Neurosteroids modulate calcium currents in hippocampal CA1 neurons via a pertussis toxin-sensitive G-protein-coupled mechanism

The inhibition of Ca2+ channel currents by endogenous brain steroids was examined in freshly dissociated pyramidal neurons from the adult guinea pig hippocampal CA1 region. The steady-state inhibition of the peak Ca2+ channel current evoked by depolarizing steps from -80 to -10 mV occurred in a concentration-dependent manner with the following IC50 values: pregnenolone sulfate (PES), 11 nM; pregnenolone (PE), 130 nM; and allotetrahydrocorticosterone (THCC), 298 nM. THCC, PE, and PES depressed a fraction of the Ca2+ channel current with a maximal inhibition of 60% of the total current. However, substitution of an acetate group for the sulfate group on PES resulted in a complete loss of activity. Progesterone had no effect (4% inhibition at 100 microM). Intracellular dialysis of PES had no effect on the Ca2+ current; concomitant extracellular perfusion of PES showed normal inhibitory activity, suggesting that the steroid binding site can only be accessed extracellularly. Analysis of tail currents at -80 mV demonstrated that THCC and PES slowed the rate of Ca2+ current activation and deactivation with no change in the voltage dependence of activation. Inhibition of the Ca2+ channel current by THCC and PES was voltage dependent. THCC primarily inhibits the omega-conotoxin (CgTX)-sensitive or N-type Ca2+ channel current. PE was nonselective in inhibiting both the CgTX- and the nifedipine (NIF)-sensitive Ca2+ channel current. These neurosteroids had no effect on the CgTX/NIF-insensitive current. In neurons isolated from pertussis toxin (PTX)-treated animals by chronic intracerebroventricular infusion (1000 ng/24 hr for 48 hr), the Ca2+ channel current inhibition by PES, PE, and THCC was significantly diminished. Intracellular dialysis with GDP-beta-S (500 microM) also significantly diminished the neurosteroid inhibition of the Ca2+ channel current. Intracellular dialysis with the general kinase inhibitors H-7 (100 microM), staurosporine (400 nM), and a 20 amino acid protein kinase inhibitor (1 microM) also significantly prevented the THCC and PES inhibition of the Ca2+ channel current. Intracellular dialysis with the more specific inhibitors of protein kinase C (PKC), the pseudosubstrate inhibitor (PKCI 19-36) (1-2 microM) and bisindolylmaleimide (1 microM) significantly diminished the THCC and PE inhibition of the Ca2+ channel current. Rp- cAMP (100 microM), a specific inhibitor of cAMP-dependent protein kinase (PKA), had no effect on the THCC and PE inhibition of the Ca2+ current.(ABSTRACT TRUNCATED AT 400 WORDS)     

Estimation and hypothesis testing of treatment effects in animal reproductive toxicology studies

Healy (1) and Dempster et al. (8) proposed statistical methods to evaluate the treatment effects in animal reproductive toxicology research. Both methods assume homogeneous variance for the dams and the pups, respectively, in all the treatment groups. In this paper, via mixed effect modeling, we propose a method to estimate the treatment effects allowing heterogeneous variances for the dams and the pups, respectively, in different treatment groups. Covariates will also be included in the model. A procedure to test the fixed effects is also discussed. An example from an animal reproductive toxicological study is used to illustrate the procedures.     

Ultrastructural immunocytochemical localization of neurotensin and the dopamine D2 receptor in the rat nucleus accumbens

The neuroleptic-like effects of neurotensin (NT) are thought to be due to interactions with dopamine (DA) acting primarily at D2 receptors within the nucleus accumbens septi (Acb). Using electron microscopic dual labeling immunocytochemistry, we sought to demonstrate cellular substrates for functional interactions involving NT and DA D2 receptors in the adult rat Acb. Peroxidase reaction product representing D2 receptor-like immunoreactivity (D2-LI) was seen along membranes of Golgi lamellae and multivesicular bodies of perikarya containing immunogold labeling representing NT-LI. Dually labeled somata usually contained highly indented nuclei, a characteristic of aspiny neurons. Dendrites also occasionally colocalized the two immunomarkers. Other somata, dendrites, and all axon terminals were singly labeled with either NT-LI or D2-LI. In distinct sets of terminals, NT-LI was commonly associated with large, dense-cored vesicles, whereas D2-LI was found along the plasmalemma and over nearby small clear vesicles. Each type of terminal comprised approximately 20% of synaptic input to NT-immunoreactive dendrites. Similar proportions of terminals containing NT-LI or D2-LI contacted unlabeled (approximately 55%) or NT-labeled (approximately 35%) dendrites and, occasionally, were observed converging onto common dendrites. Terminals containing NT-LI or D2-LI also were often closely apposed. These findings provide the first ultrastructural evidence that: (1) NT and D2 receptors are colocalized in aspiny neurons and dendrites, (2) NT may produce a direct postsynaptic effect on neurons receiving input from terminals which are presynaptically modulated by DA via D2 receptors, and (3) NT and DA acting at D2 receptors may interact through presynaptic modulation of common axon terminals.     

Zollinger-Ellison syndrome and antral G-cell hyperfunction in patients with resistant duodenal ulcer disease

We measured basal and pentagastrin-stimulated acid secretion, as well as basal and meal-stimulated plasma gastrin concentration to determine, in 67 patients affected by resistant duodenal ulcer, whether their condition could be related to gastric acid secretion and/or gastrin-related syndromes. We then compared them to 46 duodenal ulcer control patients. The outpatients were investigated consecutively. The resistant duodenal ulcer patients differed from the controls only in their higher complication rates (bleeding or perforation, P < 0.05). We identified five patients in the resistant duodenal ulcer group with Zollinger-Ellison syndrome and 12 with antral G cell hyperfunction, whereas in the control group only one patient was affected by antral G cell hyperfunction. IgG anti-Helicobacter pylori antibodies were positive for the presence of infection in 7 of the hypergastrinaemic patients. When Zollinger-Ellison syndrome or antral G cell hyperfunction were excluded, no differences could be found in gastric acid secretion, or basal and meal-stimulated plasma gastrin levels, between the resistant and control duodenal ulcer patients, except for basal acid hypersecretion (resistant duodenal ulcer 16% vs duodenal ulcer 2% P = 0.0144). In the presence of duodenal ulcer disease resistant to H2-blockers, it is mandatory to measure basal plasma gastrin concentration since it was possible to diagnose the gastrin-related syndromes, Zollinger-Ellison syndrome and antral G cell hyperfunction, in 26% of this group of patients.     

The interaction of dexamethasone with ondansetron on drug-induced emesis in the ferret

The characteristics of the five Korean isolates of Japanese encephalitis (JE) virus were compared with those of the already reported JE virus strains from Japan and China using the hemagglutination test and polymerase chain reaction direct sequencing of the JE virus genomes (capsid/premembrane, envelope region). The hemagglutination patterns of all the isolates were distinctly different from the Nakayama-NIH strain. The optimal pH of hemagglutination of all the Korean isolates was 6.6-7.0 and the reaction range was broader than that of the Nakayama-NIH strain. The 198 nucleotide sequences in the capsid/premembrane gene region of the five Korean strains indicated that they were classified into the third genotype group, the JE strains from the countries in the temperate zone including the Nakayama-NIH, JaOArS982, and Beijing-1 strains. Four of the five Korean isolates formed a unique phylogenetic tree within the third genotype group, although the last one was genetically highly related to the Nakayama-NIH strain. The 251 nucleotide sequences in the envelope region of the five isolates were more divergent than the capsid/premembrane region. Four of the five isolates showed a large nucleotide divergency as compared with the JaOArS982 strain (< or = 12.4%), but the last one was similar to the JaOArS982 strain (98% of nucleotide homology). These results suggest the evolutionary divergence of the JE viruses isolated in Korea from the Japanese and Chinese strains and that there may exist at least two antigenically different JE virus strains in Korea.     

Local mobility within villin 14T probed via heteronuclear relaxation measurements and a reduced spectral density mapping

Villin 14T, a representative domain from the actin severing and bundling protein villin, binds calcium ions and actin monomers. To begin to understand the contributions of mobility to the villin-calcium and villin-actin interactions, relaxation rates for magnetization involving the amide nitrogens and protons have been measured for 15N-labeled villin 14T in solution. Although we have measured the complete set of rates required for a full spectral density map, difficulties in the accurate measurement of relaxation rates for antiphase coherence and two-spin order led us to consider a reduced mapping formalism. From the reduced spectral density map, a model-free analysis, or directly from the measured Nx,y relaxation rates, local variations in mobility along the backbone of villin 14T have been revealed. Fast motions are evident not only at the amino and carboxyl termini but also in the turn between strands beta 4 and beta 5 of the central beta-sheet and in the turn between helix alpha 3 and strand beta 7. Slower motions are suggested for the turn between strands beta 2 and beta 3. Motions on the microsecond to millisecond time scale have been probed directly by examining the dependence of the proton transverse relaxation rate on the spin-locking field strength. Leu11 shows a strong dependence on field strength, implying conformational exchange with a time constant of 125 +/- 69 microseconds. The backbone at the actin-binding interface appears to be rather rigid.     

Production and acceleration of ion beams by laser ablation

In this work, we present a new pulsed laser ablation technique to obtain energetic ion beams. The accelerator we made is a compact device able to extract and accelerate the ionic components of plasma up to 160 keV per charge state. It is composed by a generating chamber containing an expansion chamber used like first electrode. Next, a second electrode connected to ground and a third electrode connected to negative voltage are used. The third electrode is used also as Faraday cup. By the analysis of the ion signals we studied the plume parameters such as TOF accelerated signals, charge state, and divergence.     

Processing step-related upgrading of silicon-based solar cells detected by photoluminescence spectroscopy

The aim of this work was to study the effect of solar cell processing steps on the electrical quality of the material, using photoluminescence spectroscopy as a non-contact technique. It was confirmed that consecutive steps in solar cell processing not only lead to electronic quality upgrading in Ribbon and multicrystalline silicon wafers, but also to the introduction of recombining defects. Furthermore, the importance and usefulness of photoluminescence technique on the overall improvement of the substrate material is discussed.     

Insect antifeedant activity associated with compounds isolated from species ofLonchocarpus andTephrosia

The antifeedant activity of a series of 21 chalcones, flavanes, and flavanones isolated from the generaLonchocarpus andTephrosia (Leguminosae) was assessed by behavioral and electrophysiological bioassays against larvae ofSpodoptera littoralis andS. exempta. The antifeedant activity is related to the molecular structure of the compounds, and possible modes of interaction with the insect taste receptors are discussed.