Influence of VIP on prolactinemia in turkey anterior pituitary cells: role of cAMP second messenger in VIP-induced prolactin gene expression

Vasoactive intestinal peptide (VIP) is the avian prolactin (PRL)-releasing factor. In the turkey, hypothalamic VIP immunoreactivity and mRNA content, as well as VIP levels in hypophyseal portal blood, are closely related to the state of prolactinemia and the reproductive stage. The present study investigated the role of VIP on prolactinemia in turkey anterior pituitary (AP) cells through PRL gene expression and the role of a cAMP second messenger system on VIP-induced PRL expression. In primary AP cells harvested from hens in different prolactinemic states, steady state promoter activities were positively correlated with secreted PRL levels. VIP increased PRL promoter activities in AP cells from hens with intermediate PRL levels (laying), but not in AP cells from hypoprolactinemic hens (nonphotostimulated reproductively quiescent). However, in AP cells from hyperprolactinemic hens (incubating), PRL promoter activity was down-regulated by VIP. PRL mRNA steady state levels were significantly decreased by the cAMP analogue, 8-bromo-cyclic adenosine monophosphate (8-Br-cAMP), and PRL secretion was down-regulated by the phosphodiesterase blocker, 3-isobutyl-1-methylxanthine (IBMX) in a dose-dependent manner, suggesting that the cAMP second messenger system might be involved in the inhibitory action of dopamine upon VIP-stimulated PRL secretion and gene expression at the pituitary level. In a study of VIP immediate and long-term effects on c-fos expression in relation to PRL expression, VIP dramatically induced c-fos mRNA expression within 5 min, suggesting that VIP-induced c-fos expression might be involved in VIP-stimulated PRL secretion and gene expression. These results provide additional evidence of the functional significance of VIP in PRL gene expression and suggest that changes in PRL promoter activity by VIP may be one of the important inductive mechanisms leading to prolactinemia.     

Fine structure of muscles in the tickHyalomma (Hyalomma) dromedarii (Ixodoidea: Ixodidae)

Skeletal and visceral muscles are distinguished in the unfed nymphHyalomma (Hyalomma) dromedarii according to position, structure and function. The skeletal muscles include the capitulum, dorsoventral and leg oblique muscles. Their muscle fibres have the striated pattern of successive sarcomeres whose thick myosin filaments are surrounded by orbitals of up to 12 thin actin filaments. The cell membrane invaginates into tubular system (T) extending deeply into the sarcoplasm and closely associated to cisternae of sarcoplasmic reticulum (SR). The T and SR forming two-membered dyads are considered to be the main route of calcium ions whose movements are synchronized with the motor impulse to control contraction and relaxation in most muscles. Two types of skeletal muscle fibres are recognized, and are suggested as representing different physiological phases.In the visceral-muscle fibres investing tick internal organs, the actin and myosin filaments are slightly interrupted, and the T and SR are well demonstrated. Both skeletal and visceral muscles are invaginated by tracheoles and innervated by nerve-axons containing synaptic vesicles.     

Microwave‐assisted water extraction of green tea polyphenols

Introduction – Green tea, a popular drink with beneficial health properties, is a rich source of specific flavanols (polyphenols). There is a special interest in the water extraction of green tea polyphenols since the composition of the corresponding extracts is expected to reflect the one of green tea infusions consumed worldwide. Objective – To develop a microwave‐assisted water extraction (MWE) of green tea polyphenols. Methodology – MWE of green tea polyphenols has been investigated as an alternative to water extraction under conventional heating (CWE). The experimental conditions were selected after consideration of both temperature and extraction time. The efficiency and selectivity of the process were determined in terms of extraction time, total phenolic content, chemical composition (HPLC‐MS analysis) and antioxidant activity of the extracts. Results – By MWE (80°C, 30 min), the flavanol content of the extract reached 97.46 (± 0.08) mg of catechin equivalent/g of green tea extract, vs. only 83.06 (± 0.08) by CWE (80°C, 45 min). In particular, the concentration of the most bioactive flavanol EGCG was 77.14 (± 0.26) mg of catechin equivalent/g of green tea extract obtained by MWE, vs 64.18 (± 0.26) mg/g by CWE. Conclusion – MWE appears more efficient than CWE at both 80 and 100°C, particularly for the extraction of flavanols and hydroxycinnamic acids. Although MWE at 100°C typically affords higher yields in total phenols, MWE at 80°C appears more convenient for the extraction of the green tea‐specific and chemically sensitive flavanols. Copyright © 2009 John Wiley & Sons, Ltd.     

Using Adeno-associated Virus as a Tool to Study Retinal Barriers in Disease

Müller cells are the principal glial cells of the retina. Their end-feet form the limits of the retina at the outer and inner limiting membranes (ILM), and in conjunction with astrocytes, pericytes and endothelial cells they establish the blood-retinal barrier (BRB). BRB limits material transport between the bloodstream and the retina while the ILM acts as a basement membrane that defines histologically the border between the retina and the vitreous cavity. Labeling Müller cells is particularly relevant to study the physical state of the retinal barriers, as these cells are an integral part of the BRB and ILM. Both BRB and ILM are frequently altered in retinal disease and are responsible for disease symptoms.There are several well-established methods to study the integrity of the BRB, such as the Evans blue assay or fluorescein angiography. However these methods do not provide information on the extent of BRB permeability to larger molecules, in nanometer range. Furthermore, they do not provide information on the state of other retinal barriers such as the ILM. To study BRB permeability alongside retinal ILM, we used an AAV based method that provides information on permeability of BRB to larger molecules while indicating the state of the ILM and extracellular matrix proteins in disease states. Two AAV variants are useful for such study: AAV5 and ShH10. AAV5 has a natural tropism for photoreceptors but it cannot get across to the outer retina when administered into the vitreous when the ILM is intact (i.e., in wild-type retinas). ShH10 has a strong tropism towards glial cells and will selectively label Müller glia in both healthy and diseased retinas. ShH10 provides more efficient gene delivery in retinas where ILM is compromised. These viral tools coupled with immunohistochemistry and blood-DNA analysis shed light onto the state of retinal barriers in disease.     

Comparison of the adaptive potential of the Arthrobacter oxydans and Acinetobacter lwoffii isolates from permafrost sedimentary rock and the analogous collection strains

A comparative study was conducted on the adaptive mechanisms of the strains Arthrobacter oxydans K14 and Acinetobacter lwoffii EK30A isolated from permafrost subsoil sediments and of those of the analogous collection strains (Ac-1114 Type and BSW-27, respectively). In each pair of the strains compared, the strains differed in terms of (i) growth-related, physiological, and biochemical properties; (ii) resistance to stress factors; (iii) capacity for generation of dormant forms (DFs) under growth arrest conditions, and (iv) intrapopulation production of phase variants. The strains isolated from permafrost displayed a lower growth rate but were more resistant to repeated freezing-thawing treatment than the collection strains. Under the same growth conditions, the permafrost strains formed larger numbers of cystlike anabiotic DFs, extraordinarily small cells, and forms that became nonculturable during long-term storage. Resuscitation of the nonculturable forms resulted in a 2- to-7-fold increase in the percentage of FISH-detectable metabolically active cells. The permafrost strains were also distinguished by increased genome lability. This facilitated their dissociation into intrapopulation variants with phenotypically distinct colonial and morphological properties and different antibiotic resistance. The phenotypic variability was more prominent in Arthrobacter (for which it was not reported previously) than in Acinetobacter. In the populations produced by plating the dormant bacterial forms, the qualitative and quantitative characteristics of the phase variant spectra varied depending on the formation conditions and the composition of the solid media used for the plating. Thus, the permafrost isolates of A. oxydans and Ac. lwoffii were distinguished from their collection analogs by a more manifest adaptive potential including stress resistance, the intensity of DF generation under growth arrest conditions, and increased intrapopulation variability.     

Changes in Membrane Fatty Acid Composition of Pseudomonas aeruginosa in Response to UV-C Radiations

The changes in lipid composition enable the micro-organisms to maintain membrane functions in the face of environmental fluctuations. The relationship between membrane fatty acid composition and UV-C stress was determined for mid-exponential phase and stationary phase Pseudomonas aeruginosa. The total lipids were obtained by dichloromethane/methanol (3:1) and were quantified by GC. The TLC analysis of phospholipids showed the presence of three major fractions phosphatidylethanolamine, phosphatidylglycerol, and cardiolipin. Significant modifications, as manifested by an increase of UFA, were obtained. Interestingly, this microorganism showed a remarkable capacity for recovery from the stressful effects of UV-C.