Modulation of neuritogenesis by astrocyte muscarinic receptors

Astrocytes have been shown to release factors that have promoting or inhibiting effects on neuronal development. However, mechanisms controlling the release of such factors from astrocytes are not well established. Astrocytes express muscarinic receptors whose activation stimulates a robust intracellular signaling, although the role of these receptors in glial cells is not well understood. Acetylcholine and acetylcholine receptors are present in the brain before synaptogenesis occurs and are believed to be involved in neuronal maturation. The present study was undertaken to investigate whether stimulation of muscarinic receptors in astrocytes would modulate neurite outgrowth in hippocampal neurons. Rat hippocampal neurons, co-cultured with rat cortical astrocytes previously exposed to the cholinergic agonist carbachol, displayed longer neurites. The effect of carbachol in astrocytes was due to the activation of M3 muscarinic receptors. Exposure of astrocytes to carbachol increased the expression of the extracellular matrix proteins fibronectin and laminin-1 in these cells. This effect was mediated in part by an increase in laminin-1 and fibronectin mRNA levels and in part by the up-regulation of the production and release of plasminogen activator inhibitor-1, an inhibitor of the proteolytic degradation of the extracellular matrix. The inhibition of fibronectin activity strongly reduced the effect of carbachol on the elongation of all the neurites, whereas inhibition of laminin-1 activity reduced the elongation of minor neurites only. Plasminogen activator inhibitor-1 also induced neurite elongation through a direct effect on neurons. Taken together, these results demonstrate that cholinergic muscarinic stimulation of astrocytes induces the release of permissive factors that accelerate neuronal development.     

Wild mouse lemurs revisit artificial feeding platforms: implications for field experiments on sensory and cognitive abilities in small primates

Dealing effectively with space to find important resources in a natural environment is a fundamental ability necessary for survival. Evidence has already been provided that wild gray mouse lemurs revisit stationary feeding sites regularly. In this study, we explore to what extent two sympatric mouse lemur species, Microcebus murinus and M. ravelobensis, revisited artificial feeding sites during a period of food scarcity. As the tested populations are marked with individual transponders, we built up artificial feeding platforms equipped with a transponder reader at nine different locations where mouse lemurs had been previously caught. We baited them with a liquid reward and recorded the visitors' ID, the time and frequency of their visits, as well as all encounters that occurred on the platforms. Only mouse lemurs visited platforms and a total of sixteen individuals across both species were identified. Mouse lemurs visited a platform with a frequency of 2.02 (+/-0.95, range: 1-3.4) times in a night and they revisited it on several consecutive nights following their first visit (percentage of revisits 90.6%+/-11.7, range: 73.3-100%). First visits on a platform occurred on average 44 min (+/-35; range: 13-131) after sunset. We identified encounters between mouse lemurs on platforms: all of them were agonistic and within a species. Within a dyad, chasers were significantly heavier than chasees (N=7 dyads). Our design of platform experiments offers the advantage of observing wild individually known small primates in their natural environment and of setting up controlled experiments to gain insight into their sensory and cognitive abilities.     

Autofluorescence and Ultrastructure in the Myxomycete Diachea leucopodia (Physarales)

Autofluorescence is reported for the first time in Myxomycete fruiting bodies. Ultrastructure of stalked sporangia of Diachea leucopodia (Didymiaceae, Physarales) was studied using scanning and transmission electron microscopy, energy-dispersive X-ray microanalysis, and fluorescence microscopy. External and internal properties of the peridium that surround the spores and capillitium exhibit autofluorescence. The stalk is composed of calcareous granules and energy-dispersive X-ray microanalysis demonstrates that the elemental composition of the peridium, capillitium, and stalk has varying concentrations of calcium.     

Regeneration processes of a boreal forest in Kamchatka with special reference to the contribution of sprouting to population maintenance

We studied regeneration patterns of three tree species Picea ajanensis, Betula platyphylla and Populus tremula from 1998 to 2000 in the Central Depression of the Kamchatka Peninsula. We paid special attention to the contribution of sprouting to their regeneration. P. ajanensis was the only species that regenerated by seedling. In a 40 × 40 m study plot, the density of P. ajanensis saplings < 2.0 cm in diameter at basal area (DBH) was 1132, and this was the highest among the three species studied. The number of saplings 2 cm in DBH declined sharply with size class. The spatial distribution of P. ajanensis saplings (< 2 cm in DBH) showed a significant positive correlation with that of adult trees and a negative correlation with that of gaps. These trends were not changed after re-measurement in 2000, although nearly half of the juveniles had died or been injured during the two years. These results suggest that small Picea saplings prefer habitats under the canopy of adult trees rather than in gaps for establishment. Most small individuals of B. platyphylla were produced from sprouts. The number of saplings in the smallest size class (< 2 cm in DBH) was much less than that of P. ajanensis, although the number of larger individuals did not decrease remarkably. The spatial distribution of B. platyphylla saplings showed a positive correlation with that of adult trunks and a negative correlation with that of canopy trees of P. ajanensis. These results suggest an effective contribution of sprouts to the regeneration of B. platyphylla. P. tremula was the only species that could invade big gaps and produce many root suckers efficiently. There were 181 suckers of P. tremula in the smallest size class (< 2 cm in DBH) in the study plot, although the number of saplings 2 cm in DBH declined abruptly. The spatial distribution of saplings of this species showed a slight positive correlation with that of gaps, and negative correlation with that of adult trees of B. platyphylla, P. ajanensis, and P. tremula. The root suckering strategy of P. tremula might be adaptive under severe conditions in high-latitude regions. Our data suggest, however, that it does not necessarily contribute to regeneration in mature forests. The three component species in this forest did not seem to utilize canopy gaps for regeneration; we suggest that gap dynamics do not work in this forest. The sparse canopy, which is a typical character of forests in high-latitude regions, might be a consequence of high mortalities of seedlings and root suckers inside gaps.     

Optimization of speed and accuracy of decoding in translation

The speed and accuracy of protein synthesis are fundamental parameters for understanding the fitness of living cells, the quality control of translation, and the evolution of ribosomes. In this study, we analyse the speed and accuracy of the decoding step under conditions reproducing the high speed of translation in vivo. We show that error frequency is close to 10⁻³, consistent with the values measured in vivo. Selectivity is predominantly due to the differences in k_cat values for cognate and near-cognate reactions, whereas the intrinsic affinity differences are not used for tRNA discrimination. Thus, the ribosome seems to be optimized towards high speed of translation at the cost of fidelity. Competition with near- and non-cognate ternary complexes reduces the rate of GTP hydrolysis in the cognate ternary complex, but does not appreciably affect the rate-limiting tRNA accommodation step. The GTP hydrolysis step is crucial for the optimization of both the speed and accuracy, which explains the necessity for the trade-off between the two fundamental parameters of translation.     

Evaluation of the BH3-only Protein Puma as a Direct Bak Activator

Interactions among Bcl-2 family proteins play critical roles in cellular life and death decisions. Previous studies have established the BH3-only proteins Bim, tBid, and Noxa as “direct activators” that are able to directly initiate the oligomerization and activation of Bak and/or Bax. Earlier studies of Puma have yielded equivocal results, with some concluding that it also acts as a direct activator and other studies suggesting that it acts solely as a sensitizer BH3-only protein. In the present study we examined the interaction of Puma BH3 domain or full-length protein with Bak by surface plasmon resonance, assessed Bak oligomerization status by cross-linking followed by immunoblotting, evaluated the ability of the Puma BH3 domain to induce Bak-mediated permeabilization of liposomes and mitochondria, and determined the effect of wild type and mutant Puma on cell viability in a variety of cellular contexts. Results of this analysis demonstrate high affinity (K_D = 26 ± 5 nm) binding of the Puma BH3 domain to purified Bak ex vivo, leading to Bak homo-oligomerization and membrane permeabilization. Mutations in Puma that inhibit (L141E/M144E/L148E) or enhance (M144I/A145G) Puma BH3 binding to Bak also produce corresponding alterations in Bak oligomerization, Bak-mediated membrane permeabilization and, in a cellular context, Bak-mediated killing. Collectively, these results provide strong evidence that Puma, like Bim, Noxa, and tBid, is able to act as a direct Bak activator.     

Insulin Mimetic Effect of Tungsten Compounds on Isolated Rat Adipocytes

Investigations of effective, orally active, and safe antidiabetic metallopharmaceuticals have been carried out during the last two decades. It has been reported that tungsten compounds mimic the action of insulin in intact cell systems. As insulin mimetics, the most investigated tungsten compound was sodium tungstate (ST), rarely investigated was tungstophosphoric acid (WPA), but never alanine complex of tungstophosphoric acid (WPA-A). In this study, the insulin mimetic activity of three different tungsten compounds, ST, WPA, and WPA-A, was evaluated by means of in vitro measurements of the glucose uptake and inhibition of free fatty acids release from epinephrine-treated isolated rat white adipocytes. We investigated the influence of concentration (lower and higher, 0.1 and 1.0 mM, respectively) and solvent: isotonic salt solution—saline (0.9% w/v of NaCl) and dimethyl sulfoxide (DMSO; 2% v/v), on the biological effect of tested compounds. Our experimental data showed that all of the three investigated tungsten compounds possess insulin mimetic activity in vitro on the isolated adipocytes. Influence of concentration and solvents on insulin mimetic effect for the certain tungsten compounds were: WPA was shown effect independently of concentration and solvents; higher concentration and DMSO were significant decreasing insulin mimetic effect of ST; lower concentration and saline led to decreasing effect of WPA-A. Generally, there were no differences in insulin mimetic effect of three tungsten compounds in lower concentration and dissolved in DMSO. When saline was used as solvent, it was needed higher concentration of investigated compounds to accomplish the same effect. In conclusion, our results suggest that low concentration (0.1 mM) of ST, WPA, and WPA-A dissolved in 2% DMSO could be the good candidates for in vivo investigation of their antidiabetic properties.     

Morphometric variability between and within species of Ligophorus Euzet & Suriano, 1977 (Monogenea: Ancyrocephalidae) in the Mediterranean Sea

Monogeneans belonging to the genus Ligophorus Euzet & Suriano, 1977 are parasitic on the gills of several fish species of the family Mugilidae (mullets). At present, 12 species belonging to this genus have been described in the Mediterranean Sea and each host species is infected by a number of Ligophorus species that are not found on other mullets. A multivariate study using principal component and canonical variate analyses was carried out, with the aim of investigating the extent of morphometric variability within and between the 12 Mediterranean species of Ligophorus. Significant intraspecific morphometric variation was detected between two allopatric populations of L. angustus Euzet & Suriano, 1977 parasitic on Chelon labrosus from Sardinia; however, the existence of a new species of Ligophorus needs further evidence. Analysis of interspecific variation confirms the validity of the morphological keys used at present for the identification of Ligophorus at the species level and provides a picture of the relationships between species, indicating that the two species parasitising Mugil cephalus, L. chabaudi Euzet & Suriano, 1977 and L. mugilinus (Hargis, 1955) are the most differentiated. The hypothesis that strict host-parasite ecological relationships may be the result of co-speciation processes is briefly discussed.     

Synthesis, molecular modeling and preliminary biological evaluation of a set of 3-acetyl-2,5-disubstituted-2,3-dihydro-1,3,4-oxadiazole as potential antibacterial, anti-Trypanosoma cruzi and antifungal agents

A series of 3-acetyl-2,5-disubstituted-2,3-dihydro-1,3,4-oxadiazole derivatives was synthesized and their activity screened in vitro against Staphylococcus aureus, Trypanosoma cruzi, and Candida albicans. The bioactivity was expressed as minimum inhibitory concentration (MIC) for S. aureus strains, and as fifty-percent inhibitory concentration (IC(50)) of parasite population growth for T. cruzi. A molecular modeling approach was performed to establish qualitative relationships regarding the biological data and the compounds' physicochemical properties. The 5-(4-OC(4)H(9)Ph, 5l), and 5-(4-CO(2)CH(3)Ph, 5o) derivatives were the most active compounds for S. aureus ATCC 25923 (MIC=1.95-1.25 μg/mL) and T. cruzi (IC(50)=7.91 μM), respectively. Also, a preliminary evaluation against C. albicans involving some compounds was performed and the 5-(4-CH(3)Ph, 5e) derivative was the most active compound (MIC=3.28-2.95 μg/mL). In this preliminary study, all synthesized 3-acetyl-2,5-disubstituted-2,3-dihydro-1,3,4-oxadiazole derivatives were active against all microorganisms tested.     

AQP1 expression analysis in human diseases: implications for proteomic characterization

Aquaporin (AQP)1 belongs to a ubiquitous family of water channel proteins characterized by sequence similarity and the presence of two NPA (Asp-Pro-Ala) motifs existing in almost all organs and tissues. Currently, 13 human AQPs are known and they are divided into two subgroups according to their ability to transport only water molecules, such as AQP1, or also glycerol and other small solutes. The genomic, structural and functional aspects of AQP1 are briefly described. An in-depth discussion is devoted to proteomic approaches that are useful for identifying and characterizing AQP1, mainly through electrophoretic techniques combined with different extraction procedures followed by mass spectrometry analysis. Moreover, the relevance of AQP1 in human diseases is also explained. Its role in human tumors and, in particular, those of the kidney (e.g., clear cell renal carcinoma) is discussed.