Determinants of frequency-dependent regulation of Kv1.2-containing potassium channels

Voltage-gated potassium channels are important regulators of electrical excitation in many tissues, with Kv1.2 standing out as an essential contributor in the CNS. Genetic deletion of Kv1.2 invariably leads to early lethality in mice. In humans, mutations affecting Kv1.2 function are linked to epileptic encephalopathy and movement disorders. We have demonstrated that Kv1.2 is subject to a unique regulatory mechanism in which repetitive stimulation leads to dramatic potentiation of current. In this study, we explore the properties and molecular determinants of this use-dependent potentiation/activation. First, we examine how alterations in duty cycle (depolarization and repolarization/recovery times) affect the onset and extent of use-dependent activation. Also, we use trains of repetitive depolarizations to test the effects of a variety of Thr252 (S2-S3 linker) mutations on use-dependent activation. Substitutions of Thr with some sterically similar amino acids (Ser, Val, and Met, but not Cys) retain use-dependent activation, while bulky or charged amino acid substitutions eliminate use-dependence. Introduction of Thr at the equivalent position in other Kv1 channels (1.1, 1.3, 1.4), was not sufficient to transfer the phenotype. We hypothesize that use-dependent activation of Kv1.2 channels is mediated by an extrinsic regulator that binds preferentially to the channel closed state, with Thr252 being necessary but not sufficient for this interaction to alter channel function. These findings extend the conclusions of our recent demonstration of use-dependent activation of Kv1.2-containing channels in hippocampal neurons, by adding new details about the molecular mechanism underlying this effect.     

Effect of different levels of concentrate on ruminal microorganisms and rumen fermentation in Nellore steers

The aim of this study was to investigate the effect of different dietary levels of concentrate on feed intake, digestibility, ruminal fermentation and microbial population in steers. Eight Nellore steers fitted with ruminal cannulas were used in a double 4 × 4 Latin square design experiment. The dietary treatments consist of four different proportions of concentrate to roughage: 30:70, 40:60, 60:40 and 80:20% in the dry matter, resulting in Diets 30, 40, 60 and 80, respectively. The roughage was corn silage, and the concentrate was composed of corn, soybean meal and urea. Apparent digestibility of organic matter and crude protein showed a linear association with concentrate proportion (p = 0.01), but the increased concentrate levels did not affect the digestibility of fibre. The lowest ruminal pH-values were observed in animals fed with Diet 80, remaining below pH 6.0 from 6 h after feeding, while in the other diets, the ruminal pH was below 6.0 not before 12 h after feeding. After feeding Diet 80, the ammonia concentration in the rumen was significantly the highest. Higher dietary concentrate levels resulted in a linear increase of propionic acid concentrations, a linear reduction of the ratio acetic acid to propionic acid (p < 0.01) and a linear increased synthesis of microbial nitrogen (p < 0.001). The predicted production of methane was lower in diets with greater amounts of concentrate (p = 0.032). The population of methanogens, R. flavefaciens and R. albus decreased with higher concentrate levels, while the population of S. ruminantium increased (p < 0.05). The results indicate that greater amounts of concentrate do not decrease ruminal pH-values as much as expected and inhibit some cellulolytic bacteria without impairing the dry matter intake and fibre digestibility in Nellore steers.     

A Cross-Species Analysis Reveals a General Role for Piezo2 in Mechanosensory Specialization of Trigeminal Ganglia from Tactile Specialist Birds

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Chaperone Effects on Prion and Nonprion Aggregates

Exposure to high temperature or other stresses induces a synthesis of heat shock proteins. Many of these proteins are molecular chaperones and some of them help cells to cope with heat-induced denaturation and aggregation of other proteins. In the last decade, chaperones have received increased attention in connection with their role in maintenance and propagation of the Saccharomyces cerevisiae prions, infectious or heritable agents transmitted at the protein level. Recent data suggest that functioning of the chaperones in reactivation of heat-damaged proteins and in propagation of prions is based on the same molecular mechanisms but may lead to different consequences depending on the type of aggregate. In both cases the concerted and balanced action of “chaperones'' team,” including Hsp104, Hsp70, Hsp40 and possibly other proteins, determines whether a misfolded protein is to be incorporated into an aggregate, rescued to the native state or targeted for degradation.Key Words: Amyloid, Hsp40, Hsp70, Hsp104, stress response, yeast     

CRISPR/Cas9-mediated gfp gene inactivation in Arabidopsis suspension cells

Molecular Biology Reports - Targeted genome editing using CRISPR/Cas9 is a promising technology successfully verified in various plant species; however, it has hardly been used in plant cell...     

Radioactive contamination in the upper part of the Techa river: stirring-up of bottom sediments and precipitation of suspended particles

A hydrodynamic model of the upper part of the Techa river was developed on the basis of the river valley geometry as well as data of hydrological conditions and of the granulometric composition of bottom sediments. The model describes the transport of radioactivity by suspended sediments with different granulometric compositions (clay, silt) in the early 1950s. It includes the stirring-up of bottom sediments and the precipitation of suspended sediments as a function of water discharge rate and water level in the investigated part of the river. The results allow to specify the development of the river system contamination as a result of inflow of suspended sediments contaminated with radionuclides. In the period of liquid radioactive waste (LRW) discharges, the water of the Techa river contained a large fraction of finely dispersed particles of less than 5 micro m diameter. At the site of LRW discharge 80% of the discharged activity was adsorbed to these particles. Depending on the water flow, 40-80% of the suspensions precipitated at the bottom of subsequent sedimentation reservoirs. A total of about 1.6 MCi adsorbed to the suspended particles entered the open hydrographic system of the Techa river. The conclusion that the largest part of the activity was adsorbed on the suspended particles contradicts the assumption in the Techa river dosimetry system, TRDS-2000, that most of the released activity entered the Techa river in soluble form. For a correct reconstruction of the doses received by the Techa river population it is, therefore, essential to consider hydrodynamic models that take into account the transport of radionuclides adsorbed on the suspended particles.     

Network analysis of potential migration routes for Oriental White Storks (<Emphasis Type="Italic">Ciconia boyciana</Emphasis>)

From 1998 through to 2000, we satellite-tracked the movements of 13 Oriental White Storks (Ciconia boyciana) on their autumnal migration in order to identify their important stopover sites for preserving links from the Russian Far East breeding sites to the wintering sites in south-eastern China. New analytical methods of satellite tracking data were employed to derive robust information on the locations of stay sites, the number of stopovers made during migration, and the distance traveled without making stopovers. Based on the derived information, we modeled a stay site network as an abstraction of the storks potential migration routes from their breeding sites to wintering sites. Using network analysis techniques, we explored how the loss of stopover sites could affect the connectivity of potential migration routes. The results suggested that if the seashore stopover sites facing Bohai Bay in eastern China were lost, the storks wintering sites along the Yangtze River in south-eastern China would be isolated. Among the seashore stopover sites, Jiantuozhi Gley Mire (39.185°N, 118.627°E), located on the northern seashore of Bohai Bay, was considered particularly important for migrating storks, because it was used every year by the storks we tracked. If conservation needs of this critically located site fail to be addressed, the stay site network of storks can create weak links in the chain of migration and, if broken, storks will have great difficulties in completing their autumnal migration.     

Neurotrophin action on a rapid timescale

Mechanisms underlying the fast action of neurotrophins include intracellular Ca(2+) signaling, neuronal excitation, augmentation of synaptic excitation by modulation of N-methyl-d-aspartate receptor activity and control of synaptic inhibition through the regulation of the K(+)-Cl(-) cotransporter KCC2. The fastest action of brain-derived neurotrophic factor and neurotrophin-4/5 occurs within milliseconds, and involves activation of TrkB and the opening of the Na(+) channel Na(v)1.9. Through these rapid actions, neurotrophins shape neuronal activity, modulate synaptic transmission and produce instructive signals for the induction of long-term changes in the efficacy of synaptic transmission.     

Role of protein phosphatases in hypoxic preconditioning

To find a protein kinase C (PKC)-independent preconditioning mechanism, hypoxic preconditioning (HP; i.e., 10-min anoxia and 10-min reoxygenation) was applied to isolated rat hearts before 60-min global ischemia. HP led to improved recovery of developed pressure and reduced end-diastolic pressure in the left ventricle during reperfusion. Protection was unaffected by the PKC inhibitor bisindolylmaleimide (BIM; 1 micromol/l). It was abolished by the inhibitor of protein phosphatases 1 and 2A cantharidin (20 or 5 micromol/l) and partially enhanced by the inhibitor of protein phosphatase 2A okadaic acid (5 nmol/l). In adult rat cardiomyocytes treated with BIM and exposed to 60-min simulated ischemia (anoxia, extracellular pH 6.4), HP led to attenuation of anoxic Na(+)/Ca(2+) overload and of hypercontracture, which developed on reoxygenation. This protection was prevented by treatment with cantharidin but not with okadaic acid. In conclusion, HP exerts PKC-independent protection on ischemic-reperfused rat hearts and cardiomyocytes. Protein phosphatase 1 seems a mediator of this protective mechanism.