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Rho GTPases are a class of G-proteins involved in several aspects of cellular biology, including the regulation of actin cytoskeleton. The most studied members of this family are RHOA and RAC1 that act in concert to regulate actin dynamics. Recently, Rho GTPases gained much attention as synaptic regulators in the mammalian central nervous system (CNS). In this context, ARHGAP22 protein has been previously shown to specifically inhibit RAC1 activity thus standing as critical cytoskeleton regulator in cancer cell models; however, whether this function is maintained in neurons in the CNS is unknown. Here, we generated a knockout animal model for arhgap22 and provided evidence of its role in the hippocampus. Specifically, we found that ARHGAP22 absence leads to RAC1 hyperactivity and to an increase in dendritic spine density with defects in synaptic structure, molecular composition, and plasticity. Furthermore, arhgap22 silencing causes impairment in cognition and a reduction in anxiety-like behavior in mice. We also found that inhibiting RAC1 restored synaptic plasticity in ARHGAP22 KO mice. All together, these results shed light on the specific role of ARHGAP22 in hippocampal excitatory synapse formation and function as well as in learning and memory behaviors. [ABSTRACT FROM AUTHOR]

Fragile X syndrome (FXS) is the most frequent form of inherited intellectual disability and the best-described monogenic cause of autism. CGG-repeat expansion in the FMR1 gene leads to FMR1 silencing, loss-of-expression of the Fragile X Mental Retardation Protein (FMRP), and is a common cause of FXS. Missense mutations in the FMR1 gene were also identified in FXS patients, including the recurrent FMRP-R138Q mutation. To investigate the mechanisms underlying FXS caused by this mutation, we generated a knock-in mouse model (Fmr1R138Q) expressing the FMRP-R138Q protein. We demonstrate that, in the hippocampus of the Fmr1R138Q mice, neurons show an increased spine density associated with synaptic ultrastructural defects and increased AMPA receptor-surface expression. Combining biochemical assays, high-resolution imaging, electrophysiological recordings, and behavioural testing, we also show that the R138Q mutation results in impaired hippocampal long-term potentiation and socio-cognitive deficits in mice. These findings reveal the functional impact of the FMRP-R138Q mutation in a mouse model of FXS. The R138Q mutation in the Fragile X Mental Retardation 1 (FMR1) gene has been associated with Fragile X syndrome (FXS). Here, the authors present a Fmr1R138Q Knock-In mouse model and show that R138Q mutation results in impaired long-term potentiation and socio-cognitive performance in these mice. [ABSTRACT FROM AUTHOR]

Correlative light electron microscopy (CLEM) combines the advantages of light and electron microscopy, thus making it possible to follow dynamic events in living cells at nanometre resolution. Various CLEM approaches and devices have been developed, each of which has its own advantages and technical challenges. We here describe our customized patterned glass substrates, which improve the feasibility of correlative fluorescence/confocal and scanning electron microscopy. [ABSTRACT FROM AUTHOR]

We have recently shown that oxytocin inhibits cell proliferation when the vast majority of oxytocin receptors are excluded from caveolin-1-enriched microdomains, and that, on the contrary, it has a mitogenic effect when the receptors are targeted to these plasma membrane domains. In this study, we investigated whether the receptors located inside and outside caveolar microdomains initiate different signalling pathways and how this may lead to opposite effects on cell proliferation. Our data indicate that, depending on their localization, oxytocin receptors transactivate EGFR and activate ERK1/2 using different signalling intermediates. The final outcome is a different temporal pattern of EGFR and ERK1/2 phosphorylation, which is more persistent when the receptors are located outside caveolar microdomains and inhibit cell growth, and very transient when they are located in caveolar microdomains and stimulate cell growth. Finally, only the activation of receptors located outside caveolar microdomains correlates with the activation of the cell cycle inhibitor p21WAF1/CIP1, thus suggesting that the antiproliferative OTR effects may, in this case, be achieved by a sustained activation of EGFR and MAPK leading to the induction of this cell cycle regulator.Oncogene (2003) 22, 6054-6060. doi:10.1038/sj.onc.1206612 [ABSTRACT FROM AUTHOR]

Presents a study that investigated the functional role of the localization of human oxytocin receptor in caveolin-1 enriched membrane domains. Functions of caveolae; List of signaling molecules; Impact of G protein-coupled receptors (GPRC) on caveolae; Analysis of mitogenic effects mediated by some GPRC.

Penetration of the gut mucosa by pathogens expressing invasion genes is believed to occur mainly through specialized epithelial cells, called M cells, that are located in Peyer's patches. However, Salmonella typhimurium that are deficient in invasion genes encoded by Salmonella pathogenicity island 1 (SPI1) are still able to reach the spleen after oral administration. This suggests the existence of an alternative route for bacterial invasion, one that is independent of M cells. We report here a new mechanism for bacterial uptake in the mucosa tissues that is mediated by dendritic cells (DCs). DCs open the tight junctions between epithelial cells, send dendrites outside the epithelium and directly sample bacteria. In addition, because DCs express tight-junction proteins such as occludin, claudin 1 and zonula occludens 1, the integrity of the epithelial barrier is preserved. [ABSTRACT FROM AUTHOR]

The effects of hexarelin, a growth hormone (GH) secretagogue, and human GH on the mechanical and metabolic changes measured in isolated rat hearts submitted to 5 min of Ca deprivation followed by reperfusion with Ca-containing medium, the so-called calcium paradox phenomenon, were studied. Hexarelin (80 μg/kg bid, subcutaneously) administered for 7 d to male rats effectively antagonized the sudden increase in resting tension measured in vitro on Ca repletion. Moreover, during Ca repletion the release of creatine kinase activity (an index of cell damage) in the perfusate of these hearts was reduced up to 40% compared with controls. By contrast, administration of hexarelin for 3 d or GH (400 μg/kg bid, subcutaneously) for 7 d did not affect the mechanical and metabolic alterations induced by the calcium paradox. To assess its direct and acute cardiac effects, hexarelin (8 μg/mL) was perfused in vitro in recirculating conditions for 60 min through the hearts of normal rats. In this case, hexarelin did not stimulate heart contractility and failed to prevent ventricular contracture upon Ca readmission, whereas diltiazem, a Ca channel blocker, effectively antagonized the calcium paradox phenomenon. We conclude that short-term in vivo exposure to hexarelin, but not GH, enables cardiac myocyites to prevent cytoplasmatic electrolytic unbalance and to control intracellular Ca gain, two functions largely impaired during the calcium paradox phenomenon. Moreover, because the effect of hexarelin is not acute but dependent on the length of in vivo treatment, we suggest that it requires modifications of myocardiocyte physiology. [ABSTRACT FROM AUTHOR]

The N-type voltage-operated calcium channel has been characterized over the years as a high-threshold channel, with variable inactivation kinetics, and a unique ability to bind with high affinity and specificity ω-conotoxin GVIA and related toxins. This channel is particularly expressed in some neurons and endocrine cells, where it participates in several calcium-dependent processes, including secretion. ω-conotoxin GVIA was instrumental not only for the biophysical and pharmacological characterization of N-type channels but also for the development of in vitro assays for studying N-type VOCC subcellular localization, biosynthesis, turnover, as well as short-and long-term regulation of its expression. We here summarize our studies on N-type VOCC expression in neurosecretory cells, with a major emphasis on recent data demonstrating the presence of N-type channels in intracellular secretory organelles and their recruitment to the cell surface during regulated exocytosis. [ABSTRACT FROM AUTHOR]

Shrm4, a protein expressed only in polarized tissues, is encoded by the KIAA1202 gene, whose mutations have been linked to epilepsy and intellectual disability. However, a physiological role for Shrm4 in the brain is yet to be established. Here, we report that Shrm4 is localized to synapses where it regulates dendritic spine morphology and interacts with the C terminus of GABAB receptors (GABABRs) to control their cell surface expression and intracellular trafficking via a dynein-dependent mechanism. Knockdown of Shrm4 in rat severely impairs GABABR activity causing increased anxiety-like behaviour and susceptibility to seizures. Moreover, Shrm4 influences hippocampal excitability by modulating tonic inhibition in dentate gyrus granule cells, in a process involving crosstalk between GABABRs and extrasynaptic δ-subunit-containing GABAARs. Our data highlights a role for Shrm4 in synaptogenesis and in maintaining GABABR-mediated inhibition, perturbation of which may be responsible for the involvement of Shrm4 in cognitive disorders and epilepsy. [ABSTRACT FROM AUTHOR]