A common inhibitory binding site for zinc and odorants at the voltage-gated K+ channel of rat olfactory receptor neurons

This study compared the effects of zinc and odorants on the voltage-gated K+ channel of rat olfactory neurons. Zinc reduced current magnitude, depolarized the voltage activation curve and slowed activation kinetics without affecting inactivation or deactivation kinetics. Zinc inhibition was potentiated by the NO compound, S-nitroso-cysteine. The pH- and diethylpyrocarbonate-dependence of zinc inhibition suggested that zinc acted by binding to histidine residues. Cysteine residues were eliminated as contributing to the zinc-binding site. The odorants, acetophenone and amyl acetate, also reduced current magnitude, depolarized the voltage activation curve and selectively slowed activation kinetics. Furthermore, the diethylpyrocarbonate- and pH-dependence of odorant inhibition implied that the odorants also bind to histidine residues. Zinc inhibitory potency was dramatically diminished in the presence of odorants, implying competition for a common binding site. These observations indicate that the odorants and zinc share a common inhibitory binding site on the external surface of the voltage-gated K+ channel.

An investigation into the possible inhibitory effects of white cypress pine (Callitris glaucophylla) litter on the germination and growth of associated ground cover species

White cypress-pine stands typically support sparse densities of shrubs and grasses. The commonly held opinion is that leaching of allelopathic chemical compounds from cypress-pine litter partly facilitates this exclusion. Germination and growth of cypress pine seedlings do not appear to be similarly affected. This study set out to determine whether cypress litter had a differential effect on germination and growth of cypress-pine seedlings and on associated ground-cover species. Glasshouse trials comparing seedling emergence under cypress- and artificial-litter layers were undertaken. Cypress-pine litter did not have an inhibitory effect on the germination or growth of ground-cover species. In most cases, seedling emergence was facilitated by the application of cypress-pine litter due to its ability to increase the water holding capacity of the underlying soil. Cypress litter did not promote growth of its own seedlings over its competitors except on coarse-textured soils where it provided an ameliorative function to water stress due to the soil's reduced water holding capacity. The inhibition of ground-cover species' germination and growth in pure cypress stands was suggested to be the result of high below-ground resource competition due to the pine's expansive root morphology.

Platelet inhibitory effects of the nitric oxide donor drug MAHMA NONOate in vivo in rats

The platelet inhibitory effects of the nitric oxide (NO) donor drug MAHMA NONOate ((Z-1-{N-methyl-N-[6-(N-methylammoniohexyl)amino] diazen-1-ium-1,2-diolate) were examined in anaesthetised rats and compared with those of S-nitrosoglutathione (GSNO; an S-nitrosothiol). Bolus administration of the aggregating agent ADP dose-dependently reduced the number of circulating free platelets. Intravenous infusions of MAHMA NONOate (3-30 nmol/kg/min) dose-dependently inhibited the effect of 0.3 mumol/kg ADP. MAHMA NONOate was approximately 10-fold more potent than GSNO. MAHMA NONOate (0.3-10 nmol/kg/min) also reduced systemic artery pressure and was again 10-fold more potent than GSNO. Thus MAHMA NONOate has both platelet inhibitory and vasodepressor effects in vivo. The dose ranges for these two effects overlapped, although blood pressure was affected at slightly lower doses. The platelet inhibitory effects compared favourably with those of GSNO, even though NONOates generate free radical NO which, in theory, could have been scavenged by haemoglobin. Therefore platelet inhibition may be a useful therapeutic property of NONOates. (C) 2003 Elsevier B.V. All rights reserved.

Parvalbumin-, calbindin-, and calretinin-immunoreactive neurons in the prefrontal cortex of the owl monkey (Aotus trivirgatus): A standardized quantitative comparison with sensory and motor areas

Recent studies have revealed regional variation in the density and distribution of inhibitory neurons in different cortical areas, which are thought to reflect area-specific specializations in cortical circuitry. However, there are as yet few standardized quantitative data regarding how the inhibitory circuitry in prefrontal cortex (PFC), which is thought to be involved in executive functions such as cognition, emotion and decision making, compares to that in other cortical areas. Here we used immunohistochemical techniques to determine the density and distribution of parvalbumin (PV)-, calbindin (CB)-, and calretinin (CR)-immunoreactive (ir) neurons and axon terminals in the dorsolateral and orbital PFC of the owl monkey (Aotus trivirgatus), and compared them directly with data obtained using the same techniques in 11 different visual, somatosensory and motor areas. We found marked differences in the density of PV-ir, CB-ir, and CR-ir interneurons in several cortical areas. One hundred and twenty eight of all 234 possible between-area pairwise comparisons were significantly different. The density of specific subpopulations of these cells also varied among cortical areas, as did the density of axon terminals. Comparison of PFC with other cortical areas revealed that 40 of all 66 possible statistical comparisons of the density of PV-ir, CB-ir, and CR-ir cells were significantly different. We also found evidence for heterogeneity in the pattern of labeling of PV-ir, CB-ir, and CR-ir cells and axon terminals between the dorsolateral and orbital subdivisions of PFC. These data are likely to reflect basic differences in interneuron circuitry, which are likely to influence inhibitory function in the cortex. Copyright (C) 2003 S. Karger AG, Basel.

Synthesis, characterization, electrochemical behavior and in vitro protein tyrosine kinase inhibitory activity of the cymene-halogenobenzohydroxamato [Ru(eta(6)-cymene)(bha)Cl] complexes

The ruthenium(II)-cymene complexes [Ru(eta(6)-cymene)(bha)Cl] with substituted halogenobenzohydroxamato (bha) ligands (substituents = 4-F, 4-Cl, 4-Br, 2,4-F-2, 3,4-F-2, 2,5-F-2, 2,6-F-2) have been synthesized and characterized by elemental analysis, IR, H-1 NMR, C-13 NMR, cyclic voltammetry and controlled-potential electrolysis, and density functional theory (DFT) studies. The compositions of their frontier molecular orbitals (MOs) were established by DFT calculations, and the oxidation and reduction potentials are shown to follow the orders of the estimated vertical ionization potential and electron affinity, respectively. The electrochemical E-L Lever parameter is estimated for the first time for the various bha ligands, which can thus be ordered according to their electron-donor character. All complexes exhibit very strong protein tyrosine kinase (PTK) inhibitory activity, even much higher than that of genistein, the clinically used PTK inhibitory drug. The complex containing the 2,4-difluorobenzohydroxamato ligand is the most active one, and the dependences of the PTK activity of the complexes and of their redox potentials on the ring substituents are discussed. (C) 2012 Elsevier B.V. All rights reserved.

Inhibitory effect of deferoxamine on Paracoccidioides brasiliensis survival in human monocytes: reversal by holotransferrin not by apotransferrin

The mechanisms used by Paracoccidioides brasiliensis to survive into phagocytic cells are not clear. Cellular iron metabolism is of critical importance to the growth of several intracellular pathogens whose capacity to multiply in mononuclear phagocytes is dependent on the availability of intracellular iron. Thus, the objective of this paper was to investigate the role of intracellular iron in regulating the capacity of P. brasiliensis yeast cells to survive within human monocytes. Treatment of monocytes with deferoxamine, an iron chelator, suppressed the survival of yeasts in a concentration-dependent manner. The effect of deferoxamine was reversed by iron-saturated transferrin (holotransferrin) but not by nonsaturated transferrin (apotransferrin). These results strongly suggest that P. brasiliensis survival in human monocytes is iron dependent.

The inhibitory activity of synthetic compounds and ions against transporters of multi-drug resistant bacteria

RESUMO: Sessenta e três derivados de hidantoína foram utilizados para avaliar possíveis efeitos de modulação na actividade das bombas de efluxo (BE) na Salmonella NCTC 13349 utilizando um método fluorimétrico semi-automático. Nenhum dos compostos apresentaram actividade anti-bacteriana até concentrações de 240 mg/L. Entre todos os compostos, SZ-7 demonstrou possuir propriedades de modulação de effluxo na presença de glucose. Para testar esta actividade, estirpes de Salmonella resistentes à ciprofloxacina, induzidas a elevados níveis de resistência com sobre-expressão de BE, foram expostas ao SZ-7. Este derivado afectou a susceptibilidade das estirpes à ciprofloxacina. Uma vez que os 63 compostos estudados apresentaram pouco efeito inibitório /cumulativo, apesar de serem conhecidos pelos seus efeitos moduladores de BE-dependentes de iões em eucariotas, foi questionado o papel dos iões na regulação de BE bacterianas, que poderão influenciar a eficácia de novos compostos. Para este estudo, utilizamos a Escherichia coli AG100 como modelo, devido ao extenso conhecimento no que respeita a estrutura e actividade das BE. Devido à importância de iões de cálcio (Ca2+) nos canais de transporte membranar e na actividade de ATPases, a sua actividade na modulação do efluxo foi investigada. De resultados anteriormente obtidos concluiu-se que a pH 5 o efluxo é independente de energia metabólica; contudo, a pH 8 é absolutamente dependente, sendo que o Ca2+ é indispensável para manter a actividade das ATPases bacterianas. A acumulação/effluxo de EtBr pela E. coli AG100 foi determinada na presença/ausência de Ca2+, clorpromazina (inibidor de ligação de Ca2+ a proteínas), e ácido etilenodiamino tetra-acético (quelante de Ca2+). Acumulação/effluxo aumentou a pH 8, contudo o Ca2+ reverte estes efeitos evidenciando a sua importância no funcionamento das BE bacterianas. Em resumo este trabalho colocou em evidência que muitos aspectos bioquímicos e bioenergéticos devem ser tomados em consideração no estudo da resistência bacteriana mediada por BE.------- ABSTRACT: Sixty-three hydantoin derivatives were evaluated for their modulating effects on efflux pump (EP) activity of Salmonella NCTC 13349 utilizing a semi-automatic fluorometric method. None of the compounds presented antibacterial activities at concentrations as high as 240 mg/L. Among all compounds, SZ-7 showed possible efflux modulating activity in the presence of glucose, indicative of a potential EP inhibitor. To verify its potential effects, ciprofloxacin-resistant Salmonella strains, induced to high level resistance with over-expressing EPs, were exposed to SZ-7. This derivative affected the susceptibility of the ciprofloxacin-resistant strains. Since the 63 compounds studied had very low inhibitory/accumulative effects, even though their known for being efficient in modulating ion-driven eukaryotic EPs, we questioned whether ions had a leading role in regulating bacterial EPs, influencing the effectiveness of new compounds. For this study we used Escherichia coli AG100 as a model, due to the extensive knowledge on its EPs structure and activity. Owing the importance of calcium ions (Ca2+) for membrane transport channels and activity of ATPases, the role of Ca2+ was investigated. From previous results we concluded that at pH 5 efflux is independent of metabolic energy; however, at pH 8 it is entirely dependent of metabolic energy and the Ca2+ ions are essential to maintain the activity of bacterial ATPases. Accumulation and efflux of ethidium bromide (EtBr) by E. coli AG100 was determined in the presence and absence of Ca2+, chlorpromazine (inhibitor of Ca2+-binding to proteins), and ethylenediaminetetraacetic acid (Ca2+ chelator). Accumulation of EtBr increased at pH 8; however Ca2+ reversed these effects providing information as to the importance of this ion in the regulation of bacterial EP systems. Overall this work puts in evidence that many biochemical and bioenergetic aspects related to the strains physiology need to be taken into consideration in bacterial drug resistance mediated by EPs.

Inhibitory effect of cyclophosphamide on the biosynthesis of a perchloro-soluble protein fraction of ehrlich ascites carcinoma cells

The perchloro-soluble mucroptotein fraction was determined in the cells of Ehrlich ascites carcinoma on the 10th and 12th days post-inoculation of the tumor. After 3 days of a single subcutaneous dose of cyclophosphamide (200 mg/kg) the mucoprotein levels were found considerable lower. This difference was highly significant statistically.

Nasal secretions of Neisseria lactamica carriers have an inhibitory effect on Neisseria meningitidis attachment to human oroepithelial cells

Nasal secretions of volunteers colonized by N. lactamica impaired the attachment of N. lactamica and of meningococci of groups A and B to oroepithelial cells. Bacterial adherence was found to be mediated by nonpiliated adhesins with antigen(s) which probably are shared by the strains tested. Although a strong attachment-inhibiting activity arises in their nasal secretions, volunteers remained colonized by N. lactamica. This evidence suggest that the eradication of Neisseria carriage is a multifactorial event.

Insulin secretion is regulated by the glucose-dependent production of islet beta cell macrophage migration inhibitory factor.

Macrophage migration inhibitory factor (MIF), originally identified as a cytokine secreted by T lymphocytes, was found recently to be both a pituitary hormone and a mediator released by immune cells in response to glucocorticoid stimulation. We report here that the insulin-secreting beta cell of the islets of Langerhans expresses MIF and that its production is regulated by glucose in a time- and concentration-dependent manner. MIF and insulin colocalize by immunocytochemistry within the secretory granules of the pancreatic islet beta cells, and once released, MIF appears to regulate insulin release in an autocrine fashion. In perifusion studies performed with isolated rat islets, immunoneutralization of MIF reduced the first and second phase of the glucose-induced insulin secretion response by 39% and 31%, respectively. Conversely, exogenously added recombinant MIF was found to potentiate insulin release. Constitutive expression of MIF antisense RNA in the insulin-secreting INS-1 cell line inhibited MIF protein synthesis and decreased significantly glucose-induced insulin release. MIF is therefore a glucose-dependent, islet cell product that regulates insulin secretion in a positive manner and may play an important role in carbohydrate metabolism.

Viral FLICE-inhibitory proteins (FLIPs) prevent apoptosis induced by death receptors.

Viruses have evolved many distinct strategies to avoid the host's apoptotic response. Here we describe a new family of viral inhibitors (v-FLIPs) which interfere with apoptosis signalled through death receptors and which are present in several gamma-herpesviruses (including Kaposi's-sarcoma-associated human herpesvirus-8), as well as in the tumorigenic human molluscipoxvirus. v-FLIPs contain two death-effector domains which interact with the adaptor protein FADD, and this inhibits the recruitment and activation of the protease FLICE by the CD95 death receptor. Cells expressing v-FLIPs are protected against apoptosis induced by CD95 or by the related death receptors TRAMP and TRAIL-R. The herpesvirus saimiri FLIP is detected late during the lytic viral replication cycle, at a time when host cells are partially protected from CD95-ligand-mediated apoptosis. Protection of virus-infected cells against death-receptor-induced apoptosis may lead to higher virus production and contribute to the persistence and oncogenicity of several FLIP-encoding viruses.

Macrophage Migration Inhibitory Factor Deficiency Is Associated With Impaired Killing of Gram-Negative Bacteria by Macrophages and Increased Susceptibility to Klebsiella pneumoniae Sepsis.

The cytokine macrophage migration inhibitory factor (MIF) is an important component of the early proinflammatory response of the innate immune system. However, the antimicrobial defense mechanisms mediated by MIF remain fairly mysterious. In the present study, we examined whether MIF controls bacterial uptake and clearance by professional phagocytes, using wild-type and MIF-deficient macrophages. MIF deficiency did not affect bacterial phagocytosis, but it strongly impaired the killing of gram-negative bacteria by macrophages and host defenses against gram-negative bacterial infection, as shown by increased mortality in a Klebsiella pneumonia model. Consistent with MIF's regulatory role of Toll-like 4 expression in macrophages, MIF-deficient cells stimulated with lipopolysaccharide or Escherichia coli exhibited reduced nuclear factor κB activity and tumor necrosis factor (TNF) production. Addition of recombinant MIF or TNF corrected the killing defect of MIF-deficient macrophages. Together, these data show that MIF is a key mediator of host responses against gram-negative bacteria, acting in part via a modulation of bacterial killing by macrophages.

The effect of subminimal inhibitory concentrations of penicillin on growth rate and haemolysin activity of group G Streptococcus

The influence of the subminimal inhibitory concentrations (1/3 and 1/4 of the MIC) of penicillin on growth rate and on haemolysin production of a strain of group G Streptococcus was studied. It was shown that 1/3 of the MIC almost completely inhibited the bacterial growth, but it was not able to inhibit haemolysin activity in the culture supernate. The generation time of bacteria grown in 1/4 of the MIC was approximately twice longer than that of the control culture. In all cultures, the haemolysin, after being produced (or liberated), reached a peak and decreased to low levels, which could suggest that group G Streptococcus produces some end products of metabolism that are able to inhibit haemolysin activity.

Serum factors inhibitory for in vitro development of Plasmodium falciparum blood-stage parasites

Sera from 29 individuals residing in a malaria-endemic region of Colombia were evaluated by an inhibition assay for their capacity to retard the growth of Plasmodium falciparum in vitro. The inhibitory activity was found to be independent of antibody activity. Furthermore, the degree of inhibition of parasite development was variable, depending on the parasite isolate used for the assay and the season of malaria transmission. We selected sera with high inhibitory activity and carried out partial analytical characterization by anion exchange fast protein liquid chromatography (FPLC) to identify the chemical nature of the inhibitory factor(s). The results suggested that the in vitro inhibitory activity might result from the additive effect of different molecules. It appears that these molecules could be non-specifically induced by stimulation of the immune system, they seem to play a role in the immunity to malaria.