Light-induced changes in energy metabolites, guanine nucleotides, and guanylate cyclase within frog retinal layers

Freeze-dried sections were prepared from retinas of frogs which were dark-adapted or exposed to varying periods of light. Samples of the discrete layers were dissected, weighed, and analyzed for energy metabolites, guanylate compounds, and the enzyme guanylate cyclase. ATP and P-creatine were measured in both dark- and light-adapted retinas. There was a gradient in ATP and P-creatine levels in dark-adapted retinas, with the lower concentrations in the photoreceptors, and increasing concentrations in the inner retina. After light adaptation, concentrations increased, an observation which supports the concept that transmitter release occurs in the dark and ceases in the light. The sum of GTP plus GDP, GDP, and cyclic GMP were analyzed in dark-adapted retinas and after exposure to 2 min or 2 h of room light. GDP was rather uniformly distributed in the retinal layers, was increased by 2 min of light in all layers but the outer nuclear, and remained elevated at 2 h in the inner retina. GTP values showed a marked localization in the outer nuclear layer, which increased after 2 min or 2 h of illumination; in all other layers GTP was decreased by light. Cyclic GMP in the dark was highest in the photoreceptor cells, decreasing to one-third after 2 min of light; there were significant increases in the outer plexiform and inner nuclear layers at this time. Cyclic GMP remained low in the photoreceptor cells even after 2 h of light, while the inner layers returned to dark values. Guanylate cyclase, like cyclic GMP, was largely confined to the photoreceptor cells and showed a maximal increase after 2 min of light exposure.     

Changes in fluid compartments and ionic composition in the isolated chick retina during SD

Total content of water, extracellular space (ES), Na⁺, K⁺, and Cl^– in the isolated chick retina were measured in the presence (test) or absence (control) of spreading depression (SD). During SD in medium with 0.5 mM or 2 mM MgSO₄, there is an increase in the intracellular concentration of Na⁺ and Cl^– and a decrease in the intracellular concentration of K⁺. A decrease in the ES was only found in the medium with 2 mM MgSO₄ together with a diminshed outmovement of K⁺. We suggest that a decrease in the ES is due to an increased absorption of K⁺ by the Muller cells, causing its swelling and consequently a decrease of the ES.The addition of sucrose (17 mM) to the incubation medium as the extracellular marker markedly decreased the intracellular concentration of Cl^– in control retinas, blocked the inward movement of this ion to the tissue during SD and also changed the K⁺ movement during the phenomenon in medium with 2 mM MgSO₄. We suggest that Cl^– is an important ion in the ionic balance of the Muller cells and that sucrose must have its site of action that these cells.     

QSAR modeling and transmission electron microscopy stereology of altered mitochondrial ultrastructure of white blood cells in patients diagnosed as schizophrenic and treated with antipsychotic drugs.

Treatment of patients diagnosed as schizophrenic with antipsychotic drugs (neuroleptics) is known to cause occasional unexplained depletion of white blood cells, especially neutrophil granulocytes. It has been known for many years that neuroleptics can interfere with the mitochondrial respiratory chain in vitro. Because there has been a growing interest recently in mitochondrial targeting of drugs, and since a quantitative structure-activity relationship (QSAR) model that predicts mitochondrial accumulation of neuroleptics has been published, we investigated the effects of neuroleptics on white blood cell mitochondria. Venous blood samples were collected from both patients undergoing treatment with neuroleptics and healthy volunteers. The samples were processed for transmission electron microscopy. The resulting images of white blood cells were analyzed using stereology to compare quantitatively mitochondrial morphology in the patient and control groups. We found that in patients, but not in controls, there was swelling of mitochondria and fragmentation of the mitochondrial cristae. There also were fewer mitochondria in patients than in controls, although due to the swelling of the organelles, the volume density of mitochondria in the two groups was not significantly different. Such changes are typical of a toxic insult. Consequently, it seems plausible that, since schizophrenia is not a disease considered to affect white blood cells per se, these changes probably are due to the medication.     

The role of exopolymers in the attachment of sphingomonas paucimobilis

The importance of exopolymers in the adhesion of Sphingomonas paucimobilis was established by studying the attachment to glass of three mutants with defective gellan production. The attachment assays were performed in either phosphate buffered saline (controls) or in the exopolymeric solutions produced by the mutants. The exopolymer was found to have surface active properties, changing the glass surface from hydrophilic to hydrophobic, making adhesion thermodynamically favourable. Only the cells that had a substantial polymeric layer surrounding their walls were able to significantly colonise glass coated with the exopolymer. It is hypothesised that the exopolymer bound to the glass and the exopolymer present at the surface of the bacteria bound together, overcoming the energy barrier created by the negative charge of both surfaces. It is concluded that the exopolymer from S. paucimobilis has a dual role in the process of adhesion by both coating the surface thereby strengthening adhesion and by enhancing adhesion through the establishment of polymeric bridges.     

Discrimination of bacteriophage infected cells using locked nucleic acid fluorescent in situ hybridization (LNA-FISH)

Bacteriophage–host interaction studies in biofilm structures are still challenging due to the technical limitations of traditional methods. The aim of this study was to provide a direct fluorescence in situ hybridization (FISH) method based on locked nucleic acid (LNA) probes, which targets the phage replication phase, allowing the study of population dynamics during infection. Bacteriophages specific for two biofilm-forming bacteria, Pseudomonas aeruginosa and Acinetobacter, were selected. Four LNA probes were designed and optimized for phage-specific detection and for bacterial counterstaining. To validate the method, LNA-FISH counts were compared with the traditional plaque forming unit (PFU) technique. To visualize the progression of phage infection within a biofilm, colony-biofilms were formed and infected with bacteriophages. A good correlation (r = 0.707) was observed between LNA-FISH and PFU techniques. In biofilm structures, LNA-FISH provided a good discrimination of the infected cells and also allowed the assessment of the spatial distribution of infected and non-infected populations.     

Morphogenesis control in Candida albicans and Candida dubliniensis through signaling molecules produced by planktonic and biofilm cells

Morphogenesis control by chemical signaling molecules is beginning to be highlighted in Candida biology. The present study focuses on morphogenic compounds produced in situ by Candida albicans and Candida dubliniensis during planktonic and biofilm growth that may at least partially substantiate the effect promoted by supernatants in morphogenesis. For both species, planktonic versus biofilm supernatants were analyzed by headspace-solid-phase microextraction and gas chromatography-mass spectrometry. Both planktonic cells and biofilm supernatants of C. albicans and C. dubliniensis contained isoamyl alcohol, 2-phenylethanol, 1-dodecanol, E-nerolidol, and E,E-farnesol. Alcohol secretion profiles were species, culture mode, and growth time specific. The addition of exogenous alcohols to the cultures of both species inhibited the morphological transition from the yeast to the filamentous form by up to 50%. The physiological role of these alcohols was put to evidence by comparing the effects of a 96-h cultured supernatant with synthetic mixtures containing isoamyl alcohol, 2-phenylethanol, E-nerolidol, and E,E-farnesol at concentrations determined herein. All synthetic mixtures elicited a morphological effect similar to that observed for the corresponding supernatants when used to treat C. albicans and C. dubliniensis cultures, except for the effect of the 96-h C. dubliniensis planktonic supernatant culture on C. albicans. Overall, these results reveal a group of alcohol extracellular signaling molecules that are biologically active with C. albicans and C. dubliniensis morphogenesis.     

Expression of a fungal hydrophobin in the Saccharomyces cerevisiae cell wall: effect on cell surface properties and immobilization

The aim of this work was to modify the cell surface properties of Saccharomyces cerevisiae by expression of the HFBI hydrophobin of the filamentous fungus Trichoderma reesei on the yeast cell surface. The second aim was to study the immobilization capacity of the modified cells. Fusion to the Flo1p flocculin was used to target the HFBI moiety to the cell wall. Determination of cell surface characteristics with contact angle and zeta potential measurements indicated that HFBI-producing cells are more apolar and slightly less negatively charged than the parent cells. Adsorption of the yeast cells to different commercial supports was studied. A twofold increase in the binding affinity of the hydrophobin-producing yeast to hydrophobic silicone-based materials was observed, while no improvement in the interaction with hydrophilic carriers could be seen compared to that of the parent cells. Hydrophobic interactions between the yeast cells and the support are suggested to play a major role in attachment. Also, a slight increase in the initial adsorption rate of the hydrophobin yeast was observed. Furthermore, due to the engineered cell surface, hydrophobin-producing yeast cells were efficiently separated in an aqueous two-phase system by using a nonionic polyoxyethylene detergent, C(12-18)EO(5).     

Yeast communities associated with sugarcane in Campos, Rio de Janeiro, Brazil.

Yeast communities associated with sugarcane leaves, stems and rhizosphere during different phases of plant development were studied near Campos, in Rio de Janeiro, Brazil. Atmospheric temperature, soil granulometry and pH, and sugar cane juice degree Brix and pH were determined. Yeast communities associated with sugarcane were obtained after cellular extraction by shaking, blending and shaking plus sonication, and cultured on Yeast Nitrogen Base Agar plus glucose (0.5%) and Yeast Extract-Malt Extract Agar. No significant differences in yeast counts were found among the cellular extraction treatments and culture media. 230 yeast cultures were identified according to standard methods, and distinct yeast communities were found for each substrate studied. The prevalent species isolated from sugarcane were Cryptococcus laurentii, Cryptococcus albidus, Rhodotorula mucilaginosa and Debaryomyces hansenii.     

4α‐Acetoxyamijidictyol – A New Antifeeding Dolastane Diterpene from the Brazilian Brown Alga Canistrocarpus cervicornis

Chemical investigation of the CH₂Cl₂ crude extract from the brown alga Canistrocarpus cervicornis (Dictyotaceae) led to isolation of one new ( 1 ) and four previously reported dolastane diterpenes ( 2 – 5 ). Their structures were characterized by 1D‐ and 2D‐NMR spectroscopic techniques, including a full single crystal X‐ray diffraction analysis for 1, 2 , and 4 . In addition, the new structure 1 was assayed as chemical defense inhibiting the feeding by the sea urchin Lytechinus variegatus. This study constitutes an additional report broadening the known spectrum of action and defensive roles of secondary metabolites of the C. cervicornis and Dictyotales species.