Dynamic visual information plays a critical role for spatial navigation in water but not on solid ground.

In the Morris water maze (MWM) task, proprioceptive information is likely to have a poor accuracy due to movement inertia. Hence, in this condition, dynamic visual information providing information on linear and angular acceleration would play a critical role in spatial navigation. To investigate this assumption we compared rat's spatial performance in the MWM and in the homing hole board (HB) tasks using a 1.5 Hz stroboscopic illumination. In the MWM, rats trained in the stroboscopic condition needed more time than those trained in a continuous light condition to reach the hidden platform. They expressed also little accuracy during the probe trial. In the HB task, in contrast, place learning remained unaffected by the stroboscopic light condition. The deficit in the MWM was thus complete, affecting both escape latency and discrimination of the reinforced area, and was thus task specific. This dissociation confirms that dynamic visual information is crucial to spatial navigation in the MWM whereas spatial navigation on solid ground is mediated by a multisensory integration, and thus less dependent on visual information.

Renal tubular NEDD4-2 deficiency causes NCC-mediated salt-dependent hypertension.

The E3 ubiquitin ligase NEDD4-2 (encoded by the Nedd4L gene) regulates the amiloride-sensitive epithelial Na+ channel (ENaC/SCNN1) to mediate Na+ homeostasis. Mutations in the human β/γENaC subunits that block NEDD4-2 binding or constitutive ablation of exons 6-8 of Nedd4L in mice both result in salt-sensitive hypertension and elevated ENaC activity (Liddle syndrome). To determine the role of renal tubular NEDD4-2 in adult mice, we generated tetracycline-inducible, nephron-specific Nedd4L KO mice. Under standard and high-Na+ diets, conditional KO mice displayed decreased plasma aldosterone but normal Na+/K+ balance. Under a high-Na+ diet, KO mice exhibited hypercalciuria and increased blood pressure, which were reversed by thiazide treatment. Protein expression of βENaC, γENaC, the renal outer medullary K+ channel (ROMK), and total and phosphorylated thiazide-sensitive Na+Cl- cotransporter (NCC) levels were increased in KO kidneys. Unexpectedly, Scnn1a mRNA, which encodes the αENaC subunit, was reduced and proteolytic cleavage of αENaC decreased. Taken together, these results demonstrate that loss of NEDD4-2 in adult renal tubules causes a new form of mild, salt-sensitive hypertension without hyperkalemia that is characterized by upregulation of NCC, elevation of β/γENaC, but not αENaC, and a normal Na+/K+ balance maintained by downregulation of ENaC activity and upregulation of ROMK.

Monitoring time-dependent degradation of phospholipids in sectioned tissues by MALDI imaging mass spectrometry.

Imaging mass spectrometry (IMS) is useful for visualizing the localization of phospholipids on biological tissue surfaces creating great opportunities for IMS in lipidomic investigations. With advancements in IMS of lipids, there is a demand for large-scale tissue studies necessitating stable, efficient and well-defined sample handling procedures. Our work within this article shows the effects of different storage conditions on the phospholipid composition of sectioned tissues from mouse organs. We have taken serial sections from mouse brain, kidney and liver thaw mounted unto ITO-coated glass slides and stored them under various conditions later analyzing them at fixed time points. A global decrease in phospholipid signal intensity is shown to occur and to be a function of time and temperature. Contrary to the global decrease, oxidized phospholipid and lysophospholipid species are found to increase within 2 h and 24 h, respectively, when mounted sections are kept at ambient room conditions. Imaging experiments reveal that degradation products increase globally across the tissue. Degradation is shown to be inhibited by cold temperatures, with sample integrity maintained up to a week after storage in −80 °C freezer under N2 atmosphere. Overall, the results demonstrate a timeline of the effects of lipid degradation specific to sectioned tissues and provide several lipid species which can serve as markers of degradation. Importantly, the timeline demonstrates oxidative sample degradation begins appearing within the normal timescale of IMS sample preparation of lipids (i.e. 1-2 h) and that long-term degradation is global. Taken together, these results strengthen the notion that standardized procedures are required for phospholipid IMS of large sample sets, or in studies where many serial sections are prepared together but analyzed over time such as in 3-D IMS reconstruction experiments.

Age-dependent gain of alternative splice forms and biased duplication explain the relation between splicing and duplication.

We analyze here the relation between alternative splicing and gene duplication in light of recent genomic data, with a focus on the human genome. We show that the previously reported negative correlation between level of alternative splicing and family size no longer holds true. We clarify this pattern and show that it is sufficiently explained by two factors. First, genes progressively gain new splice variants with time. The gain is consistent with a selectively relaxed regime, until purifying selection slows it down as aging genes accumulate a large number of variants. Second, we show that duplication does not lead to a loss of splice forms, but rather that genes with low levels of alternative splicing tend to duplicate more frequently. This leads us to reconsider the role of alternative splicing in duplicate retention.

Intestinal bacteria condition dendritic cells to promote IgA production.

Immunoglobulin (Ig) A represents the predominant antibody isotype produced at the intestinal mucosa, where it plays an important role in limiting the penetration of commensal intestinal bacteria and opportunistic pathogens. We show in mice that Peyer's Patch-derived dendritic cells (PP-DC) exhibit a specialized phenotype allowing the promotion of IgA production by B2 cells. This phenotype included increased expression of the retinaldehyde dehydrogenase 1 (RALDH1), inducible nitric oxide synthase (iNOS), B cell activating factor of the tumor necrosis family (BAFF), a proliferation-inducing ligand (APRIL), and receptors for the neuropeptide vasoactive intestinal peptide (VIP). The ability of PP-DC to promote anti-CD40 dependent IgA was partially dependent on retinoic acid (RA) and transforming growth factor (TGF)-beta, whilst BAFF and APRIL signaling were not required. Signals delivered by BAFF and APRIL were crucial for CD40 independent IgA production, although the contribution of B2 cells to this pathway was minimal. The unique ability of PP-DC to instruct naïve B cells to differentiate into IgA producing plasma cells was mainly imparted by the presence of intestinal commensal bacteria, and could be mimicked by the addition of LPS to the culture. These data indicate that exposure to pathogen-associated molecular patterns present on intestinal commensal bacteria condition DC to express a unique molecular footprint that in turn allows them to promote IgA production.

Stage of change of cigarette smoking in drug dependent patients

Résumé En Suisse, les programmes de désaccoutumance au tabac se réfèrent généralement au modèle de préparation au changement de Prochaska et DiClemente (1983), Les patients atteints de maladies somatiques liées au tabagisme comme les pathologies cardiovasculaires ou pulmonaires accèdent facilement à ces programmes, contrairement aux patients présentant une dépendance à des drogues illicites. La prévalence de fumeurs dans cette population est pourtant élevée et les problèmes engendrés par le tabac sont importants, non seulement d'un point de vue individuel mais aussi en terme de santé publique. Il est par conséquent intéressant d'évaluer la motivation concernant la désaccoutumance au tabac de patients toxicomanes entreprenant un sevrage de drogues illicites. Dans cette étude, nous avons évalué les stades de préparation au changement concernant la dépendance au tabac chez 100 patients toxicomanes hospitalisés sur un mode volontaire dans le cadre d'un programme de sevrage à des drogues illégales. L'évaluation s'est faite à l'aide d'un auto-questionnaire dont les résultats indiquent qu'une minorité de patients sont décidés à interrompre la consommation de tabac. En effet, seul 15% des patients se trouvaient aux stades de contemplation ou de décision. De plus, 93% des sujets considéraient l'arrêt du tabac comme difficile ou très difficile. Ces données montrent qu'il existe un décalage important entre la motivation relative au sevrage de drogues illégales et la motivation liées à l'arrêt du tabac. En effet, malgré leur motivation élevée pour se sevrer de drogues illicites, la proportion de patients restant au stade de précontemplation concernant la désaccoutumance au tabac reste élevée. Diverses hypothèses permettent d'expliquer ces résultats, notamment la perception que la désaccoutumance au tabac est plus difficile à réaliser que le sevrage de substances illicites. Abstract Nicotine cessation programmes in Switzerland, which are commonly based on the stage of change model of Prochaska and DiClemente (1983), are rarely offered to patients with illicit drug dependence. This stands in contrast to the high smoking rates and the heavy burden of tobacco-related problems in these patients. The stage of change was therefore assessed by self-administered questionnaire in 100 inpatients attending an illegal drug withdrawal programme. Only 15% of the patients were in the contemplation or decision stage. 93% considered smoking cessation to be difficult or very difficult. These data show a discrepancy between the motivation to change illegal drug consumption habits and the motivation for smoking cessation. The high pro-portion of patients remaining in the precontemplation stage for smoking cessation, in spite of their motivation for illicit drug detoxification, may be due to the perception that cessation of smoking is more difficult than illicit drug abuse cessation.

Environment-dependent microevolution in a Mediterranean pine (Pinus pinaster Aiton).

BACKGROUND: A central question for understanding the evolutionary responses of plant species to rapidly changing environments is the assessment of their potential for short-term (in one or a few generations) genetic change. In our study, we consider the case of Pinus pinaster Aiton (maritime pine), a widespread Mediterranean tree, and (i) test, under different experimental conditions (growth chamber and semi-natural), whether higher recruitment in the wild from the most successful mothers is due to better performance of their offspring; and (ii) evaluate genetic change in quantitative traits across generations at two different life stages (mature trees and seedlings) that are known to be under strong selection pressure in forest trees. RESULTS: Genetic control was high for most traits (h2 = 0.137-0.876) under the milder conditions of the growth chamber, but only for ontogenetic change (0.276), total height (0.415) and survival (0.719) under the more stressful semi-natural conditions. Significant phenotypic selection gradients were found in mature trees for traits related to seed quality (germination rate and number of empty seeds). Moreover, female relative reproductive success was significantly correlated with offspring performance for specific leaf area (SLA) in the growth chamber experiment, and stem mass fraction (SMF) in the experiment under semi-natural conditions, two adaptive traits related to abiotic stress-response in pines. Selection gradients based on genetic covariance of seedling traits and responses to selection at this stage involved traits related to biomass allocation (SMF) and growth (as decomposed by a Gompertz model) or delayed ontogenetic change, depending also on the testing environment. CONCLUSIONS: Despite the evidence of microevolutionary change in adaptive traits in maritime pine, directional or disruptive changes are difficult to predict due to variable selection at different life stages and environments. At mature-tree stages, higher female effective reproductive success can be explained by differences in their production of offspring (due to seed quality) and, to a lesser extent, by seemingly better adapted seedlings. Selection gradients and responses to selection for seedlings also differed across experimental conditions. The distinct processes involved at the two life stages (mature trees or seedlings) together with environment-specific responses advice caution when predicting likely evolutionary responses to environmental change in Mediterranean forest trees.

Maturation-dependent neurotoxicity of lead acetate in vitro: implication of glial reactions.

Despite a wealth of data on the neurotoxic effects of lead at the cellular and molecular levels, the reasons for its development-dependent neurotoxicity are still unclear. Here, the maturation-dependent effects of lead acetate were analyzed in immature and differentiated brain cells cultured in aggregates. Markers of general cytotoxicity as well as cell-type-specific markers of glial and neuronal cells showed that immature brain cells were more sensitive to lead than the differentiated counterparts, demonstrating that the development-dependent neurotoxicity of lead can be reproduced in aggregating brain cell cultures. After 10 days of treatment, astrocytes were found to be more affected by lead acetate than neurons in immature cultures, and microglial cells were strongly activated. Eleven days after cessation of the treatment, lead acetate caused a partial loss of astrocytes and an intense reactivity of the remaining ones. Furthermore, microglial cells expressed a macrophagic phenotype, and the loss of activity of neuron-specific enzymes was aggravated. In differentiated cultures, no reactive gliosis was found. It is hypothetized that the intense glial reactions (microgliosis and astrogliosis) observed in immature cultures contribute to the development-dependent neurotoxicity of lead.

CD8 beta increases CD8 coreceptor function and participation in TCR-ligand binding.

To study the role of CD8 beta in T cell function, we derived a CD8 alpha/beta-(CD8-/-) T cell hybridoma of the H-2Kd-restricted N9 cytotoxic T lymphocyte clone specific for a photoreactive derivative of the Plasmodium berghei circumsporozoite peptide PbCS 252-260. This hybridoma was transfected either with CD8 alpha alone or together with CD8 beta. All three hybridomas released interleukin 2 upon incubation with L cells expressing Kd-peptide derivative complexes, though CD8 alpha/beta cells did so more efficiently than CD8 alpha/alpha and especially CD8-/- cells. More strikingly, only CD8 alpha/beta cells were able to recognize a weak agonist peptide derivative variant. This recognition was abolished by Fab' fragments of the anti-Kd alpha 3 monoclonal antibody SF1-1.1.1 or substitution of Kd D-227 with K, both conditions known to impair CD8 coreceptor function. T cell receptor (TCR) photoaffinity labeling indicated that TCR-ligand binding on CD8 alpha/beta cells was approximately 5- and 20-fold more avid than on CD8 alpha/a and CD8-/- cells, respectively. SF1-1.1.1 Fab' or Kd mutation D227K reduced the TCR photoaffinity labeling on CD8 alpha/beta cells to approximately the same low levels observed on CD8-/- cells. These results indicate that CD8 alpha/beta is a more efficient coreceptor than CD8alpha/alpha, because it more avidly strengthens TCR-ligand binding.

Differential roles of T cell receptor alpha and beta chains in ligand binding among H-2Kd-restricted cytolytic T lymphocyte clones specific for a photoreactive Plasmodium berghei circumsporozoite peptide derivative.

To study the interaction of T cell receptor with its ligand, a complex of a major histocompatibility complex molecule and a peptide, we derived H-2Kd-restricted cytolytic T lymphocyte clones from mice immunized with a Plasmodium berghei circumsporozoite peptide (PbCS) 252-260 (SYIPSAEKI) derivative containing photoreactive Nepsilon-[4-azidobenzoyl] lysine in place of Pro-255. This residue and Lys-259 were essential parts of the epitope recognized by these clones. Most of the clones expressed BV1S1A1 encoded beta chains along with specific complementary determining region (CDR) 3beta regions but diverse alpha chain sequences. Surprisingly, all T cell receptors were preferentially photoaffinity labeled on the alpha chain. For a representative T cell receptor, the photoaffinity labeled site was located in the Valpha C-strand. Computer modeling suggested the presence of a hydrophobic pocket, which is formed by parts of the Valpha/Jalpha C-, F-, and G-strands and adjacent CDR3alpha residues and structured to be able to avidly bind the photoreactive ligand side chain. We previously found that a T cell receptor specific for a PbCS peptide derivative containing this photoreactive side chain in position 259 similarly used a hydrophobic pocket located between the junctional CDR3 loops. We propose that this nonpolar domain in these locations allow T cell receptors to avidly and specifically bind epitopes containing non-peptidic side chains.

SOCS-1 protein prevents Janus Kinase/STAT-dependent inhibition of beta cell insulin gene transcription and secretion in response to interferon-gamma.

In the pathogenesis of type I diabetes mellitus, activated leukocytes infiltrate pancreatic islets and induce beta cell dysfunction and destruction. Interferon (IFN)-gamma, tumor necrosis factor-alpha and interleukin (IL)-1 beta play important, although not completely defined, roles in these mechanisms. Here, using the highly differentiated beta Tc-Tet insulin-secreting cell line, we showed that IFN-gamma dose- and time-dependently suppressed insulin synthesis and glucose-stimulated secretion. As described previously IFN-gamma, in combination with IL-1 beta, also induces inducible NO synthase expression and apoptosis (Dupraz, P., Cottet, S., Hamburger, F., Dolci, W., Felley-Bosco, E., and Thorens, B. (2000) J. Biol. Chem. 275, 37672--37678). To assess the role of the Janus kinase/signal transducer and activator of transcription (STAT) pathway in IFN-gamma intracellular signaling, we stably overexpressed SOCS-1 (suppressor of cytokine signaling-1) in the beta cell line. We demonstrated that SOCS-1 suppressed cytokine-induced STAT-1 phosphorylation and increased cellular accumulation. This was accompanied by a suppression of the effect of IFN-gamma on: (i) reduction in insulin promoter-luciferase reporter gene transcription, (ii) decrease in insulin mRNA and peptide content, and (iii) suppression of glucose-stimulated insulin secretion. Furthermore, SOCS-1 also suppressed the cellular effects that require the combined presence of IL-1 beta and IFN-gamma: induction of nitric oxide production and apoptosis. Together our data demonstrate that IFN-gamma is responsible for the cytokine-induced defect in insulin gene expression and secretion and that this effect can be completely blocked by constitutive inhibition of the Janus kinase/STAT pathway.

Dissociation between vasodilation and Leishmania infection-enhancing effects of sand fly saliva and maxadilan

In this study, the ability of maxadilan and Lutzomyia longipalpis salivary gland lysate to enhance the infection of CBA mice by Leishmania major and of BALB/c mice by L. braziliensis was tested. No difference was observed between sizes of lesion in CBA mice infected with L. major and treated or not with salivary gland lysate or maxadilan, although they were injected in concentrations that induced cutaneous vasodilation. Although parasites were more frequently observed in foot pads and spleens of animals treated with maxadilan than in the animals treated with salivary gland lysate or saline, the differences were small and not statistically significant. The lesions in BALB/c mice infected with L. braziliensis and treated with maxadilan were slightly larger than in animals that received Leishmania alone. Such differences disappeared 14 weeks after infection, and were statistically significant only in one of two experiments.

Preliminary results on the effects of CD40/CD40L interactions and SAC-induction on IFN-gamma expression in human schistosomiasis

In this communication the authors analyzed the pattern of expression of IFN-gamma as a surrogate type 1 response in different clinical forms of schistosomiasis in response to stimulation involving T-cell dependent and T-cell independent pathways, to investigate which pathways were functional in human schistosomiasis, and to further characterize the nature of Th1 response impairment in this parasitic disease.