Assessing visual requirements for social context-dependent activation of the songbird song system

Social context has been shown to have a profound influence on brain activation in a wide range of vertebrate species. Best studied in songbirds, when males sing undirected song, the level of neural activity and expression of immediate early genes (IEGs) in several song nuclei is dramatically higher or lower than when they sing directed song to other birds, particularly females. This differential social context-dependent activation is independent of auditory input and is not simply dependent on the motor act of singing. These findings suggested that the critical sensory modality driving social context-dependent differences in the brain could be visual cues. Here, we tested this hypothesis by examining IEG activation in song nuclei in hemispheres to which visual input was normal or blocked. We found that covering one eye blocked visually induced IEG expression throughout both contralateral visual pathways of the brain, and reduced activation of the contralateral ventral tegmental area, a non-visual midbrain motivation-related area affected by social context. However, blocking visual input had no effect on the social context-dependent activation of the contralateral song nuclei during female-directed singing. Our findings suggest that individual sensory modalities are not direct driving forces for the social context differences in song nuclei during singing. Rather, these social context differences in brain activation appear to depend more on the general sense that another individual is present.     

A potential relationship between the acrosome response characteristics of bovine spermatozoa and their in vitro fertilizing ability

The aim of the work was to study a potential relationship between acrosome response characteristics of bovine spermatozoa and their ability to fertilize oocytes and produce in vitro embryos. Sperm of artificial insemination bulls with a high rate (22.0 +/- 4.1%, group A, n = 7) or a low rate (10.3 +/- 4.1%, group B, n = 8) of embryos were used. For acrosome assessment, motile spermatozoa from a Percoll gradient were incubated with or without heparin and examined by the fix-vital sperm assay (FVSA). The differences between the heparin-treated (H+) and the non-treated (H-) spermatozoa were significant (p < 0.01) in all bulls at all tested intervals. According to the kinetics of the heparin response, the bulls fell into three categories: fast (FR, n = 7), moderate (MR, n = 5) or slow (SR, n = 3) acrosome responses (p < 0.01). Five MR bulls were found in group A in comparison with two MR bulls in group B (57.1 vs 12.5%; p < 0.05). Intensity of the acrosome response (response index) was significantly higher in bull group A compared with bull group B (7.0 vs 4.6, p < 0.01). A positive correlation was recorded between response index and embryo rate (r = 0.668, p < 0.01). In conclusion (a) the kinetics of spermatozoa response to heparin may be important for in vitro fertilization, bulls with a moderate response appearing to be most suitable for embryo production; (b) greater spermatozoa response to heparin was related to more effective embryo production.     

Brain Under Stress and Alzheimer’s Disease

Modern society is characterized by the ubiquity of stressors that affect every individual to different extents. Furthermore, experimental, clinical, and epidemiological data have shown that chronic activation of the stress response may participate in the development of various somatic as well as neuropsychiatric diseases. Surprisingly, the role that stress plays in the etiopathogenesis of Alzheimer’s disease (AD) has not yet been studied in detail and is therefore not well understood. However, accumulated data have shown that neuroendocrine and behavioral changes accompanying the stress response affect neuronal homeostasis and compromise several key neuronal processes. Mediators of the neuroendocrine stress response, if elevated repeatedly or chronically, exert direct detrimental effects on the brain by impairing neuronal metabolism, plasticity, and survival. Stress-induced hormonal and behavioral reactions may also participate in the development of hypertension, atherosclerosis, insulin resistance, and other peripheral disturbances that may indirectly induce neuropathological processes participating in the development and progression of AD. Importantly, stress-induced detrimental effects as etiological factors of AD are attractive because they can be reduced by several approaches including behavioral and pharmacological interventions. These interventions may therefore represent an important strategy for prevention or attenuation of the progression of AD.     

The genetic incorporation of p-azidomethyl-l-phenylalanine into proteins in yeast

The noncanonical amino acid p-azidomethyl-l-phenylalanine can be genetically incorporated into proteins in bacteria, and has been used both as a spectroscopic probe and for the selective modification of proteins by alkynes using click chemistry. Here we report identification of Escherichia coli tyrosyl tRNA synthetase mutants that allow incorporation of p-azidomethyl-l-phenylalanine into proteins in yeast. When expressed together with the cognate E. coli tRNA_CUA^Tyr, the new mutant tyrosyl tRNA synthetases directed robust incorporation of p-azidomethyl-l-phenylalanine into a model protein, human superoxide dismutase, in response to the UAG amber nonsense codon. Mass spectrometry analysis of purified superoxide dismutase proteins confirmed the efficient site-specific incorporation of p-azidomethyl-l-phenylalanine. This work provides an additional tool for the selective modification of proteins in eukaryotic cells.     

A Ca(v)3.2/syntaxin-1A signaling complex controls T-type channel activity and low-threshold exocytosis

T-type calcium channels represent a key pathway for Ca(2+) entry near the resting membrane potential. Increasing evidence supports a unique role of these channels in fast and low-threshold exocytosis in an action potential-independent manner, but the underlying molecular mechanisms have remained unknown. Here, we report the existence of a syntaxin-1A/Ca(v)3.2 T-type calcium channel signaling complex that relies on molecular determinants that are distinct from the synaptic protein interaction site (synprint) found in synaptic high voltage-activated calcium channels. This interaction potently modulated Ca(v)3.2 channel activity, by reducing channel availability. Other members of the T-type calcium channel family were also regulated by syntaxin-1A, but to a smaller extent. Overexpression of Ca(v)3.2 channels in MPC 9/3L-AH chromaffin cells induced low-threshold secretion that could be prevented by uncoupling the channels from syntaxin-1A. Altogether, our findings provide compelling evidence for the existence of a syntaxin-1A/T-type Ca(2+) channel signaling complex and provide new insights into the molecular mechanism by which these channels control low-threshold exocytosis.     

Surface heparinization of polyurethane via bromoalkylation of hard segment nitrogens

Previous research from our group has demonstrated that bromoalkylation of polyurethane elastomers via base mediated activation of the urethane-hard segment nitrogen groups can be used to either attach bisphosphonate groups to confer calcification resistance or append cholesterol to promote endothelial cell adhesion. In the present studies we further explore the potential of this chemical approach by investigating bulk carboxylation of polyurethanes via bromoalkylation to enable surface heparinization for thromboresistance. Thus, polyurethane (PU) was modified with pendant 7-carboxy-5-thiaheptyl groups using a polymer-analogous reaction of bromobutylated PU with tetrabutylammonium 3-mercaptopropionate in mild conditions. The grafting of polyallylamine (PAA) onto the surface of carboxylated PU via direct coupling of amino and carboxy groups resulted in high levels of PAA (up to 8 mug/cm(2)). The surface-aminated PU was further covalently modified with unfractionated heparin as confirmed by FTIR. Fluorescence labeling of PAA hydrochloride and heparin with BODIPY-FL was used to quantify the extent of surface modifications. Heparin was covalently bound at a high level (1.11 +/- 0.06 mug/cm(2)) and was shown to be active, with demonstrable Factor Xa inhibition and platelet factor IV binding. It is concluded that surface amination of bulk-carboxylated PU represents a novel approach for heparinizing PU; carboxylation followed by surface amination represents another important dimension of bromo-alkyl activation of polyurethane hard segments, thereby enabling heparinization.     

Ferritin oxidation and proteasomal degradation: protection by antioxidants

The accumulation of oxidatively damaged proteins is a well-known hallmark of aging and several neurodegenerative diseases including Alzheimer's, Parkinson's and Huntigton's diseases. These highly oxidized protein aggregates are in general not degradable by the main intracellular proteolytic machinery, the proteasomal system. One possible strategy to reduce the accumulation of such oxidized protein aggregates is the prevention of the formation of oxidized protein derivatives or to reduce the protein oxidation to a degree that can be handled by the proteasome. To do so an antioxidative strategy might be successful. Therefore, we undertook the present study to test whether antioxidants are able to prevent the protein oxidation and to influence the proteasomal degradation of moderate oxidized proteins. As a model protein we choose ferritin. H₂O₂ induced a concentration dependent increase of protein oxidation accompanied by an increased proteolytic susceptibility. This increase of proteolytic susceptibility is limited to moderate hydrogen peroxide concentrations, whereas higher concentrations are accompanied by protein aggregate formation.

Protective effects of the vitamin E derivative Trolox, the pyridoindole derivative Stobadine and of the standardized extracts of flavonoids from bark of Pinus Pinaster Pycnogenol^® and from leaves of Ginkgo biloba (EGb 761) were studied on moderate damaged ferritin.