Motivational Modulation of Self-Initiated and Externally Triggered Movement Speed Induced by Threat of Shock: Experimental Evidence for Paradoxical Kinesis in Parkinson’s Disease

Background

Paradoxical kinesis has been observed in bradykinetic people with Parkinson’s disease. Paradoxical kinesis occurs in situations where an individual is strongly motivated or influenced by relevant external cues. Our aim was to induce paradoxical kinesis in the laboratory. We tested whether the motivation of avoiding a mild electric shock was sufficient to induce paradoxical kinesis in externally-triggered and self-initiated conditions in people with Parkinson’s disease tested on medication and in age-matched controls.

Methods

Participants completed a shock avoidance behavioural paradigm in which half of the trials could result in a mild electric shock if the participant did not move fast enough. Half of the trials of each type were self-initiated and half were externally-triggered. The criterion for avoiding shock was a maximum movement time, adjusted according to each participant’s performance on previous trials using a staircase tracking procedure.

Results

On trials with threat of shock, both patients with Parkinson’s disease and controls had faster movement times compared to no potential shock trials, in both self-initiated and externally-triggered conditions. The magnitude of improvement of movement time from no potential shock to potential shock trials was positively correlated with anxiety ratings.

Conclusions

When motivated to avoid mild electric shock, patients with Parkinson’s disease, similar to healthy controls, showed significant speeding of movement execution. This was observed in both self-initiated and externally-triggered versions of the task. Nevertheless, in the ET condition the improvement of reaction times induced by motivation to avoid shocks was greater for the PD patients than controls, highlighting the value of external cues for movement initiation in PD patients. The magnitude of improvement from the no potential shock to the potential shock trials was associated with the threat-induced anxiety. This demonstration of paradoxical kinesis in the laboratory under both self-initiated and externally-triggered conditions has implications for motivational and attentional enhancement of movement speed in Parkinson’s disease.     

The role of the coypu (Myocastor coypus), an invasive aquatic rodent species,in the epidemiological cycle of leptospirosis: a study in two wetlands in the East of France

Leptospirosis is caused by pathogenic species of the Leptospira genus. Animals can have two roles in the epidemiological cycle: they can be an accidental host and suffer of the disease or a reservoir host which does not express any clinical sign and shed bacteria in their urine. Some of the most known reservoirs for leptospirosis are certain rodent species, but the situation is less clear for aquatic rodents, especially for coypu (Myocastor coypus). It has been shown that this species can have kidney carriage for leptospirosis, but the relationship between carriage and individuals or population health has not been investigated yet. We trapped 133 coypus in two wetlands in the East of France during 3 years. For each animal, a complete necropsy, leptospirosis serology, and a specific real-time quantitative PCR (qPCR) for pathogenic leptospires were performed; in addition, for some animals, a specific kidney culture for leptospires and histology on kidney were performed. In spite of a high seroprevalence (respectively 76 % and 64 %) and of a significant prevalence of kidney carriage in both areas (respectively 12.1 % and 8.0 % of positive qPCR on kidney), the trapped animals seemed in good health, and the population did not seem to be affected by the circulation of the bacteria. These findings are concurring arguments to consider coypu as a real reservoir for leptospirosis.     

Migrant birds disperse haemosporidian parasites and affect their transmission in avian communities

Migration has an important impact on the transmission of pathogens. Migratory birds disperse parasites through their routes and may consequently introduce them to new areas and hosts. Hence, haemosporidian parasites, which are among the most prevalent, diverse and important bird pathogens, are potentially dispersed when infecting migrant hosts. Further, migrant hosts could enhance local parasite prevalence and richness by transporting new parasite strains to new areas. Here, we hypothesize and aim to evaluate if 1) migratory birds spread parasite lineages along their routes, and 2) localities crossed by more migratory birds have greater prevalence and richness of haemosporidians. For the first hypothesis, we tested whether parasite lineages found 1) in both migrants and residents, and 2) only in residents, differ in their frequencies of occurrence among localities. For the second hypothesis, we tested for a relationship among localities between the overall local haemosporidian parasite richness and prevalence, and the proportion of migratory bird individuals present in a locality. We combined a dataset on 13 200 bird samples with additional data from the MalAvi database (total: ~2800 sequenced parasites comprising 675 distinct lineages, from 506 host species and 156 localities) from South America, and used Bayesian multi-level models to test our hypotheses. We demonstrate that parasites shared between resident and migratory species are the most spatially widespread, highlighting the potential of migrants to carry and transmit haemosporidians. Further, the presence of migrants in a locality was negatively related to local parasite richness, but not associated with local prevalence. Here, we confirm that migrants can contribute to parasite dispersal and visiting migrants are present in regions with lower Plasmodium prevalence. Also, we observed their presence might raise Haemoproteus community prevalence. Therefore, we demonstrate migrants enhance pathogens spread and their presence may influence parasite community transmission.     

Identification of a Probable Pore-Forming Domain in the Multimeric Vacuolar Anion Channel AtALMT9

Aluminum-activated malate transporters (ALMTs) form an important family of anion channels involved in fundamental physiological processes in plants. Because of their importance, the role of ALMTs in plant physiology is studied extensively. In contrast, the structural basis of their functional properties is largely unknown. This lack of information limits the understanding of the functional and physiological differences between ALMTs and their impact on anion transport in plants. This study aimed at investigating the structural organization of the transmembrane domain of the Arabidopsis (Arabidopsis thaliana) vacuolar channel AtALMT9. For that purpose, we performed a large-scale mutagenesis analysis and found two residues that form a salt bridge between the first and second putative transmembrane α-helices (TMα1 and TMα2). Furthermore, using a combination of pharmacological and mutagenesis approaches, we identified citrate as an “open channel blocker” of AtALMT9 and used this tool to examine the inhibition sensitivity of different point mutants of highly conserved amino acid residues. By this means, we found a stretch within the cytosolic moiety of the TMα5 that is a probable pore-forming domain. Moreover, using a citrate-insensitive AtALMT9 mutant and biochemical approaches, we could demonstrate that AtALMT9 forms a multimeric complex that is supposedly composed of four subunits. In summary, our data provide, to our knowledge, the first evidence about the structural organization of an ion channel of the ALMT family. We suggest that AtALMT9 is a tetramer and that the TMα5 domains of the subunits contribute to form the pore of this anion channel.The transport of ions across cellular membranes is mediated by specialized proteins that catalyze the transfer of charged molecules across hydrophobic lipid bilayers. Based on the thermodynamics, two major classes of transport systems can be distinguished: (1) passive transporters such as ion channels, which catalyze the flux of solutes down the electrochemical gradient, and (2) active transporters like pumps and antiporters, which transport molecules against their electrochemical gradient. Independent of the nature of the transport system, the flux of ions across membranes is crucial for a wide range of physiological functions in plants. Among others, ion transport is involved in intracellular pH regulation, metal tolerance, stomatal movement, cellular signaling, plant nutrition, and cell expansion (Roelfsema and Hedrich, 2005; Kim et al., 2010; Barbier-Brygoo et al., 2011). Despite the importance of anion transport in plant physiology, only in the last decade has the molecular identity of anion transport proteins started to be unveiled by identifying the chloride channel (CLC), slow anion channel (SLAC), and aluminum-activated malate transporter (ALMT) families. Their discovery has been a fundamental breakthrough in understanding the molecular mechanisms of anion homeostasis and its roles in various aspects of plant cell physiology (Ward et al., 2009; Barbier-Brygoo et al., 2011; Hedrich, 2012; Martinoia et al., 2012).The CLC family consists of both anion channels and secondary active transporters, which are ubiquitously expressed in all living organisms. In Arabidopsis (Arabidopsis thaliana), the first identified and characterized member of the family was AtCLCa (Hechenberger et al., 1996; Geelen et al., 2000). AtCLCa is targeted to the tonoplast and acts as a 2NO₃⁻/H⁺ antiporter (De Angeli et al., 2006). In planta, AtCLCa represents a major vacuolar nitrate transporter driving the accumulation of this anion into the vacuole. Subsequent studies revealed that all other Arabidopsis CLCs are likewise localized in intracellular membranes but feature different cellular functions (Barbier-Brygoo et al., 2011).The SLAC protein family was identified in the last decade (Negi et al., 2008; Vahisalu et al., 2008). Despite its recent discovery, the characterization of this plant anion transporter family proceeded rapidly (Negi et al., 2008; Vahisalu et al., 2008; Geiger et al., 2009, 2010; Brandt et al., 2012). SLAC1, the first identified member of the family, is involved in slow-type anion currents across the plasma membrane of plant cells (Negi et al., 2008; Vahisalu et al., 2008). This ion channel is expressed in guard cells, where it mediates the efflux of anions into the apoplast, a process that is fundamental for stomata closure. SLAC1 regulates the stomatal aperture in response to different stimuli such as abscisic acid and high CO₂ and ozone concentrations (Negi et al., 2008; Vahisalu et al., 2008). In addition, the activity of SLAC1 is controlled by different kinases (Geiger et al., 2009, 2010) that are part of various signaling pathways. This multiple regulation of SLAC1 suggests that it plays a critical role in the integration of different environmental stimuli.ALMTs are membrane proteins exclusive to plants. In Arabidopsis, this family consists of 14 members that can be grouped into three clades (Kovermann et al., 2007). The first member of the ALMT family, TaALMT1, was identified in wheat (Triticum aestivum) by Sasaki et al. (2004) when screening for genes associated with aluminum resistance. They provided evidence that TaALMT1 as well as AtALMT1, its homolog in Arabidopsis, are channels that catalyze the efflux of malate across the plasma membrane of root cells (Yamaguchi et al., 2005; Hoekenga et al., 2006). This exudation of organic acids into the soil facilitates the detoxification of environmental Al³⁺. Besides contributing to Al³⁺ tolerance, ALMTs have been found to exhibit other important physiological functions. AtALMT12 has been proposed to mediate rapid anion currents across the plasma membrane of guard cells in order to induce stomata closure (Meyer et al., 2010). AtALMT9 and AtALMT6 have been shown to be channels localized in the tonoplast that mediate the export of malate into the vacuole (Kovermann et al., 2007; Meyer et al., 2011). AtALMT6 is predominantly expressed in guard cells, where its activity is regulated by cytosolic Ca²⁺ and vacuolar pH (Meyer et al., 2011). In contrast, AtALMT9 is widely expressed in several plant tissues, such as the mesophyll and guard cells. Recently, AtALMT9 was shown to play a crucial role in stomata movement, where it functions as a malate-activated chloride channel (De Angeli et al., 2013).The knowledge about ion channel structures has expanded considerably in the last 20 years. Notably, various three-dimensional structures of such proteins have been solved (Choe, 2002; Jentsch, 2008; Traynelis et al., 2010). This has boosted the research into and the understanding of structure-function relations in transport systems. Among the anion channel families described above, the structure has been determined for CLCs (Dutzler et al., 2002) and SLACs (Chen et al., 2010). Additionally, large structure-function analyses have been conducted, providing detailed knowledge on the molecular basis underlying the ion channel functionality of these families. In contrast, little information was revealed about the structure of ALMTs by describing an important phosphorylation site (Ligaba et al., 2009; Furuichi et al., 2010) and by providing data on the topology (Motoda et al., 2007; Dreyer et al., 2012). However, the proposed models in these studies do not entirely coincide regarding the number of transmembrane-spanning domains, the cellular orientation of the N terminus, and the organization of the C-terminal domain. Therefore, the structural organization of ALMTs is still ambiguous.In this study, we performed a large-scale mutagenesis analysis of the transmembrane domain (TMD) of Arabidopsis ALMTs using the vacuolar channel AtALMT9 as a model. The aim was to identify regions of the TMD that potentially exhibit functional relevance by forming the pore or the voltage sensor. For that purpose, we took advantage of citrate, which we identified as an open channel blocker of AtALMT9. The use of this blocker allowed elucidation of the structural details of ion channels, such as the quaternary organization and pore-forming domains, when no crystal structure was available (MacKinnon, 1991; Yellen et al., 1991; Ferrer-Montiel and Montal, 1996; Linsdell, 2005). By this means, it is possible to show, for instance, that potassium channels are tetramers and to identify their “selectivity filter” domain (MacKinnon and Yellen, 1990; MacKinnon, 1991). Thus, by using citrate, we pharmacologically investigated structure-function relations in AtALMT9. We identified a region adjacent to and within the fifth putative TMD that is supposedly involved in forming the permeation pathway of AtALMT9. Moreover, we demonstrated that AtALMT9 is a multimeric channel of probably four subunits in which the monomers participate in forming the pore.     

Vital role for Plasmodium berghei Kinesin8B in axoneme assembly during male gamete formation and mosquito transmission

Sexual development is an essential phase in the Plasmodium life cycle, where male gametogenesis is an unusual and extraordinarily rapid process. It produces 8 haploid motile microgametes, from a microgametocyte within 15 minutes. Its unique achievement lies in linking the assembly of 8 axonemes in the cytoplasm to the three rounds of intranuclear genome replication, forming motile microgametes, which are expelled in a process called exflagellation. Surprisingly little is known about the actors involved in these processes. We are interested in kinesins, molecular motors that could play potential roles in male gametogenesis. We have undertaken a functional characterization in Plasmodium berghei of kinesin‐8B (PbKIN8B) expressed specifically in male gametocytes and gametes. By generating Pbkin8B‐gfp parasites, we show that PbKIN8B is specifically expressed during male gametogenesis and is associated with the axoneme. We created a ΔPbkin8B knockout cell line and analysed the consequences of the absence of PbKIN8B on male gametogenesis. We show that the ability to produce sexually differentiated gametocytes is not affected in ΔPbkin8B parasites and that the 3 rounds of genome replication occur normally. Nevertheless, the development to free motile microgametes is halted and the life cycle is interrupted in vivo. Ultrastructural analysis revealed that intranuclear mitoses are unaffected whereas cytoplasmic microtubules, although assembled in doublets and elongated, fail to assemble in the normal axonemal ‘9+2' structure and become motile. Absence of a functional axoneme prevented microgamete assembly and release from the microgametocyte, severely reducing infection of the mosquito vector. This is the first functional study of a kinesin involved in male gametogenesis. These results reveal a previously unknown role for PbKIN8B in male gametogenesis, providing new insights into Plasmodium flagellar formation.     

IL-10 is excluded from the functional cytokine memory of human CD4+ memory T lymphocytes

Epigenetic modifications, including DNA methylation, profoundly influence gene expression of CD4(+) Th-specific cells thereby shaping memory Th cell function. We demonstrate here a correlation between a lacking fixed potential of human memory Th cells to re-express the immunoregulatory cytokine gene IL10 and its DNA methylation status. Memory Th cells secreting IL-10 or IFN-gamma were directly isolated ex vivo from peripheral blood of healthy volunteers, and the DNA methylation status of IL10 and IFNG was assessed. Limited difference in methylation was found for the IL10 gene locus in IL-10-secreting Th cells, as compared with Th cells not secreting IL-10 isolated directly ex vivo or from in vitro-established human Th1 and Th2 clones. In contrast, in IFN-gamma(+) memory Th cells the promoter of the IFNG gene was hypomethylated, as compared with IFN-gamma-nonsecreting memory Th cells. In accordance with the lack of epigenetic memory, almost 90% of ex vivo-isolated IL-10-secreting Th cells lacked a functional memory for IL-10 re-expression after restimulation. Our data indicate that IL10 does not become epigenetically marked in human memory Th cells unlike effector cytokine genes such as IFNG. The exclusion of IL-10, but not effector cytokines, from the functional memory of human CD4(+) T lymphocytes ex vivo may reflect the need for appropriate regulation of IL-10 secretion, due to its potent immunoregulatory potential.     

Aluminium blunts the proliferative response and increases apoptosis of cultured human cells: putative relationship to Alzheimer's disease

Aluminium (Al) has been investigated as a neurotoxic substance. Al ranks among the potential environmental risk factors for Alzheimer's disease (AD). Epidemiological studies tested the relationship between Al in drinking water and AD, showing a significant correlation between elevated levels of monomeric Al in water and AD, although data to date remain inconclusive with respect to total Al. The aim of this study was to test whether or not Al exacerbates cellular toxicity mediated by the amyloid beta (Abeta) peptide. We evaluated the role of Al in modulating programmed cell death (apoptosis) in human cell cultures. We used the osteosarcoma cell line monolayer (SaOs-2) to demonstrate that treatment of SaOs-2 cultures with the Abeta peptide mid-fragment (25 to 35) at nano M, followed by co-incubation with physiological concentrations of aluminium chloride, which release monomeric Al in solution, led to marked expression of caspase 3, but not caspase 9, key markers of the apoptotic process. The same experimental conditions were shown to blunt significantly the proliferative response of normal human peripheral blood mononuclear cells (PBMC) to phytohemagglutinin (PHA) stimulation. Our observations support the hypothesis that Al significantly impairs certain cellular immune responses, and confirm that Al-mediated cell toxicity may play an important role in AD.     

Experimental Colitis Is Associated with Transcriptional Inhibition of Na+/Ca2+ Exchanger Isoform 1 (NCX1) Expression by Interferon γ in the Renal Distal Convoluted Tubules

NCX1 is a Na⁺/Ca²⁺ exchanger, which is believed to provide a key route for basolateral Ca²⁺ efflux in the renal epithelia, thus contributing to renal Ca²⁺ reabsorption. Altered mineral homeostasis, including intestinal and renal Ca²⁺ transport may represent a significant component of the pathophysiology of the bone mineral density loss associated with Inflammatory Bowel Diseases (IBD). The objective of our research was to investigate the effects of TNBS and DSS colitis and related inflammatory mediators on renal Ncx1 expression. Colitis was associated with decreased renal Ncx1 expression, as examined by real-time RT-PCR, Western blotting, and immunofluorescence. In mIMCD3 cells, IFNγ significantly reduced Ncx1 mRNA and protein expression. Similar effects were observed in cells transiently transfected with a reporter construct bearing the promoter region of the kidney-specific Ncx1 gene. This inhibitory effect of IFNγ is mediated by STAT1 recruitment to the proximal promoter region of Ncx1. Further in vivo study with Stat1^−/− mice confirmed that STAT1 is indeed required for the IFNγ mediated Ncx1 gene regulation. These results strongly support the hypothesis that impaired renal Ca²⁺ handling occurs in experimental colitis. Negative regulation of NCX1- mediated renal Ca²⁺ absorption by IFNγ may significantly contribute to the altered Ca²⁺ homeostasis in IBD patients and to IBD-associated loss of bone mineral density.