The surface free energy of Leishmania mexicana amazonensis.

Surface charge of Leishmania mexicana amazonensis was investigated by direct zeta-potential determination and ultrastructural cytochemistry, and its surface tension was studied by measurements of the advancing contact angle formed by the parasite monolayers with drops of liquids of different polarities. Both virulent and avirulent promastigotes exhibited negatively charged surfaces with a zeta-potential of about -15 mV. Treatment of these cells with trypsin, alkaline phosphatase, or phospholipase C rendered their surfaces less negatively charged, whereas neuraminidase did not alter the parasite negativeness. Cytochemically, we could observe a reduction in the cationized ferritin binding after the parasite treatment with each of the former enzymes, but not with neuraminidase. The surface free energy of parasites was calculated by taken to account the London dispersion, the Keeson dipole-dipole, and the Debye dipole-induced forces, as well as the surface polarity of the parasites and their zeta-potentials, by considering their adhesion to polystyrene surfaces. The delta G values of -6.4 and -18.1 mJ.m-2 were obtained for avirulent and virulent promstigotes, respectively.     

Internalization of surface anionic sites and phagosome-lysosome fusion during interaction of Toxoplasma gondii with macrophages

The participation of cell surface anionic sites on the interaction between tachyzoites of Toxoplasma gondii and macrophages and the process of phagosome-lysosome fusion were analyzed using cationized ferritin as a marker of cell surface anionic sites and albumin-colloidal gold as a marker for secondary lysosomes. Incubation of either the macrophages or the parasites with cationized ferritin before the interaction increased the ingestion of parasites by macrophages. Anionic sites of the macrophage's surface, labeled with cationized ferritin before the interaction, were internalized together with untreated parasites. However, after interaction with glutaraldehyde-fixed or specific antibody-coated parasites, the cationized ferritin particles were observed in endocytic vacuoles which did not contain parasites. Macrophages previously labeled with albumin-gold at 37 degrees C, were incubated in the presence of cationized ferritin at 4 degrees C and then incubated with untreated or specific antibody-coated parasites. After interaction with opsonized parasites, the colloidal gold particles were observed in the parasitophorous vacuoles while the cationized ferritin particles were observed in cytoplasmic vesicles. However, when the interaction was carried out with untreated parasites, the parasitophorous vacuoles exhibited ferritin particles while the colloidal gold particles were observed in cytoplasmic vesicles. These observations, in association with studies previously reported, suggest that the state of the parasite surface determines the mechanism of parasite entry into the macrophage, the composition of the membrane lining the parasitophorous vacuole and the ability of lysosomes to fuse with the vacuoles.     

Increased Neostriatal Tyrosine Hydroxylation During Stress: Role of Extracellular Dopamine and Excitatory Amino Acids

Abstract: We examined the regulation of neostriatal tyrosine hydroxylation during acute stress, testing the hypothesis that excitatory amino acids (EAAs) contribute to the stress-evoked increase in dopamine (DA) synthesis. Dialysis probes implanted into neostriatum permitted delivery of drugs and sampling of extracellular fluid. Rats were exposed to 30 min of intermittent tail shock during infusion of an inhibitor of aromatic amino acid decarboxylase (AAAD), NSD-1015 (100 µM), and DOPA was measured in the dialysate. Tail shock was applied beginning either 15 min after the onset of NSD-1015 treatment (the initial rate of DOPA accumulation) or 75 min after the onset of treatment (when DOPA had approached steady state). Tail shock increased the steady-state levels of extracellular DOPA in neostriatum (+40%). However, there was no change in the initial rate of DOPA accumulation unless animals also received the D₂ receptor antagonist eticlopride (50 nM), in which case an increase was observed (+228%). The impact of tail shock on the steady-state level of DOPA was attenuated by the D₂ agonist quinpirole (100 µM), or by 2-amino-5-phosphonovalerate (APV) (100 µM) or 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) (100 µM), EAA antagonists acting at NMDA or d ,l -α-amino-3-hydroxy-5-methyl-4-isoxazole-4-propionate (AMPA) receptors, respectively. These data suggest that acute stress normally has little effect on tyrosine hydroxylation in neostriatum due to the inhibitory influence of DA in the extracellular fluid. However, when that influence is absent (e.g., during extended inhibition of DOPA decarboxylation or blockade of DA receptors), stress increases tyrosine hydroxylation via EAAs acting on NMDA and AMPA receptors. Thus, EAAs released from corticostriatal projections may stimulate DA synthesis and thereby restore dopaminergic activity under conditions in which the availability of DA for release has been compromised.     

Two Components of Glutamate Exocytosis Differentially Affected by Presynaptic Modulation

Abstract: The total Ca²⁺-dependent release of glutamate induced by depolarization of cerebrocortical nerve terminals with KCl was analyzed into a fast and a slow component. The fast component exhibited a decay time of <1 s and accounted for 0.95 ± 0.10 nmol of glutamate, whereas the slow component, which exhibited a decay time of 52 ± 7 s, accounted for the release of 2.48 ± 0.19 nmol of glutamate. These two components were differentially affected by the Ca²⁺ chelator BAPTA, the divalent cation Sr²⁺, or the botulinum neurotoxin A. The adenosine A₁ receptor agonist N ⁶-cyclohexyladenosine strongly reduced the fast component without altering the slow component. In contrast, the inhibitory effect of arachidonic acid and the facilitatory action of the metabotropic glutamate receptor agonist (1 S ,3 R )-1-aminocyclopentane-1,3-dicarboxylic acid were observed as a decrease and an increase, respectively, in the two components. It is concluded, first, that the fast and slow components correspond to the release of docked and mobilized vesicles, respectively, and second, that presynaptic modulation more significantly alters the fast component of release.     

Cyclic beta-(1,2)-glucan synthesis in Rhizobiaceae: roles of the 319-kilodalton protein intermediate.

Cyclic beta-(1,2)-glucans are synthesized by members of the Rhizobiaceae family through protein-linked oligosaccharides as intermediates. The protein moiety is a large inner membrane molecule of about 319 kDa. In Agrobacterium tumefaciens and in Rhizobium meliloti the protein is termed ChvB and NdvB, respectively. Inner membranes of R. meliloti 102F34 and A. tumefaciens A348 were first incubated with UDP-[14C]Glc and then solubilized with Triton X-100 and analyzed by polyacrylamide gel electrophoresis under native conditions. A radioactive band corresponding to the 319-kDa protein was detected in both bacteria. Triton-solubilized inner membranes of A. tumefaciens were submitted to native electrophoresis and then assayed for oligosaccharide-protein intermediate formation in situ by incubating the gel with UDP-[14C]Glc. A [14C]glucose-labeled protein with an electrophoretic mobility identical to that corresponding to the 319-kDa [14C]glucan protein intermediate was detected. In addition, protein-linked radioactivity was partially chased when the gel was incubated with unlabeled UDP-Glc. A heterogeneous family of cyclic beta-(1,2)-glucans was formed upon incubation of the gel portion containing the 319-kDa protein intermediate with UDP-[14C]Glc. A protein with an electrophoretic behavior similar to the 319-kDa protein intermediate was "in gel" labeled by using Triton-solubilized inner membranes of an A. tumefaciens exoC mutant, which contains a protein intermediate without nascent glucan. These results indicate that initiation (protein glucosylation), elongation, and cyclization were catalyzed in situ. Therefore, the three enzymatic activities detected in situ reside in a unique protein component (i.e., cyclic beta-(1,2)-glucan synthase). It is suggested that the protein component is the 319-kDa protein intermediate, which might catalyze the overall cyclic beta-(1,2)-glucan synthesis.     

Effects of brain ischemia on intermediate filaments of rat hippocampus

Neurofilaments subunits (NF-H, NF-M, NF-L) and glial fibrillary acidic protein (GFAP) were investigated in the hippocampus of rats after distinct periods of reperfusion (1 to 15 days) following 20 min of transient global forebrain ischemia in the rat. In vitro [¹⁴Ca]leucine incorporation was not altered until 48 h after the ischemic insult, however concentration of intermediate filament subunits significantly decreased in this period. Three days after the insult, leucine incorporation significantly increased while the concentration NF-H, NF-M, and NF-L were still diminished after 15 days of reperfusion. In vitro incorporation of³²P into NF-M and NF-L suffered immediately after ischemia, but returned to control values after two days of reperfusion. GFAP levels decreased immediately after ischemia but quickly recovered and significantly peaked from 7 to 10 days after the insult. These results suggest that transient ischemia followed by reperfusion causes proteolysis of intermediate filaments in the hippocampus, and that proteolysis could be facilitated by diminished phosphorylation levels of NF-M and NF-L.     

Changes in the lower premolar-size sequence during hominid evolution. Physlogenetic implications

The size relationship between the crown area of the lower canines (Cs), third (P3s) and fourth premolars (P4s), and first molars (M1s) in hominids is examined by means of the regression analysis. The lower P3 seem to be under the influence of those factors that control both the size of the anterior and posterior teeth, and the P4:P3 size ratio is related to the relative size of the anterior and posterior dentitions. So, the P4>P3 sequence is associated with the megadontia and hipermegadontia of the posterior teeth, whereas the expansion of the anterior teeth produces the P3>P4 sequence. We consider the P4:P3 size ratio as an excellent indicator of the taxonomic and philogenetic status of fossil hominids.     

Variable influence of ferric and cupric ions on Saccharomyces cerevisiae strains used in asymmetric organic synthesis

Summary Fe (III) and Cu (II) each at 50 M in four commercial strains of Saccharomyces cerevisiae induced an increase of NAD(P)⁺ reduction in one strain (Turkish), but two others (Chilean and Brazilian), the presence of Fe(III) and/or Cu(II) diminished NAD(P)⁺ reduction presumably due to free radicais formation inside these living cells. Suprisingly, in the American strain, Fe(III) induced a decrease and Cu (II) an increase of NAD(P)⁺ reduction.     

A population analysis of Robertsonian and Ag-NOR polymorphisms in brown trout (Salmo trutta)

An analysis of Robertsonian polymorphism and variation in the number of active NORs has been carried out in several populations of brown trout (Salmo trutta) from Northwestern Spain. The karyotype of this species appears to be soundly established, and essentially no variation has been found in chromosome number. Interindividual and interpopulation variation in arm number was detected, with figures ranging between 100 and 102 among individuals, and between 100.10 and 100.80 among populations. This variation in arm number is solely attributable to the polymorphism of the short arm of the main NOR-bearing pair 11, which can appear from acrocentric to metacentric in different individuals. Most populations analyzed showed the standard distribution of active NORs previously observed in this species. The Miño drainage basin, and specially the Chamoso population, showed a multi-chromosomal distribution of active NORs, with several new locations, always telomeric. In most cases no concordance was observed between previously detected rDNA sites in S. trutta and the new Ag-NOR locations. This fact suggests a transposition mechanism rather than an activation of silent rDNA sites to explain this multichromosomal NOR pattern.