Dynamic protein S-palmitoylation mediates parasite life cycle progression and diverse mechanisms of virulence.

Eukaryotic parasites possess complex life cycles and utilize an assortment of molecular mechanisms to overcome physical barriers, suppress and/or bypass the host immune response, including invading host cells where they can replicate in a protected intracellular niche. Protein S-palmitoylation is a dynamic post-translational modification in which the fatty acid palmitate is covalently linked to cysteine residues on proteins by the enzyme palmitoyl acyltransferase (PAT) and can be removed by lysosomal palmitoyl-protein thioesterase (PPT) or cytosolic acyl-protein thioesterase (APT). In addition to anchoring proteins to intracellular membranes, functions of dynamic palmitoylation include - targeting proteins to specific intracellular compartments via trafficking pathways, regulating the cycling of proteins between membranes, modulating protein function and regulating protein stability. Recent studies in the eukaryotic parasites - Plasmodium falciparum, Toxoplasma gondii, Trypanosoma brucei, Cryptococcus neoformans and Giardia lamblia - have identified large families of PATs and palmitoylated proteins. Many palmitoylated proteins are important for diverse aspects of pathogenesis, including differentiation into infective life cycle stages, biogenesis and tethering of secretory organelles, assembling the machinery powering motility and targeting virulence factors to the plasma membrane. This review aims to summarize our current knowledge of palmitoylation in eukaryotic parasites, highlighting five exemplary mechanisms of parasite virulence dependent on palmitoylation.

Growth of Plasmodium falciparum in response to a rotating magnetic field.

BACKGROUND: Plasmodium falciparum is the deadliest strain of malaria and the mortality rate is increasing because of pathogen drug resistance. Increasing knowledge of the parasite life cycle and mechanism of infection may provide new models for improved treatment paradigms. This study sought to investigate the paramagnetic nature of the parasite's haemozoin to inhibit parasite viability. RESULTS: Paramagnetic haemozoin crystals, a byproduct of the parasite's haemoglobin digestion, interact with a rotating magnetic field, which prevents their complete formation, causing the accumulation of free haem, which is lethal to the parasites. Plasmodium falciparum cultures of different stages of intraerythrocytic growth (rings, trophozoites, and schizonts) were exposed to a magnetic field of 0.46 T at frequencies of 0 Hz (static), 1, 5, and 10 Hz for 48 h. The numbers of parasites were counted over the course of one intraerythrocytic life cycle via flow cytometry. At 10 Hz the schizont life stage was most affected by the rotating magnetic fields (p = 0.0075) as compared to a static magnetic field of the same strength. Parasite growth in the presence of a static magnetic field appears to aid parasite growth. CONCLUSIONS: Sequestration of the toxic haem resulting from haemoglobin digestion is key for the parasites' survival and the focus of almost all existing anti-malarial drugs. Understanding how the parasites create the haemozoin molecule and the disruption of its creation aids in the development of drugs to combat this disease.

The long-term uncertainty of biodegradable mulch film residues and associated microplastics pollution on plant-soil health.

Biodegradable mulch film potentially offers an encouraging alternative to conventional (petroleum-based) plastic films. Since biodegradable films are more susceptible to rapid degradation, more microplastics (MPs) are likely to be generated than conventional films within the same time frame, probably leading to more severe MPs pollution and associated effects. However, the effect of biodegradable mulch film residues and associated MPs pollution on plant-soil health remains uncertainty. Here, we evaluated the potential effect of bio-MPs pollution on soil carbon (C) and nutrient (i.e., N and P) cycling, soil biology (microorganisms and mesofauna), and plant health, as these are crucial to agroecosystem functioning and the delivery of key ecosystem services. Unlike the inert (and therefore recalcitrant) C contained within petroleum-based MPs, at least 80% of the C from bio-MPs is converted to CO2, with up to 20% immobilized in living microbial biomass (i.e., < 0.05 t C ha-1). Although biodegradable films are unlikely to be important in promoting soil C storage, they may accelerate microbial biomass turnover in the short term, as well as CO2 production. Compared to conventional MPs, bio-MPs degradation is more pronounced, thereby inducing greater alterations in microbial diversity and community composition. This may further alter N2O and CH4 emissions, and ultimately resulting in unpredictable consequences for global climate warming. The extent to which this may occur, however, has yet to be shown in either laboratory or field studies. In addition, bio-MPs have a large chance of forming nanoplastics, potentially causing a stronger toxic effect on plants relative to conventional MPs. Consequently, this would influence plant health, crop productivity, and food safety, leading to potential health risks. It is unclear, however, if these are direct effects on key plant processes (e.g. signaling, cell expansion) or indirect effects (e.g. nutrient deficiency or acidification). Overall, the question as to whether biodegradable mulch films offer a promising alternative to solve the conventional plastic legacy in soil over the long term remains unclear.

Field application of biodegradable microplastics has no significant effect on plant and soil health in the short term.

Bioplastics (biodegradable plastics) potentially offer an encouraging alternative to conventional (petroleum-based) plastics. In practice, bioplastics inevitably generate a large number of bio-microplastics (bio-MPs, diameter <5 mm) during the degradation progress. However, the impact of bio-MPs on plant and soil health within agroecosystems remains incomplete. Here, a field study was conducted to investigate the effect of two shapes (fiber and powder) of pure polylactic acid (PLA) bio-MPs on oat (Avena sativa L.) and soybean (Glycinemax (L.) Merr.) growth and soil health. Our results showed that PLA application at a representative soil loading rate of 0.2% (w/w) had no significant effect on soil enzyme activities, soil physicochemical properties (soil water content, pH, etc.), root characteristics, plant biomass, and crop yield. Thus, we conclude that soil quality, plant health, and ecosystem multifunctionality were not affected by PLA over one growing season (5 months) in the presence of either bio-MP shape (fiber and powder) for either crop species (oat and soybean). Overall, PLA based bio-MPs may not pose a significant threat to agroecosystem functions in the short term (days to months) in the field, thus may provide a viable environmentally benign solution to replace traditional non-biodegradable plastics in agroecosystems.

Biochar application to temperate grasslands: challenges and opportunities for delivering multiple ecosystem services.

Grasslands (natural, semi-natural and improved) occupy approximately one-third of the terrestrial biosphere and are key for global ecosystem service provision, storing up to 30% of soil organic carbon (SOC). To date, most research on soil carbon (C) sequestration has focused on croplands where the levels of native soil organic matter (SOM) are typically low and significant potential exists to replenish SOM stocks. However, with the renewed push to achieve "net zero" C emissions by 2050, grasslands may offer an additional C store, utilising tools such as biochar. Here, we critically evaluate the potential for biochar as a technology for increasing grassland C stocks, identifying a number of practical, economic, social and legislative challenges that need to be addressed before the widescale adoption of biochar may be achieved. We critically assess the current knowledge within the field of grassland biochar research in the context of ecosystem service provision and provide opinions on the applicability of biochar as an amendment to different types of grassland (improved, semi-improved and unimproved) and the potential effect on ecosystem provision using a range of application techniques in the topsoil and subsoil. We concluded that the key question remains, is it possible for managed grasslands to store more C, without causing a loss in additional ecosystem services? To address this question future research must take a more multidisciplinary and holistic approach when evaluating the potential role of biochar at sequestering C in grasslands to mitigate climate change. Supplementary Information: The online version contains supplementary material available at 10.1007/s42773-023-00232-y.

Earthworms mediate the influence of polyethylene (PE) and polylactic acid (PLA) microplastics on soil bacterial communities.

There is a growing body of evidence that suggests that both biodegradable and conventional (non-degradable) microplastics (MP) are hazardous to soil health by affecting the delivery of key ecological functions such as litter decomposition, nutrient cycling and water retention. Specifically, soil fauna may be harmed by the presence of MPs while also being involved in their disintegration, degradation, migration and transfer in soil. Therefore, a comprehensive understanding of the interactions between MPs and soil fauna is essential. Here, we conducted a 120-day soil microcosm experiment applying polyethylene (PE) and polylactic acid (PLA), in the absence/presence of the earthworm Eisenia nordenskioldi to estimate the relative singular and combined impact of MPs and earthworms on the soil bacterial community. Our findings revealed contrasting effects of PE and PLA on the composition and diversity of soil bacteria. All treatments affected the community and network structure of the soil bacterial community. Compared to the control (no MPs or earthworms), PE decreased bacterial alpha diversity, while PLA increased it. Patescibacteria were found to be significantly abundant in the PE group whereas Actinobacteria and Gemmatimonadetes were more abundant in PE, and PLA and earthworms groups. The presence of earthworms appeared to mediate the impact of PE/PLA on soil bacteria, potentially through bacterial consumption or by altering soil properties (e.g., pH, aeration, C availability). Earthworm presence also appeared to promote the chemical aging of PLA. Collectively, our results provide novel insights into the soil-fauna-driven impact of degradable/nondegradable MPs exposure on the long-term environmental risks associated with soil microorganisms.

Trypanosoma brucei Acyl-Protein Thioesterase-like (TbAPT-L) Is a Lipase with Esterase Activity for Short and Medium-Chain Fatty Acids but Has No Depalmitoylation Activity.

Dynamic post-translational modifications allow the rapid, specific, and tunable regulation of protein functions in eukaryotic cells. S-acylation is the only reversible lipid modification of proteins, in which a fatty acid, usually palmitate, is covalently attached to a cysteine residue of a protein by a zDHHC palmitoyl acyltransferase enzyme. Depalmitoylation is required for acylation homeostasis and is catalyzed by an enzyme from the alpha/beta hydrolase family of proteins usually acyl-protein thioesterase (APT1). The enzyme responsible for depalmitoylation in Trypanosoma brucei parasites is currently unknown. We demonstrate depalmitoylation activity in live bloodstream and procyclic form trypanosomes sensitive to dose-dependent inhibition with the depalmitoylation inhibitor, palmostatin B. We identified a homologue of human APT1 in Trypanosoma brucei which we named TbAPT-like (TbAPT-L). Epitope-tagging of TbAPT-L at N- and C- termini indicated a cytoplasmic localization. Knockdown or over-expression of TbAPT-L in bloodstream forms led to robust changes in TbAPT-L mRNA and protein expression but had no effect on parasite growth in vitro, or cellular depalmitoylation activity. Esterase activity in cell lysates was also unchanged when TbAPT-L was modulated. Unexpectedly, recombinant TbAPT-L possesses esterase activity with specificity for short- and medium-chain fatty acid substrates, leading to the conclusion, TbAPT-L is a lipase, not a depalmitoylase.

Increased nicotinic acetylcholine receptor protein underlies chronic nicotine-induced up-regulation of nicotinic agonist binding sites in mouse brain.

Chronic nicotine treatment elicits a brain region-selective increase in the number of high-affinity agonist binding sites, a phenomenon termed up-regulation. Nicotine-induced up-regulation of α4ß2-nicotinic acetylcholine receptors (nAChRs) in cell cultures results from increased assembly and/or decreased degradation of nAChRs, leading to increased nAChR protein levels. To evaluate whether the increased binding in mouse brain results from an increase in nAChR subunit proteins, C57BL/6 mice were treated with nicotine by chronic intravenous infusion. Tissue sections were prepared, and binding of [(125)I]3-((2S)-azetidinylmethoxy)-5-iodo-pyridine (A85380) to ß2*-nAChR sites, [(125)I]monoclonal antibody (mAb) 299 to α4 nAChR subunits, and [(125)I]mAb 270 to ß2 nAChR subunits was determined by quantitative autoradiography. Chronic nicotine treatment dose-dependently increased binding of all three ligands. In regions that express α4ß2-nAChR almost exclusively, binding of all three ligands increased coordinately. However, in brain regions containing significant ß2*-nAChR without α4 subunits, relatively less increase in mAb 270 binding to ß2 subunits was observed. Signal intensity measured with the mAbs was lower than that with [(125)I]A85380, perhaps because the small ligand penetrated deeply into the sections, whereas the much larger mAbs encountered permeability barriers. Immunoprecipitation of [(125)I]epibatidine binding sites with mAb 270 in select regions of nicotine-treated mice was nearly quantitative, although somewhat less so with mAb 299, confirming that the mAbs effectively recognize their targets. The patterns of change measured using immunoprecipitation were comparable with those determined autoradiographically. Thus, increases in α4ß2*-nAChR binding sites after chronic nicotine treatment reflect increased nAChR protein.

A high-dynamic-range digital RF-over-fiber link for MRI receive coils using delta-sigma modulation.

Coaxial cables commonly used to connect radio-frequency (RF) coil arrays with the control console of an MRI scanner are susceptible to electromagnetic coupling. As the number of RF channels increases, such coupling could result in severe heating and pose a safety concern. Non-conductive transmission solutions based on fiber-optic cables are considered to be one of the alternatives but are limited by the high dynamic range (>80 dB) of typical MRI signals. A new digital fiber-optic transmission system based on delta-sigma modulation (DSM) is developed to address this problem. A DSM-based optical link is prototyped using off-the-shelf components and bench-tested at different signal oversampling rates (OSRs). An end-to-end dynamic range (DR) of 81 dB, which is sufficient for typical MRI signals, is obtained over a bandwidth of 200 kHz, which corresponds to OSR = 50. A fully integrated custom fourth-order continuous-time DSM is designed in 180 nm CMOS technology to enable transmission of full-bandwidth MRI signals (up to 1 MHz) with an adequate DR. Initial electrical test results from this custom chip are also presented.

Moonlighting enzymes in parasitic protozoa.

Enzymes moonlight in a non-enzymatic capacity in a diverse variety of cellular processes. The discovery of these non-enzymatic functions is generally unexpected, and moonlighting enzymes are known in both prokaryotes and eukaryotes. Importantly, this unexpected multi-functionality indicates that caution might be needed on some occasions in interpreting phenotypes that result from the deletion or gene-silencing of some enzymes, including some of the best known enzymes from classic intermediary metabolism. Here, we provide an overview of enzyme moonlighting in parasitic protists. Unequivocal and putative examples of moonlighting are discussed, together with the possibility that the unusual biological characteristics of some parasites either limit opportunities for moonlighting to arise or perhaps contribute to the evolution of novel proteins with clear metabolic ancestry.

Pharmacological and immunochemical characterization of alpha2* nicotinic acetylcholine receptors (nAChRs) in mouse brain.

AIM: alpha2 nAChR subunit mRNA expression in mice is most intense in the olfactory bulbs and interpeduncular nucleus. We aimed to investigate the properties of alpha2* nAChRs in these mouse brain regions. METHODS: alpha2 nAChR subunit-null mutant mice were engineered. Pharmacological and immunoprecipitation studies were used to determine the composition of alpha2 subunit-containing (alpha2*) nAChRs in these two regions. RESULTS: [(125)I]Epibatidine (200 pmol/L) autoradiography and saturation binding demonstrated that alpha2 deletion reduces nAChR expression in both olfactory bulbs and interpeduncular nucleus (by 4.8+/-1.7 and 92+/-26 fmol mg(-1) protein, respectively). Pharmacological characterization using the beta2-selective drug A85380 to inhibit [(125)I]epibatidine binding proved inconclusive, so immunoprecipitation methods were used to further characterize alpha2* nAChRs. Protocols were established to immunoprecipitate beta2 and beta4 nAChRs. Immunoprecipitation specificity was ascertained using tissue from beta2- and beta4-null mutant mice, and efficacy was good (>90% of beta2* and >80% of beta4* nAChRs were routinely recovered). CONCLUSION: Immunoprecipitation experiments indicated that interpeduncular nucleus alpha2* nAChRs predominantly contain beta2 subunits, while those in olfactory bulbs contain mainly beta4 subunits. In addition, the immunoprecipitation evidence indicated that both nuclei, but especially the interpeduncular nucleus, express nAChR complexes containing both beta2 and beta4 subunits.

Effects of surface EMG rectification on power and coherence analyses: an EEG and MEG study.

Coherence between electromyography (EMG) and electroencephalography (EEG) or magnetoencephalography (MEG) is frequently examined to gain insights on neuromuscular binding. Commonly, EMG signals are rectified before coherence is computed. However, the appropriateness of EMG rectification in computing EMG-EEG/MEG coherence has never been validated. Since rectification is a non-linear operation and alters the EMG power spectrum, such a validation is important to ensure the accuracy of coherence calculation. In this study we experimentally investigated the effects of EMG rectification on EMG power spectra and its coherence with EEG/MEG signals. Subjects performed sustained isometric index finger abduction at approximately 5-10% maximal voluntary force (in both EEG-EMG and MEG-EMG experiments) and index finger tapping at approximately 2-4Hz (in EEG-EMG experiment only). Bipolar surface EMG data from the first dorsal interosseus (FDI) and EEG/MEG signals from the contralateral primary sensorimotor area (C3) were recorded simultaneously. Power spectra and coherence with the EEG/MEG were calculated before and after EMG rectification. The results show that rectification shifts EMG power to lower frequencies, possibly enhancing peaks of motor unit firing. Coherences with the EEG/MEG signals were not significantly changed by EMG rectification, indicating EMG rectification is overall an appropriate procedure in power and coherence analyses.

Conceptual designs of conduction cooled MgB2 magnets for 1.5 and 3.0T full body MRI systems.

Conceptual designs of 1.5 and 3.0 T full-body magnetic resonance imaging (MRI) magnets using conduction cooled MgB2 superconductor are presented. The sizes, locations, and number of turns in the eight coil bundles are determined using optimization methods that minimize the amount of superconducting wire and produce magnetic fields with an inhomogeneity of less than 10 ppm over a 45 cm diameter spherical volume. MgB2 superconducting wire is assessed in terms of the transport, thermal, and mechanical properties for these magnet designs. Careful calculations of the normal zone propagation velocity and minimum quench energies provide support for the necessity of active quench protection instead of passive protection for medium temperature superconductors such as MgB2. A new 'active' protection scheme for medium Tc based MRI magnets is presented and simulations demonstrate that the magnet can be protected. Recent progress on persistent joints for multifilamentary MgB2 wire is presented. Finite difference calculations of the quench propagation and temperature rise during a quench conclude that active intervention is needed to reduce the temperature rise in the coil bundles and prevent damage to the superconductor. Comprehensive multiphysics and multiscale analytical and finite element analysis of the mechanical stress and strain in the MgB2 wire and epoxy for these designs are presented for the first time. From mechanical and thermal analysis of our designs we conclude there would be no damage to such a magnet during the manufacturing or operating stages, and that the magnet would survive various quench scenarios. This comprehensive set of magnet design considerations and analyses demonstrate the overall viability of 1.5 and 3.0 T MgB2 magnet designs.

Immunolabeling demonstrates the interdependence of mouse brain alpha4 and beta2 nicotinic acetylcholine receptor subunit expression.

Immunolabeling of beta2 and alpha4 subunits was quantitated in brain sections (14 mum) using [(125)I]mAb 270 and [(125)I]mAb 299, respectively. Specificity was demonstrated by signal loss in beta2(-/-) and alpha4(-/-) brain sections, respectively. Even mild paraformaldehyde fixation severely affected immunolabeling, so this study used unfixed sections. Immunolabeling autoradiography was used to map and quantitate the effects of beta2 and alpha4 subunit-null mutations on their putative partner subunits' protein expression. [(125)I]mAb 299 labeling was nearly eliminated in beta2(-/-) sections, although dorsal interpeduncular nucleus (IPN) retained a faint signal. Therefore, alpha4 subunit expression is almost universally beta2-dependent. In contrast, alpha4-null mutation effects on [(125)I]mAb 270 immunolabeling varied widely among brain regions. In corticothalamic regions, [(125)I]mAb 270 labeling was eliminated. However, in habenulopeduncular regions, alpha4 genotype had no effect. Other (predominantly dopaminergic and optic tract) nuclei also retained reduced [(125)I]mAb 270 labeling in alpha4(-/-) sections. Thus, although most beta2 subunit protein expression is alpha4-dependent, this dependence is not universal. Presumably, residual beta2 subunits are found in non-alpha4* subtypes. Together, these results show that immunolabeling is applicable to reliable, quantitative investigations of neuronal nAChRs, and that subunit-null mutants can be appropriate controls for such experiments. In situ mRNA hybridization was also performed to determine if altered mRNA transcription mediated the interdependence of alpha4 and beta2 subunit expression. alpha4-Null mutation did not affect beta2 mRNA expression, nor did beta2 genotype affect alpha4 mRNA expression. Consequently, it seems that the two subunits' effects on each other's expression are mediated at the protein, rather than gene expression level.

Systematic Survey of Serine Hydrolase Activity in Mycobacterium tuberculosis Defines Changes Associated with Persistence.

The transition from replication to non-replication underlies much of Mycobacterium tuberculosis (Mtb) pathogenesis, as non- or slowly replicating Mtb are responsible for persistence and poor treatment outcomes. Therapeutic targeting of non-replicating populations is a priority for tuberculosis treatment, but few drug targets in non-replicating Mtb are currently known. Here, we directly measured the activity of the highly diverse and druggable serine hydrolases (SHs) during active replication and non-replication using activity-based proteomics. We predict SH activity for 78 proteins, including 27 proteins with unknown function, and identify 37 SHs that remain active in the absence of replication, providing a set of candidate persistence targets. Non-replication was associated with major shifts in SH activity. These activity changes were largely independent of SH abundance, indicating extensive post-translational regulation of SHs. By probing a large cross-section of druggable Mtb enzyme space during replication and non-replication, we identify new SHs and suggest new persistence targets.

Nonlinear cortical modulation of muscle fatigue: a functional MRI study.

Muscle fatigue has been studied for over a century, but almost no data are available to indicate how the brain perceives fatigue and modulates its signals to the fatiguing muscle. In this study, brain activation was measured by functional magnetic resonance imaging (fMRI) during a sustained (2-min) maximal-effort handgrip contraction while handgrip force and finger muscle electromyographic (EMG) data were recorded simultaneously by a magnetic resonance environment-adapted force-EMG measurement system. The results showed decoupled progresses in brain and muscle activities when muscle was fatigued and correlated behaviors among the cortical areas being analyzed. While handgrip force and EMG signals declined in parallel during the course of muscle fatigue, fMRI-measured brain activities first substantially increased and then decreased. This similar signal modulation occurred not only in the primary sensorimotor areas but also in the secondary and association cortices (supplementary motor, prefrontal, and cingulate areas). The nonlinear changes of brain signal may reflect an early adjustment to strengthen the descending command for force-loss compensation and subsequent inhibition by sensory feedback as fatigue became more severe. The close association in the activation pattern in many cortical regions may reflect integrated processing of information in the brain.

Evidence for loss of a partial flagellar glycolytic pathway during trypanosomatid evolution.

Classically viewed as a cytosolic pathway, glycolysis is increasingly recognized as a metabolic pathway exhibiting surprisingly wide-ranging variations in compartmentalization within eukaryotic cells. Trypanosomatid parasites provide an extreme view of glycolytic enzyme compartmentalization as several glycolytic enzymes are found exclusively in peroxisomes. Here, we characterize Trypanosoma brucei flagellar proteins resembling glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and phosphoglycerate kinase (PGK): we show the latter associates with the axoneme and the former is a novel paraflagellar rod component. The paraflagellar rod is an essential extra-axonemal structure in trypanosomes and related protists, providing a platform into which metabolic activities can be built. Yet, bioinformatics interrogation and structural modelling indicate neither the trypanosome PGK-like nor the GAPDH-like protein is catalytically active. Orthologs are present in a free-living ancestor of the trypanosomatids, Bodo saltans: the PGK-like protein from B. saltans also lacks key catalytic residues, but its GAPDH-like protein is predicted to be catalytically competent. We discuss the likelihood that the trypanosome GAPDH-like and PGK-like proteins constitute molecular evidence for evolutionary loss of a flagellar glycolytic pathway, either as a consequence of niche adaptation or the re-localization of glycolytic enzymes to peroxisomes and the extensive changes to glycolytic flux regulation that accompanied this re-localization. Evidence indicating loss of localized ATP provision via glycolytic enzymes therefore provides a novel contribution to an emerging theme of hidden diversity with respect to compartmentalization of the ubiquitous glycolytic pathway in eukaryotes. A possibility that trypanosome GAPDH-like protein additionally represents a degenerate example of a moonlighting protein is also discussed.

Calmodulin is required for paraflagellar rod assembly and flagellum-cell body attachment in trypanosomes.

In the flagellum of the African sleeping sickness parasite Trypanosoma brucei calmodulin (CaM) is found within the paraflagellar rod (PFR), an elaborate extra-axonemal structure, and the axoneme. In dissecting mechanisms of motility regulation we analysed CaM function using RNAi. Unexpectedly CaM depletion resulted in total and catastrophic failure in PFR assembly; even connections linking axoneme to PFR failed to form following CaM depletion. This provides an intriguing parallel with the role in the green alga Chlamydomonas of a CaM-related protein in docking outer-dynein arms to axoneme outer-doublet microtubules. Absence of CaM had no discernible effect on axoneme assembly, but the failure in PFR assembly was further compounded by loss of the normal linkage between PFR and axoneme to the flagellum attachment zone of the cell body. Thus, flagellum detachment was a secondary, time-dependent consequence of CaM RNAi, and coincided with the loss of normal trypomastigote morphology, thereby linking the presence of PFR architecture with maintenance of cell form, as well as cell motility. Finally, wider comparison between the flagellum detachment phenotypes of RNAi mutants for CaM and the FLA1 glycoprotein potentially provides new perspective into the function of the latter into establishing and maintaining flagellum-cell body attachment.

86Rb+ efflux mediated by alpha4beta2*-nicotinic acetylcholine receptors with high and low-sensitivity to stimulation by acetylcholine display similar agonist-induced desensitization.

The nicotinic acetylcholine receptors (nAChR) assembled from alpha4 and beta2 subunits are the most densely expressed subtype in the brain. Concentration-effect curves for agonist activation of alpha4beta2*-nAChR are biphasic. This biphasic agonist sensitivity is ascribed to differences in subunit stoichiometry. The studies described here evaluated desensitization elicited by low concentrations of epibatidine, nicotine, cytisine or methylcarbachol of brain alpha4beta2-nAChR function measured with acetylcholine-stimulated (86)Rb(+) efflux from mouse thalamic synaptosomes. Each agonist elicited concentration-dependent desensitization. The agonists differed in potency. However, IC(50) values for each agonist for desensitization of (86)Rb(+) efflux both with high (EC(50) approximately 3 microM) and low (EC(50) approximately 150 microM) acetylcholine sensitivity were not significantly different. Concentrations required to elicit desensitization were higher that their respective K(D) values for receptor binding. Even though the two components of alpha4beta2*-nAChR-mediated (86)Rb(+) efflux from mouse brain differ markedly in EC(50) values for agonist activation, they are equally sensitive to desensitization by exposure to low agonist concentrations. Mice were also chronically treated with nicotine by continuous infusion of 0, 0.5 or 4.0mg/kg/h and desensitization induced by nicotine was evaluated. Consistent with previous results, chronic nicotine treatment increased the density of epibatidine binding sites. Acute exposure to nicotine also elicited concentration-dependent desensitization of both high-sensitivity and low-sensitivity acetylcholine-stimulated (86)Rb(+) efflux from cortical and thalamic synaptosomes. Although chronic nicotine treatment reduced maximal (86)Rb(+) efflux from thalamus, IC(50) values in both brain regions were unaffected by chronic nicotine treatment.

Nonlinear features of surface EEG showing systematic brain signal adaptations with muscle force and fatigue.

Nonlinear dynamics has been introduced to the analysis of biological data and increasingly recognized to be functionally relevant. The purpose of this study was to examine chaotic properties of human scalp EEG signals associated with voluntary motor tasks using the largest Lyapunov exponent (L1). 64-channel scalp EEG data were recorded from eight healthy subjects in two tasks: (1) intermittent handgrip contractions at 20, 40, 60, and 80% of maximal voluntary contraction (MVC) with 20 trials at each level. No significant fatigue were induced; (2) intermittent handgrip MVCs (100 trials) that resulted in significant fatigue. The L1 values of all EEG channels were calculated in each trial first then averaged across the 20 trials at each force level (Task 1) or over each of the 5-trial blocks (Task 2) before the group means were obtained. A multivariate statistical model was used to examine the effect of force and fatigue on L1. L1 values were greater with higher force (Task 1), and decreased significantly with fatigue (Task 2). The L1 of the EEG signals changes systematically and correlates significantly with muscle force and fatigue. The results suggest that nonlinear chaotic index L1 may serve as a quantitative measure for motor control-related cortical signal adaptations.