Noninvasive diffusive optical imaging of the auditory response to birdsong in the zebra finch

Songbirds communicate by learned vocalizations with concomitant changes in neurophysiological and genomic activities in discrete parts of the brain. Here, we tested a novel implementation of diffusive optical imaging (also known as diffuse optical imaging, DOI) for monitoring brain physiology associated with vocal signal perception. DOI noninvasively measures brain activity using red and near-infrared light delivered through optic fibers (optodes) resting on the scalp. DOI does not harm subjects, so it raises the possibility of repeatedly measuring brain activity and the effects of accumulated experience in the same subject over an entire life span, all while leaving tissue intact for further study. We developed a custom-made apparatus for interfacing optodes to the zebra finch (Taeniopygia guttata) head using 3D modeling software and rapid prototyping technology, and applied it to record responses to presentations of birdsong in isoflurane-anesthetized zebra finches. We discovered a subtle but significant difference between the hemoglobin spectra of zebra finches and mammals which has a major impact in how hemodynamic responses are interpreted in the zebra finch. Our measured responses to birdsong playback were robust, highly repeatable, and readily observed in single trials. Responses were complex in shape and closely paralleled responses described in mammals. They were localized to the caudal medial portion of the brain, consistent with response localization from prior gene expression, electrophysiological, and functional magnetic resonance imaging studies. These results define an approach for collecting neurophysiological data from songbirds that should be applicable to diverse species and adaptable for studies in awake behaving animals.     

Anxiolytic Properties of New Chemical Entity, 5TIO1

2-[(2,6-dichlorobenzylidene)amino]-5,6-dihydro-4H-cyclopenta[b]thiophene-3-carbonitrile), 5TIO1, is a new 2-aminothiophene derivative with a promising pharmacological activity. The aim of this work was to evaluate the potential anxiolytic effect of 5TIO1 in animal models. In the elevated plus-maze test, 5TIO1 (0.1, 1.0 and 10.0 mg/kg, i.p) increased the time of permanence and the number of entries in the open arms. In the light/dark box test, 5TIO1 at dose of 0.1 mg/kg (i.p) also showed anxiolytic-like effect indicated by an increase in the time spent in the light box, similar to diazepam 2.0 mg/kg (i.p). 5TIO1 groups did not change locomotor and coordination activities in open field and rotarod tests, respectively, when compared to vehicle. Dose dependent process was not observed and the anxiolytic effects demonstrated were not completely reversed by flumazenil 25 mg/kg (i.p). Our results suggest that 5TIO1 can bind with other receptors, besides the benzodiazepine site of the GABA receptor in mouse brain.     

Amphiphilic block copolymer/poly(dimethylsiloxane) (PDMS) blends and nanocomposites for improved fouling-release

Amphiphilic diblock copolymers, Sz6 and Sz12, consisting of a poly(dimethylsiloxane) block (average degree of polymerisation = 132) and a PEGylated-fluoroalkyl modified polystyrene block (Sz, average degree of polymerisation = 6, 12) were prepared by atom transfer radical polymerization (ATRP). Coatings were obtained from blends of either block copolymer (1–10 wt%) with a poly(dimethylsiloxane) (PDMS) matrix. The coating surface presented a simultaneous hydrophobic and lipophobic character, owing to the strong surface segregation of the lowest surface energy fluoroalkyl chains of the block copolymer. Surface chemical composition and wettability of the films were affected by exposure to water. Block copolymer Sz6 was also blended with PDMS and a 0.1 wt% amount of multiwall carbon nanotubes (CNT). The excellent fouling-release (FR) properties of these new coatings against the macroalga Ulva linza essentially resulted from the inclusion of the amphiphilic block copolymer, while the addition of CNT did not appear to improve the FR properties.     

Flow cytometry and live confocal analysis for the evaluation of the uptake and intracellular distribution of FITC-ODN into HaCaT cells

In this study, the mechanism of the internalization and the cellular distribution of 59 fluorescein conjugated PS-ODN (FITC-ODN) after transfection with different mixed lipidic vesicles/oligo complexes (lipoplexes) have been investigated. Mixed lipidic vesicles were prepared with one of the most used cationic lipid (DOTAP) and different amounts of a cholic acid (UDCA) to release the oligo into HaCaT cells. Using flow cytometry, the cellular uptake of the oligo was studied with and without different inhibitors able to block selectively the different pathways involved in the internalization mechanism. The intracellular distribution of the oligo was analyzed by confocal laser scanning microscopy (CLSM), treating the cells with the lipoplexes and directly observing without any fixing procedure. To better carry out the colocalization studies, fluorescent-labeled markers, specific for the different cellular compartments, were coincubated with 59 fluorescein-conjugated 29-mer phosphorotioate oligonucleotide (FITC-ODN). The different lipidic vesicles affect the internalization mechanism of FITC-ODN. After using the inhibitors, the uptake of complexes involved a different internalization mechanism. The live CLSM analysis demonstrated that, after 1 hour from the complex incubation, the oligo was transferred into cells and localized into the endosomes; after 24 hours, the oligo was intracellularly localized close to the nuclear structure in a punctuate pattern. However, the results from fusion experiments showed also a binding of a quite low amount of oligo with the cell membranes.     

Structural characterization of MG and pre-MG states of proteins by MD simulations,NMR, and other techniques

Almost all proteins fold via a number of partially structured intermediates such as molten globule (MG) and pre-molten globule states. Understanding the structure of these intermediates at atomic level is often a challenge, as these states are observed under extreme conditions of pH, temperature, and chemical denaturants. Furthermore, several other processes such as chemical modification, site-directed mutagenesis (or point mutation), and cleavage of covalent bond of natural proteins often lead to MG like partially unfolded conformation. However, the dynamic nature of proteins in these states makes them unsuitable for most structure determination at atomic level. Intermediate states studied so far have been characterized mostly by circular dichroism, fluorescence, viscosity, dynamic light scattering measurements, dye binding, infrared techniques, molecular dynamics simulations, etc. There is a limited amount of structural data available on these intermediate states by nuclear magnetic resonance (NMR) and hence there is a need to characterize these states at the molecular level. In this review, we present characterization of equilibrium intermediates by biophysical techniques with special reference to NMR.     

Zap70 inhibits Syk‐mediated osteoclast function

The αvβ3 integrin stimulates the resorptive capacity of the differentiated osteoclast (OC) by organizing its cytoskeleton via the tyrosine kinase, Syk. Thus, Syk‐deficient OCs fails to spread or form actin rings, in vitro and in vivo. The Syk family of tyrosine kinases consists of Syk itself and Zap70 which are expressed by different cell types. Because of their structural similarity, and its compensatory properties in other cells, we asked if Zap70 can substitute for absence of Syk in OCs. While expression of Syk, as expected, normalizes the cytoskeletal abnormalities of Syk^?/? OCs, Zap70 fails do so. In keeping with this observation, Syk, but not Zap70, rescues αvβ3 integrin‐induced SLP76 phosphorylation in Syk^?/? OCs. Furthermore the kinase sequence of Syk partially rescues the Syk^?/? phenotype but full normalization also requires its SH2 domains. Surprisingly, expression of Zap70 inhibits WT OC spreading, actin ring formation and bone resorptive activity, but not differentiation. In keeping with arrested cytoskeletal organization, Zap70 blocks integrin‐activated endogenous Syk and Vav3, SLP76 phosphorylation. Such inhibition requires Zap70 kinase activity, as it is abolished by mutation of the Zap70 kinase domain. Thus, while the kinase domain of Syk is uniquely required for OC function that of Zap70 inhibits it. J. Cell. Biochem. 114: 1871–1878, 2013. © 2013 Wiley Periodicals, Inc.     

Depression in Medical School: The Influence of Morningness‐Eveningness

Medical students are at higher risk for depression, affecting not only their lives but also patient care. This article studied a population of medical students engaged in lecture‐based learning regarding the presence of depressive symptoms and its relation to morningness‐eveningness. Depressive symptoms were assessed by the Beck Depressive Inventory scale (BDI>10), and diurnal preference was assessed by the Horne & Östberg Morningness/Eveningness Questionnaire (MEQ). Family history of depression and involvement in regular physical activity were also investigated. A total of 161 students, 77 (47.8%) males, aged 19 to 30 yrs (22.1±2.1) living in a city close to the equator were evaluated. Fifty‐three individuals (32.9%) had depressive symptoms. Depressive individuals showed a trend to be female (p=0.07). Also, female gender showed a non‐significant shift toward morningness. Fifty‐eight (36.0%) subjects participated in regular physical activity. In 57 cases (35.4%), there was a history of depression in the family. Fifteen individuals (9.3%) were definitely evening type, 42 (26.1%) were moderately evening type, 44 (27.3%) were indifferent, 42 (26.1%) were moderately morning type, and 18 (11.2%) were definitely morning type. Family history of depression (OR=0.29, 95% CI=1.37–6.12) and sedentary life (OR=0.28, 95% CI=0.12–0.65) were associated with depressive symptoms. Eveningness was associated with depressive symptoms (OR=0.66, 95% CI=0.50–0.88), and this association remained significant after adjusting for the presence of familial depression and physical activity (OR=0.71, 95% CI=0.52–0.95). In conclusion, depressive symptoms are independently associated with “eveningness” in medical students. These results should be confirmed by future studies involving a larger number of subjects.