Antimalarial drugs disrupt ion homeostasis in malarial parasites

Plasmodium chabaudi malaria parasite organelles are major elements for ion homeostasis and cellular signaling and also target for antimalarial drugs. By using confocal imaging of intraerythrocytic parasites we demonstrated that the dye acridine orange (AO) is accumulated into P. chabaudi subcellular compartments. The AO could be released from the parasite organelles by collapsing the pH gradient with the K+/H+ ionophore nigericin (20 µM), or by inhibiting the H+-pump with bafilomycin (4 µM). Similarly, in isolated parasites loaded with calcium indicator Fluo 3-AM, bafilomycin caused calcium mobilization of the acidic calcium pool that could also be release with nigericin. Interestingly after complete release of the acidic compartments, addition of thapsigargin at 10 µM was still effective in releasing parasite intracellular calcium stores in parasites at trophozoite stage. The addition of antimalarial drugs chloroquine and artemisinin resulted in AO release from acidic compartments and also affected maintenance of calcium in ER store by using different drug concentrations.

What new cell biology findings could bring to therapeutics: is it time for a phenome-project in Toxoplasma gondii?

"If you know the enemy and know yourself, you need not fear the result of a hundred battles. If you know yourself but not the enemy, for every victory gained you will also suffer a defeat" (SunTzu the Art of War, 544-496 BC). Although written for the managing of conflicts and winning clear victories, this basic guideline can be directly transferred to our battle against apicomplexan parasites and how to focus future basic research in order to transfer the gained knowledge to a therapeutic intervention stratey. Over the last two decades the establishment of several key-technologies, by different groups working on Toxoplasma gondii, made this important human pathogen accessible to modern approaches in molecular cell biology. In fact more and more researchers get attracted to this easy accessible model organism to study specific biological questions, unique to apicomplexans. This fascinating, unique biology might provide us with new therapeutic options in our battle against apicomplexan parasites by finding its Achilles' heel. In this article we argue that in the absence of a powerful high throughput technology for the characterisation of essential gene of interests a coordinated effort should be undertaken to convert our knowledge of the genome into one of the phenome.

Antifungal activity of extracts from Atacama Desert fungi againstParacoccidioides brasiliensis and identification ofAspergillus felis as a promising source of natural bioactive compounds

Fungi of the genus Paracoccidioides are responsible for paracoccidioidomycosis. The occurrence of drug toxicity and relapse in this disease justify the development of new antifungal agents. Compounds extracted from fungal extract have showing antifungal activity. Extracts of 78 fungi isolated from rocks of the Atacama Desert were tested in a microdilution assay against Paracoccidioides brasiliensis Pb18. Approximately 18% (5) of the extracts showed minimum inhibitory concentration (MIC) values≤ 125.0 µg/mL. Among these, extract from the fungus UFMGCB 8030 demonstrated the best results, with an MIC of 15.6 µg/mL. This isolate was identified as Aspergillus felis (by macro and micromorphologies, and internal transcribed spacer, β-tubulin, and ribosomal polymerase II gene analyses) and was grown in five different culture media and extracted with various solvents to optimise its antifungal activity. Potato dextrose agar culture and dichloromethane extraction resulted in an MIC of 1.9 µg/mL against P. brasiliensis and did not show cytotoxicity at the concentrations tested in normal mammalian cell (Vero). This extract was subjected to bioassay-guided fractionation using analytical C18RP-high-performance liquid chromatography (HPLC) and an antifungal assay using P. brasiliensis. Analysis of the active fractions by HPLC-high resolution mass spectrometry allowed us to identify the antifungal agents present in the A. felis extracts cytochalasins. These results reveal the potential of A. felis as a producer of bioactive compounds with antifungal activity.

Handling sticky resin by stingless bees (Hymenoptera, Apidae)

For their nest defense, stingless bees (Meliponini) collect plant resins which they stick on intruders like ants or cleptobiotic robber bees causing their immobilization. The aim of this article is to identify all parts of stingless bee workers contacting these sticky resins. Of special interest are those body parts with anti-adhesive properties to resin, where it can be removed without residues. For that, extensive behavioral observations during foraging flight, handling and application of the resin have been carried out. When handling the resin, all tarsi touch the resin while walking above it. For transportation from plants to the nest during foraging flight, the resin is packed to the corbicula via tarsi and basitarsi of front and middle legs. Once stuck to the resin or after the corbicula had been unloaded, the bee's legs have to be cleaned thoroughly. Only the tips of the mandibles, that form, cut and apply the sticky resin, seem to have at least temporarily resin-rejecting properties.

Habilidade competitiva relativa de milhã em convivência com arroz irrigado e soja

O objetivo deste trabalho foi comparar a habilidade competitiva relativa de milhã com as culturas de arroz irrigado e de soja. Realizaram-se cinco experimentos, nas safras de 2009/2010 e 2010/2011, em delineamento inteiramente casualizado, com quatro repetições. Os tratamentos consistiram de proporções de plantas de milhã e arroz irrigado ou soja (100:0, 75:25, 50:50, 25:75 e 0:100%). A área foliar e a massa de matéria seca da parte aérea foram avaliadas aos 45 dias após a emergência. A competição entre milhã e arroz irrigado ou soja ocorre pelos mesmos recursos do ambiente. Plantas de milhã apresentam menor habilidade competitiva por indivíduo do que o arroz irrigado e a soja, quando ocorrem na mesma proporção de plantas. Quando as culturas do arroz irrigado ou da soja estão associadas ao milhã, a competição intraespecífica predomina, enquanto, para a planta daninha, prevalece a competição interespecífica como a mais prejudicial.

Implementação do protocolo TRS-398 para feixes de elétrons no Hospital Israelita Albert Einstein

OBJETIVO: A fim de simplificar a utilização das tabelas correspondentes aos fatores de qualidade no TRS-398 da International Atomic Energy Agency na calibração cruzada da câmara de placas paralelas por comparação com uma câmara cilíndrica calibrada em um feixe de elétrons de qualidade Qcross, é introduzida uma energia intermediária e arbitrária Qint na qual a câmara de placas paralelas deverá ser calibrada. O objetivo deste trabalho consiste em avaliar a escolha desta energia intermediária. MATERIAIS E MÉTODOS: Uma câmara de placas paralelas da Scanditronix, modelo NACP-02, e uma câmara PTW Markus foram calibradas em dois aceleradores lineares da Varian, Clinac 2100C e Clinac 23EX, nas energias de 16 e 20 MeV, respectivamente. Como câmara de referência foi utilizada uma câmara da Nuclear Enterprises, modelo 2571, previamente calibrada em termos de Dw no Instituto de Pesquisas Energéticas e Nucleares, para um feixe de 60Co. RESULTADOS: Os fatores de calibração N D,w assim obtidos apresentam uma variação de 0,07%, o que pode ser considerado desprezível. Os valores de dose absorvida na água determinados no Clinac 2100C para uma Qint de 16 MeV apresentam uma variação de 0,04% para a menor energia (4 MeV) e de 2,6% para a maior energia (16 MeV), em relação ao protocolo TRS-381. Já para uma Qint de 20 MeV, as variações observadas são de 1,3% e 3,5%. Por outro lado, as doses determinadas no Clinac 23EX para as Qint de 16 MeV e de 20 MeV são de 1,9% e 2,0% e de 2,8% e 3,0%, respectivamente. CONCLUSÃO: A escolha de Qint não é tão crítica assim, principalmente quando não de dispõem de feixes de elétrons que tenham na prática um valor de R50 exatamente igual a 7,5 g.cm-2.

PRESERVATION OF PRIMARY FOREST CHARACTERISTICS DESPITE FRAGMENTATION AND ISOLATION IN A FOREST REMNANT FROM VIÇOSA, MG, BRAZIL1

According to its owners, the Forest of Seu Nico (FSN) from the Viçosa municipality, Minas Gerais, Brazil, never has been logged and is therefore considered a primary forest. Nevertheless, the forest patch suffered impacts due to selective wood and non-timber extraction, fragmentation and isolation. Aim of this study was to test if the FSN, despite impacts, preserved characteristics of primary forests, which are elevated percentages of non-pioneer (>90%), animal-dispersed (>80 %), understory (>50%) and endemic species (~40%). For that, all trees with diameter at breast height equal or major than 3.2 cm within a plot of 100 x 100 m were identified. With 218 tree species found within this hectare, the FSN's species richness is outstanding for the region. The percentages of non-pioneer (92 %), animal-dispersed (85 %), understory (55 %) and endemic species (39.2 %) from the FSN fulfill the criteria proposed for primary forest. Therefore, we conclude that the FSN maintained its characteristics as a primary forest which highlights its importance for the conservation of biotic resources in the region, where similar fragments are lacking or not described yet.

ANTAGONISM IS THE PREDOMINANT EFFECT OF HERBICIDE MIXTURES USED FOR IMIDAZOLINONE-RESISTANT BARNYARDGRASS (Echinochloa crus-galli) CONTROL

ABSTRACTHerbicides mixtures are used in many situations without the adequate knowledge related with the effect on major target weeds. The objective of this study was to evaluate the effects of different herbicides mixtures used in irrigated rice in order to establish the adequate combinations for the prevention and management of herbicide resistance in barnyardgrass (Echinochloa crus-galli). Three experiments were performed at field conditions with all major post-emergent herbicides used in irrigated rice in Brazil. The first experiment was performed with barnyardgrass resistant to imidazolinone herbicides and herbicides applied at label rates. The second and third experiments were performed with barnyardgrass resistant and susceptible to imidazolinone herbicides applied at doses of 50 or 75% of the label rates. The occurrence of additive, synergistic and antagonistic effects was identified at 18, 18 and 64%, respectively, among the total of 50 different associations of herbicide and rates evaluated. In general, the mixture of ACCase inhibitors with ALS inhibitors, quinclorac, clomazone + propanil or thiobencarb resulted in antagonism. Sinergic mixtures were found in clomazone with propanil + thiobencarb, profoxydim with cyhalofop-butyl or clomazone, and quinclorac with imazapyr + imazapic, bispyribac-sodium or cyhalofop-butyl. The mixtures of quinclorac with profoxydim were antagonic. Rice grain yield varied according to the efficiency of weed control. Seveveral mixtures were effective for imidazolinone resistant barnyardgrass control.

Melatonin modulation of presynaptic nicotinic acetylcholine receptors located on short noradrenergic neurons of the rat vas deferens: a pharmacological characterization

Melatonin, the pineal hormone produced during the dark phase of the light-dark cycle, modulates neuronal acetylcholine receptors located presynaptically on nerve terminals of the rat vas deferens. Recently we showed the presence of high affinity nicotine-binding sites during the light phase, and low and high affinity binding sites during the dark phase. The appearance of the low affinity binding sites was due to the nocturnal melatonin surge and could be mimicked by exposure to melatonin in vitro. The aim of the present research was to identify the receptor subtypes responsible for the functional response during the light and the dark phase. The rank order of potency of agonists was dimethylphenylpiperazinium (DMPP) = cytisine > nicotine > carbachol and DMPP = nicotine = cytisine > carbachol, during the light and dark phase, respectively, due to an increase in apparent affinity for nicotine. Mecamylamine similarly blocked the DMPP response during the light and the dark phase, while the response to nicotine was more efficiently blocked during the light phase. In contrast, methyllycaconitine inhibited the nicotine-induced response only at 21:00 h. Since a7 nicotinic acetylcholine receptors (nAChRs) have low affinity for nicotine in binding assays, we suggest that a mixed population composed of a3ß4 - plus a7-bearing nAChR subtypes is present at night. This plasticity in receptor subtypes is probably driven by melatonin since nicotine-induced contraction in organs from animals sacrificed at 15:00 h and incubated with melatonin (100 pg/ml, 4 h) is not totally blocked by mecamylamine. Thus melatonin, by acting directly on the short adrenergic neurons that innervate the rat vas deferens, induces the appearance of the low affinity binding site, probably an a7 nAChR subtype.

The brain decade in debate: IV. Chronobiology

The present article is the adapted version of an electronic symposium organized by the Brazilian Society of Neuroscience and Behavior (SBNeC) which took place on June 14, 2000. The text is divided into three sections: I. The main issues, II. Chronodrugs, and III. Methods. The first section is dedicated to the perspectives of chronobiology for the next decade, with opinions about the trends of future research being emitted and discussed. The second section deals mostly with drugs acting or potentially acting on the organism's timing systems. In the third section there are considerations about relevant methodological issues concerning data analysis.

Evaluation of the cholinomimetic actions of trimethylsulfonium, a compound present in the midgut gland of the sea hare Aplysia brasiliana (Gastropoda, Opisthobranchia)

Trimethylsulfonium, a compound present in the midgut gland of the sea hare Aplysia brasiliana, negatively modulates vagal response, indicating a probable ability to inhibit cholinergic responses. In the present study, the pharmacological profile of trimethylsulfonium was characterized on muscarinic and nicotinic acetylcholine receptors. In rat jejunum the contractile response induced by trimethylsulfonium (pD2 = 2.46 ± 0.12 and maximal response = 2.14 ± 0.32 g) was not antagonized competitively by atropine. The maximal response (Emax) to trimethylsulfonium was diminished in the presence of increasing doses of atropine (P<0.05), suggesting that trimethylsulfonium-induced contraction was not related to muscarinic stimulation, but might be caused by acetylcholine release due to presynaptic stimulation. Trimethylsulfonium displaced [³H]-quinuclidinyl benzilate from rat cortex membranes with a low affinity (Ki = 0.5 mM). Furthermore, it caused contraction of frog rectus abdominis muscles (pD2 = 2.70 ± 0.06 and Emax = 4.16 ± 0.9 g), which was competitively antagonized by d-tubocurarine (1, 3 or 10 µM) with a pA2 of 5.79, suggesting a positive interaction with nicotinic receptors. In fact, trimethylsulfonium displaced [³H]-nicotine from rat diaphragm muscle membranes with a Ki of 27.1 µM. These results suggest that trimethylsulfonium acts as an agonist on nicotinic receptors, and thus contracts frog skeletal rectus abdominis muscle and rat jejunum smooth muscle via stimulation of postjunctional and neuronal prejunctional nicotinic cholinoreceptors, respectively.

Melatonin and N-acetyl-serotonin cross the red blood cell membrane and evoke calcium mobilization in malarial parasites

The duration of the intraerythrocytic cycle of Plasmodium is a key factor in the pathogenicity of this parasite. The simultaneous attack of the host red blood cells by the parasites depends on the synchronicity of their development. Unraveling the signals at the basis of this synchronicity represents a challenging biological question and may be very important to develop alternative strategies for therapeutic approaches. Recently, we reported that the synchrony of Plasmodium is modulated by melatonin, a host hormone that is synthesized only during the dark phases. Here we report that N-acetyl-serotonin, a melatonin precursor, also releases Ca2+ from isolated P. chabaudi parasites at micro- and nanomolar concentrations and that the release is blocked by 250 mM luzindole, an antagonist of melatonin receptors, and 20 mM U73122, a phospholipase C inhibitor. On the basis of confocal microscopy, we also report the ability of 0.1 µM melatonin and 0.1 µM N-acetyl-serotonin to cross the red blood cell membrane and to mobilize intracellular calcium in parasites previously loaded with the fluorescent calcium indicator Fluo-3 AM. The present data represent a step forward into the understanding of the signal transduction process in the host-parasite relationship by supporting the idea that the host hormone melatonin and N-acetyl-serotonin generate IP3 and therefore mobilize intracellular Ca2+ in Plasmodium inside red blood cells.

Influence of melatonin on the development of functional nicotinic acetylcholine receptors in cultured chick retinal cells

The influence of melatonin on the developmental pattern of functional nicotinic acetylcholine receptors was investigated in embryonic 8-day-old chick retinal cells in culture. The functional response to acetylcholine was measured in cultured retina cells by microphysiometry. The maximal functional response to acetylcholine increased 2.7 times between the 4th and 5th day in vitro (DIV4, DIV5), while the Bmax value for [125I]-alpha-bungarotoxin was reduced. Despite the presence of alpha8-like immunoreactivity at DIV4, functional responses mediated by alpha-bungarotoxin-sensitive nicotinic acetylcholine receptors were observed only at DIV5. Mecamylamine (100 µM) was essentially without effect at DIV4 and DIV5, while dihydro-ß-erythroidine (10-100 µM) blocked the response to acetylcholine (3.0 nM-2.0 µM) only at DIV4, with no effect at DIV5. Inhibition of melatonin receptors with the antagonist luzindole, or melatonin synthesis by stimulation of D4 dopamine receptors blocked the appearance of the alpha-bungarotoxin-sensitive response at DIV5. Therefore, alpha-bungarotoxin-sensitive receptors were expressed in retinal cells as early as at DIV4, but they reacted to acetylcholine only after DIV5. The development of an alpha-bungarotoxin-sensitive response is dependent on the production of melatonin by the retinal culture. Melatonin, which is produced in a tonic manner by this culture, and is a key hormone in the temporal organization of vertebrates, also potentiates responses mediated by alpha-bungarotoxin-sensitive receptors in rat vas deferens and cerebellum. This common pattern of action on different cell models that express alpha-bungarotoxin-sensitive receptors probably reflects a more general mechanism of regulation of these receptors.

Para uma Dialéctica Constelar: Theodor W. Adorno à entrada do Século XXI1

RESUMO: Interrogando-se sobre o lugar da filosofia de Theodor W. Adorno no âmbito do pensamento crítico contemporâneo, o presente artigo procura dar conta dos revezes da recepção político-filosófica da dialéctica negativa (das posturas críticas de Habermas, Lyotard ou Agamben às mais favoráveis de Jameson e Holloway) e discutir a sua relevância actual. Defender-se-á que a politização do pensamento adorniano é possível, muito embora as suas valências críticas não se restrinjam a essa possibilidade. Hoje, a dialéctica negativa funcionaria também como antídoto contra os atalhos tomados pelas correntes "voluntarista" (Peter Hallward), "messiânica" (Agamben) e "ontológica" (realismo especulativo) da filosofia, à entrada do século XXI. Contudo, atendendo a que a relação entre teoria e prática é complexa em Adorno, a sua relevância actual ressaltaria em relação com as reacções críticas que o movimento do "realismo especulativo" tem suscitado. Em diálogo com alguns dos seus interlocutores (Markus Gabriel e Adrian Johnston), sugere-se que o desenvolvimento de uma "dialéctica constelar" depende da introdução de um elemento destotalizador no seio do diagnóstico radical - a um só tempo materialista e transcendental - da dialéctica negativa.