genotypes of the PPP3CC-CC33 polymorphism increased risk to develop BPAD comparing to carry 'CC' genotype (OR = 1.8 [1.01–3.0]; p = 0.05). For the PPP3CC-CCS3 polymorphism, 'AG' or 'GG' carriers have an increased risk to develop BPAD than 'AA' carriers (OR = 2.8 [1.5–5.2]). The CC33 and CCS3 polymorphisms were observed in significant linkage disequilibrium (D' = 0.91, r²= 0.72). Haplotype frequencies were significantly different in BPAD patients than in controls (p = 0.03), with a significant over-transmission of the 'TG' haplotype in BPAD patients (p = 0.001).</p> <p><b>Conclusion:</b></p> <p>We suggest that the PPP3CC gene might be a susceptibility gene for BPAD, in accordance with current neurobiological hypotheses that implicate dysregulation of signal-transduction pathways, such as those regulated by calcineurin, in the etiology of affective disorders.</p

Construction and validation of a dimensional scale exploring mood disorders: MAThyS (Multidimensional Assessment of Thymic States)

<p>Abstract</p> <p>Background</p> <p>The boundaries between mood states in bipolar disorders are not clear when they are associated with mixed characteristics. This leads to some confusion to define appropriate therapeutic strategies. A dimensional approach might help to better define bipolar moods states and more specifically those with mixed features.</p> <p>Therefore, we proposed a new tool based on a dimensional approach, built with a priori five sub-scales and focus on emotional reactivity rather than exclusively on mood tonality. This study was designed to validate this MAThyS Scale (Multidimensional Assessment of Thymic States).</p> <p>Methods</p> <p>One hundred and ninety six subjects were included: 44 controls and 152 bipolar patients in various states: euthymic, manic or depressed. The MAThyS is a visual analogic scale consisting of 20 items. These items corresponded to five quantitative dimensions ranging from inhibition to excitation: emotional reactivity, thought processes, psychomotor function, motivation and sensory perception. They were selected as they represent clinically relevant quantitative traits.</p> <p>Results</p> <p>Confirmatory analyses demonstrated a good validity for this scale, and a good internal consistency (Cronbach's alpha coefficient = 0.95). The MathyS scale is moderately correlated of both the MADRS scale (depressive score; r = -0.45) and the MAS scale (manic score; r = 0.56).</p> <p>When considering the Kaiser-Guttman rule and the scree plot, our model of 5 factors seems to be valid. The four first factors have an eigenvalue greater than 1.0 and the eigenvalue of the factor five is 0.97. In the scree plot, the "elbow", or the point at which the curve bends, indicates 5 factors to extract. This 5 factors structure explains 68 per cent of variance.</p> <p>Conclusion</p> <p>The characterisation of bipolar mood states based on a global score assessing inhibition/activation process (total score of the MATHyS) associated with descriptive analysis on sub-scores such as emotional reactivity (rather than the classical opposition euphoria/sadness) can be useful to better understand the broad spectrum of mixed states.</p

Number and mode of inheritance of QTL influencing backfat thickness on SSC2p in Sino-European pig pedigrees

<p>Abstract</p> <p>Background</p> <p>In the pig, multiple QTL associated with growth and fatness traits have been mapped to chromosome 2 (SSC2) and among these, at least one shows paternal expression due to the IGF2-intron3-G3072A substitution. Previously published results on the position and imprinting status of this QTL disagree between analyses from French and Dutch F2 crossbred pig populations obtained with the same breeds (Meishan crossed with Large White or Landrace).</p> <p>Methods</p> <p>To study the role of paternal and maternal alleles at the IGF2 locus and to test the hypothesis of a second QTL affecting backfat thickness on the short arm of SSC2 (SSC2p), a QTL mapping analysis was carried out on a combined pedigree including both the French and Dutch F2 populations, on the progeny of F1 males that were heterozygous (A/G) and homozygous (G/G) at the IGF2 locus. Simulations were performed to clarify the relations between the two QTL and to understand to what extent they can explain the discrepancies previously reported.</p> <p>Results</p> <p>The QTL analyses showed the segregation of at least two QTL on chromosome 2 in both pedigrees, i.e. the IGF2 locus and a second QTL segregating at least in the G/G F1 males and located between positions 30 and 51 cM. Statistical analyses highlighted that the maternally inherited allele at the IGF2 locus had a significant effect but simulation studies showed that this is probably a spurious effect due to the segregation of the second QTL.</p> <p>Conclusions</p> <p>Our results show that two QTL on SSC2p affect backfat thickness. Differences in the pedigree structures and in the number of heterozygous females at the IGF2 locus result in different imprinting statuses in the two pedigrees studied. The spurious effect observed when a maternally allele is present at the IGF2 locus, is in fact due to the presence of a second closely located QTL. This work confirms that pig chromosome 2 is a major region associated with fattening traits.</p

Differential Impact of EGFR-Targeted Therapies on Hypoxia Responses: Implications for Treatment Sensitivity in Triple-Negative Metastatic Breast Cancer

In solid tumors, such as breast cancer, cells are exposed to hypoxia. Cancer cells adapt their metabolism by activating hypoxia-inducible factors (HIFs) that promote the transcription of genes involved in processes such as cell survival, drug resistance and metastasis. HIF-1 is also induced in an oxygen-independent manner through the activation of epidermal growth factor receptor tyrosine kinase (EGFR-TK). Triple-negative breast cancer (TNBC) is a subtype of invasive breast cancer characterized by negative expression of hormonal and HER2 receptors, and this subtype generally overexpresses EGFR. Sensitivity to three EGFR inhibitors (cetuximab, gefitinib and lapatinib, an HER2/EGFR-TK inhibitor) was evaluated in a metastatic TNBC cell model (MDA-MB-231), and the impact of these drugs on the activity and stability of HIF was assessed.MDA-MB-231 cells were genetically modified to stably express an enhanced green fluorescent protein (EGFP) induced by hypoxia; the Ca9-GFP cell model reports HIF activity, whereas GFP-P564 reports HIF stability. The reporter signal was monitored by flow cytometry. HIF-1 DNA-binding activity, cell migration and viability were also evaluated in response to EGFR inhibitors. Cell fluorescence signals strongly increased under hypoxic conditions (> 30-fold). Cetuximab and lapatinib did not affect the signal induced by hypoxia, whereas gefitinib sharply reduced its intensity in both cell models and also diminished HIF-1 alpha levels and HIF-1 DNA-binding activity in MDA-MB-231 cells. This gefitinib feature was associated with its ability to inhibit MDA-MB-231 cell migration and to induce cell mortality in a dose-dependent manner. Cetuximab and lapatinib had no effect on cell migration or cell viability.Resistance to cetuximab and lapatinib and sensitivity to gefitinib were associated with their ability to modulate HIF activity and stability. In conclusion, downregulation of HIF-1 through EGFR signaling seems to be required for the induction of a positive response to EGFR-targeted therapies in TNBC

Local Cattle and Badger Populations Affect the Risk of Confirmed Tuberculosis in British Cattle Herds

Background: The control of bovine tuberculosis (bTB) remains a priority on the public health agenda in Great Britain, after launching in 1998 the Randomised Badger Culling Trial (RBCT) to evaluate the effectiveness of badger (Meles meles) culling as a control strategy. Our study complements previous analyses of the RBCT data (focusing on treatment effects) by presenting analyses of herd-level risks factors associated with the probability of a confirmed bTB breakdown in herds within each treatment: repeated widespread proactive culling, localized reactive culling and no culling (survey-only). Methodology/Principal Findings: New cases of bTB breakdowns were monitored inside the RBCT areas from the end of the first proactive badger cull to one year after the last proactive cull. The risk of a herd bTB breakdown was modeled using logistic regression and proportional hazard models adjusting for local farm-level risk factors. Inside survey-only and reactive areas, increased numbers of active badger setts and cattle herds within 1500 m of a farm were associated with an increased bTB risk. Inside proactive areas, the number of M. bovis positive badgers initially culled within 1500 m of a farm was the strongest predictor of the risk of a confirmed bTB breakdown. Conclusions/Significance: The use of herd-based models provide insights into how local cattle and badger populations affect the bTB breakdown risks of individual cattle herds in the absence of and in the presence of badger culling. These measures of local bTB risks could be integrated into a risk-based herd testing programme to improve the targeting o