ROS-GC subfamily membrane guanylate cyclase-linked transduction systems: taste, pineal gland and hippocampus

In the continuous efforts to test the validity of the theme that the Ca²⁺-modulated ROS-GC subfamily system is a universal transduction component of the sensory and sensory-linked network of neurons, this article focuses on the presence and variant biochemical forms of this transduction system in the gustatory epithelium, the site of gustatory transduction; in the pineal, a light-sensitive gland; and in the hippocampus neurons, linked with the perception of all SENSES.     

Effects of pre- and post- natal exposure to flutamide on connexin 43 expression in testes and ovaries of prepubertal pigs

The aim of this study was to show whether connexin43 (C×43) expression in gonads is affected by an anti-androgen action. To perform this test, pigs were prenatally (on gestational days 20–28 and 80–88; GD20, GD80) and postnatally (on days 2–10 after birth; PD2) exposed to flutamide, which was given in five doses every second day and its effect was observed in prepubertal gilts and boars. Morphology and expression of C×43 was investigated in testes and ovaries by means of routine histology, immunohistochemistry, Western blotting, and RT-PCR. In boars exposed to flutamide varying degrees of seminiferous tubule abnormality, including reduced number of Sertoli cells, tubules with severely dilated lumina and multinucleated germ cells were observed, whereas in gilts, the administration of flutamide at GD20 resulted in delayed folliculogenesis. Only follicles at the preantral stage were observed. Qualitative analysis of immunohistochemical staining for C×43 was confirmed by quantitative image analysis, where the staining intensity was expressed as relative optical density of diaminobenzidine deposits. After flutamide exposure, statistically significant increase in C×43 signal intensity was observed between interstitial tissue of GD20 and control pigs (**P<0.01), between seminiferous tubules of PD2 and control boars (**P<0.01) and between theca cells of GD80, of PD2 and control gilts (**P<0.01). In contrast, statistically significant decrease in C×43 signal intensity was found between granulosa cells of GD20, of PD2 and control gilts (**P<0.01 and *P<0.05, respectively) and between theca cells of GD20 and control gilts (**P<0.01). Since we demonstrated changes in gonad morphology and in the expression of C×43 at the level of protein of prepubertal boars and gilts, it seems possible that flutamide, through blocking androgen action, causes delayed gonadal maturation in later postnatal life and, among other factors, may be involved in the regulation of C×43 gene expression in pig gonads.Key words: C×43 gene expression, testis, ovary, flutamide, prepubertal pigs.     

Solution conformation of laminaribioside and (1 → 3)-β-D-glucan from optical rotation

A chiroptical method of conformational analysis is applied to linear (1 → 3)-β-D -glucans and the dimeric analogues β- and α-laminaribioside. The method is based on a recently developed semiempirical calculational model for saccharide optical activity. We conclude that disaccharide conformational energy maps in the literature represent the effective potential energy surface in aqueous solution well. The positive optical rotation observed with long chains in dilute alkaline solution is not characteristic of any single–chain conformation, and must reflect chain association.     

Obtaining and characterization of DNA-containing micromummies of yeasts and gram-positive bacteria with enhanced cell wall permeability: Application in PCR

The procedure of obtaining DNA-containing cell envelopes (“micromummies”) of bacteria, yeasts, and fungi using chaotropic salts has been developed previously and the possibility of their direct application in PCR has been demonstrated. The fine structure of micromummies has been studied by electron microscopic methods. This work has demonstrated that additional treatment of micromummies of yeasts and gram-positive bacteria with proteinase K results in hydrolytic degradation of cell proteins and drastic enhancement of cell wall permeability for macromolecules (DNA). Thus, the efficiency of PCR ex situ using resultant micromummies after washing off the products of protein hydrolysis and proteinase K can be increased. The results of electron microscopic study of ultrathin sections of yeasts (Pichia pastoris, Saccharomyces cerevisiae) and gram-positive bacteria (Micrococcus luteus, Arthrobacter globiformis, Bacillus subtilis) support the biochemical data that treatment with chaotropic salts and proteinase K results in the loosening of microbial cell walls and in a decrease in the intracellular protein content. At the same time, cell walls generally maintain their integrity (continuity) and initial spherical or rodlike shape. The optimal modes of treatment of the cells of different microbial species with chaotropic salts and proteinase K have been selected to obtain permeabilized cell envelopes containing denatured or native DNA.     

Effects of geographical heterogeneity in species interactions on the evolution of venom genes

Geographical heterogeneity in the composition of biotic interactions can create a mosaic of selection regimes that may drive the differentiation of phenotypes that operate at the interface of these interactions. Nonetheless, little is known about effects of these geographical mosaics on the evolution of genes encoding traits associated with species interactions. Predatory marine snails of the family Conidae use venom, a cocktail of conotoxins, to capture prey. We characterized patterns of geographical variation at five conotoxin genes of a vermivorous species, Conus ebraeus, at Hawaii, Guam and American Samoa, and evaluated how these patterns of variation are associated with geographical heterogeneity in prey utilization. All populations show distinct patterns of prey utilization. Three ‘highly polymorphic’ conotoxin genes showed significant geographical differences in allelic frequency, and appear to be affected by different modes of selection among populations. Two genes exhibited low levels of diversity and a general lack of differentiation among populations. Levels of diversity of ‘highly polymorphic’ genes exhibit a positive relationship with dietary breadth. The different patterns of evolution exhibited by conotoxin genes suggest that these genes play different roles in prey capture, and that some genes are more greatly affected by differences in predator–prey interactions than others. Moreover, differences in dietary breadth appear to have a greater influence on the differentiation of venoms than differences in the species of prey.     

Signatures of miR-181a on the Renal Transcriptome and Blood Pressure

MicroRNA-181a binds to the 3′ untranslated region of messenger RNA (mRNA) for renin, a rate-limiting enzyme of the renin-angiotensin system. Our objective was to determine whether this molecular interaction translates into a clinically meaningful effect on blood pressure and whether circulating miR-181a is a measurable proxy of blood pressure. In 200 human kidneys from the TRANScriptome of renaL humAn TissuE (TRANSLATE) study, renal miR-181a was the sole negative predictor of renin mRNA and a strong correlate of circulating miR-181a. Elevated miR-181a levels correlated positively with systolic and diastolic blood pressure in TRANSLATE, and this association was independent of circulating renin. The association between serum miR-181a and systolic blood pressure was replicated in 199 subjects from the Genetic Regulation of Arterial Pressure of Humans In the Community (GRAPHIC) study. Renal immunohistochemistry and in situ hybridization showed that colocalization of miR-181a and renin was most prominent in collecting ducts where renin is not released into the systemic circulation. Analysis of 69 human kidneys characterized by RNA sequencing revealed that miR-181a was associated with downregulation of four mitochondrial pathways and upregulation of 41 signaling cascades of adaptive immunity and inflammation. We conclude that renal miR-181a has pleiotropic effects on pathways relevant to blood pressure regulation and that circulating levels of miR-181a are both a measurable proxy of renal miR-181a expression and a novel biochemical correlate of blood pressure.     

Traumatic brain injury and bone healing: radiographic and biomechanical analyses of bone formation and stability in a combined murine trauma model

Introduction:The combination of traumatic brain injury (TBI) and long-bone fractures has previously been reported to lead to exuberant callus formation. The aim of this experimental study was to radiographically and biomechanically study the effect of TBI on bone healing in a mouse model.Materials and methods:138 female C57/Black6N mice were assigned to four groups (fracture (Fx) / TBI / combined trauma (Fx/TBI) / controls). Femoral osteotomy and TBI served as variables: osteotomies were stabilized with external fixators, TBI was induced with controlled cortical impact injury. During an observation period of four weeks, in vivo micro-CT scans of femora were performed on a weekly basis. Biomechanical testing of femora was performed ex vivo.Results:The combined-trauma group showed increased bone volume, higher mineral density, and a higher rate of gap bridging compared to the fracture group. The combined-trauma group showed increased torsional strength at four weeks.Discussion:TBI results in an increased formation of callus and mineral density compared to normal bone healing in mice. This fact combined with a tendency towards accelerated gap bridging leads to increased torsional strength. The present study underscores the empirical clinical evidence that TBI stimulates bone healing. Identification of underlying pathways could lead to new strategies for bone-stimulating approaches in fracture care.     

A resource of Cre driver lines for genetic targeting of GABAergic neurons in cerebral cortex

A key obstacle to understanding neural circuits in the?cerebral cortex is that of unraveling the diversity of GABAergic interneurons. This diversity poses general questions for neural circuit analysis: how are these interneuron cell types generated and assembled into stereotyped local circuits and how do they differentially contribute to circuit operations that underlie cortical functions ranging from perception to cognition? Using genetic engineering in mice, we have generated and characterized approximately 20 Cre and inducible CreER knockin driver lines that reliably target major classes and lineages of GABAergic neurons. More select populations are captured by intersection of Cre and Flp drivers. Genetic targeting allows reliable identification, monitoring, and manipulation of cortical GABAergic neurons, thereby enabling a systematic and comprehensive analysis from cell fate specification, migration, and connectivity, to their functions in network dynamics and behavior. As such, this approach will accelerate the study of GABAergic circuits throughout the mammalian brain.