Quinine blocks 5-HT and 5-HT3 receptor mediated peristalsis in both guinea pig and mouse ileum tissue

Introduction. Quinine is commonly used to treat malaria; however one of the principal side effects is gastrointestinal disturbances (White, 1992). 5-HT3 receptors modulate gut peristalsis (Chetty et al., 2006), and, as quinine has been shown to act as a 5-HT3 receptor antagonist (Thompson and Lummis, 2008) it is possible that these side effects result from actions at gut 5-HT3 receptors. To address this question, we examined the ability of quinine to antagonise 5-HT and 5-HT3 mediated peristalsis in guinea pig and mouse ileum. Methods. Ileum was excised from male guinea pigs (200-300g) and C57BL/6 mice (25-35g) following cervical dislocation. Ileum segments (3-5 cm) were mounted in 50 ml organ baths containing Tryode’s solution at 35-37 °C. Concentration-response curves were constructed for 5-HT and the selective 5-HT3 agonist 2-Me-5-HT (non-cumulative doses). Quinine was pre-applied for 10 min and inhibition measured using agonist concentrations that elicited a submaximal response. Results. Concentration-dependent contractions produced by 5-HT (pEC50 = 5.45 ± 0.17, n = 8) and the selective 5-HT3 agonist 2-Me-5-HT (5.01 ± 0.17, n = 11) were not significantly different (Student’s t-test, t = 0.619, df = 17, p = 0.544) in guinea pig ileum. Increasing concentrations of quinine were able to antagonise the activities of both 5-HT (pIC50 = 5.03 ± 0.2, n = 6) and 2-Me-5HT (pIC50 = 4.59 ± 0.26, n = 4). At mouse ileum, 5-HT (pEC50 = 7.57 ± 0.33, n = 9) was more potent (Student’s t-test, t = 3.6, df = 12, p = 0.004) than 2-Me-5-HT (pEC50 = 5.45 ± 0.58, n = 5). Quinine antagonised both the 5-HT (pIC50 = 4.87 ± 0.31, n = 7) and 2-Me-5-HT-induced (pIC50 = 6.18 ± 1.14, n = 4) contractions. Conclusions. These results support previous electrophysiological studies that identified quinine as an antagonist at recombinant 5-HT3 receptors with IC50 values comparable with those reported here (pIC50 = 4.87, Thompson et al., 2007). Further, we found that quinine completely blocked 5-HT induced contractions in mouse and guinea pig, raising the possibility that quinine targets other 5-HT receptors in the gut (e.g., 5-HT4 receptors) and may influence intestinal function

The large‐scale freshwater cycle of the Arctic

This paper synthesizes our understanding of the Arctic\u27s large‐scale freshwater cycle. It combines terrestrial and oceanic observations with insights gained from the ERA‐40 reanalysis and land surface and ice‐ocean models. Annual mean freshwater input to the Arctic Ocean is dominated by river discharge (38%), inflow through Bering Strait (30%), and net precipitation (24%). Total freshwater export from the Arctic Ocean to the North Atlantic is dominated by transports through the Canadian Arctic Archipelago (35%) and via Fram Strait as liquid (26%) and sea ice (25%). All terms are computed relative to a reference salinity of 34.8. Compared to earlier estimates, our budget features larger import of freshwater through Bering Strait and larger liquid phase export through Fram Strait. While there is no reason to expect a steady state, error analysis indicates that the difference between annual mean oceanic inflows and outflows (∼8% of the total inflow) is indistinguishable from zero. Freshwater in the Arctic Ocean has a mean residence time of about a decade. This is understood in that annual freshwater input, while large (∼8500 km3), is an order of magnitude smaller than oceanic freshwater storage of ∼84,000 km3. Freshwater in the atmosphere, as water vapor, has a residence time of about a week. Seasonality in Arctic Ocean freshwater storage is nevertheless highly uncertain, reflecting both sparse hydrographic data and insufficient information on sea ice volume. Uncertainties mask seasonal storage changes forced by freshwater fluxes. Of flux terms with sufficient data for analysis, Fram Strait ice outflow shows the largest interannual variability

Metabolomic profiling of macrophages determines the discrete metabolomic signature and metabolomic interactome triggered by polarising immune stimuli

Priming and activating immune stimuli have profound effects on macrophages, however, studies generally evaluate stimuli in isolation rather than in combination. In this study we have investigated the effects of pro-inflammatory and anti-inflammatory stimuli either alone or in combination on macrophage metabolism. These stimuli include host factors such as IFNγ and ovalbumin-immunoglobulin immune complexes, or pathogen factors such as LPS. Untargeted LC-MS based metabolomics provided an in-depth profile of the macrophage metabolome, and revealed specific changes in metabolite abundance upon either individual stimuli or combined stimuli. Here, by factoring in an interaction term in the linear model, we define the metabolome interactome. This approach allowed us to determine whether stimuli interact in a synergistic or antagonistic manner. In conclusion this study demonstrates a robust approach to interrogate immune-metabolism, especially systems that model host-pathogen interactions

Host reticulocytes provide metabolic reservoirs that can be exploited by malaria parasites

Human malaria parasites proliferate in different erythroid cell types during infection. Whilst Plasmodium vivax exhibits a strong preference for immature reticulocytes, the more pathogenic P. falciparum primarily infects mature erythrocytes. In order to assess if these two cell types offer different growth conditions and relate them to parasite preference, we compared the metabolomes of human and rodent reticulocytes with those of their mature erythrocyte counterparts. Reticulocytes were found to have a more complex, enriched metabolic profile than mature erythrocytes and a higher level of metabolic overlap between reticulocyte resident parasite stages and their host cell. This redundancy was assessed by generating a panel of mutants of the rodent malaria parasite P. berghei with defects in intermediary carbon metabolism (ICM) and pyrimidine biosynthesis known to be important for P. falciparum growth and survival in vitro in mature erythrocytes. P. berghei ICM mutants (pbpepc-, phosphoenolpyruvate carboxylase and pbmdh-, malate dehydrogenase) multiplied in reticulocytes and committed to sexual development like wild type parasites. However, P. berghei pyrimidine biosynthesis mutants (pboprt-, orotate phosphoribosyltransferase and pbompdc-, orotidine 5′-monophosphate decarboxylase) were restricted to growth in the youngest forms of reticulocytes and had a severe slow growth phenotype in part resulting from reduced merozoite production. The pbpepc-, pboprt- and pbompdc- mutants retained virulence in mice implying that malaria parasites can partially salvage pyrimidines but failed to complete differentiation to various stages in mosquitoes. These findings suggest that species-specific differences in Plasmodium host cell tropism result in marked differences in the necessity for parasite intrinsic metabolism. These data have implications for drug design when targeting mature erythrocyte or reticulocyte resident parasites

Small polar hits against <i>S. aureus</i>:Screening, initial hit optimization and metabolomic studies

The global prevalence of antibacterial resistance requires new antibacterial drugs with novel chemical scaffolds and modes of action. It is also vital to design compounds with optimal physicochemical properties to permeate the bacterial cell envelope. We described an approach of combining and integrating whole cell screening and metabolomics into early antibacterial drug discovery using a library of small polar compounds. Whole cell screening of a diverse library of small polar compounds against Staphylococcus aureus gave compound 2. Hit expansion was carried out to determine structure–activity relationships. A selection of compounds from this series, together with other screened active compounds, was subjected to an initial metabolomics study to provide a metabolic fingerprint of the mode of action. It was found that compound 2 and its analogues have a different mode of action from some of the known antibacterial compounds tested. This early study highlighted the potential of whole cell screening and metabolomics in early antibacterial drug discovery. Future works will require improving potency and performing orthogonal studies to confirm the modes of action

Drug resistance and treatment failure in leishmaniasis: A 21st century challenge

Reevaluation of treatment guidelines for Old and New World leishmaniasis is urgently needed on a global basis because treatment failure is an increasing problem. Drug resistance is a fundamental determinant of treatment failure, although other factors also contribute to this phenomenon, including the global HIV/AIDS epidemic with its accompanying impact on the immune system. Pentavalent antimonials have been used successfully worldwide for the treatment of leishmaniasis since the first half of the 20th century, but the last 10 to 20 years have witnessed an increase in clinical resistance, e.g., in North Bihar in India. In this review, we discuss the meaning of “resistance” related to leishmaniasis and discuss its molecular epidemiology, particularly for Leishmania donovani that causes visceral leishmaniasis. We also discuss how resistance can affect drug combination therapies. Molecular mechanisms known to contribute to resistance to antimonials, amphotericin B, and miltefosine are also outlined

Untargeted metabolomics to understand the basis of phenotypic differences in amphotericin B-resistant

Background: Protozoan Leishmania parasites are responsible for a range of clinical infections that represent a substantial challenge for global health. Amphotericin B (AmB) is increasingly used to treat Leishmania infection, so understanding the potential for resistance to this drug is an important priority. Previously we described four independently-derived AmB-resistant L. mexicana lines that exhibited resistance-associated genetic lesions resulting in altered sterol content. However, substantial phenotypic variation between these lines, including differences in virulence attributes, were not fully explained by these changes. Methods: To identify alterations in cellular metabolism potentially related to phenotypic differences between wild-type and AmB-resistant lines, we extracted metabolites and performed untargeted metabolomics by liquid chromatography-mass spectrometry. Results: We observed substantial differences in metabolite abundance between lines, arising in an apparently stochastic manner. Concerted remodeling of central carbon metabolism was not observed; however, in three lines, decreased abundance of several oligohexoses was observed. Given that the oligomannose mannogen is an important virulence factor in Leishmania, this could relate to loss of virulence in these lines. Increased abundance of the reduced forms of the oxidative stress-protective thiols trypanothione and glutathione was also observed in multiple lines. Conclusions: This dataset will provide a useful resource for understanding the molecular basis of drug resistance in Leishmania, and suggests a role for metabolic changes separate from the primary mechanism of drug resistance in determining the phenotypic profile of parasite lines subjected to experimental selection of resistance

The early transcriptomic response to interleukin 1β and interleukin 33 in rat neonatal cardiomyocytes

In the heart, inflammatory cytokines including interleukin (IL) 1β are implicated in regulating adaptive and maladaptive changes, whereas IL33 negatively regulates cardiomyocyte hypertrophy and promotes cardioprotection. These agonists signal through a common co-receptor but, in cardiomyocytes, IL1β more potently activates mitogen-activated protein kinases and NFκB, pathways that regulate gene expression. We compared the effects of external application of IL1β and IL33 on the cardiomyocyte transcriptome. Neonatal rat cardiomyocytes were exposed to IL1β or IL33 (0.5, 1 or 2h). Transcriptomic profiles were determined using Affymetrix rat genome 230 2.0 microarrays and data were validated by quantitative PCR. IL1β induced significant changes in more RNAs than IL33 and, generally, to a greater degree. It also had a significantly greater effect in downregulating mRNAs and in regulating mRNAs associated with selected pathways. IL33 had a greater effect on a small, select group of specific transcripts. Thus, differences in intensity of intracellular signals can deliver qualitatively different responses. Quantitatively different responses in production of receptor agonists and transcription factors may contribute to qualitative differences at later times resulting in different phenotypic cellular responses