Noradrenergic afferents and receptors in the medial preoptic area: neuroanatomical and neurochemical links between the regulation of sleep and body temperature

Several studies have shown the importance of the medial preoptic area in the regulation of sleep-wakefulness and of body temperature. The medial preoptic area has a rich noradrenergic innervation, coming mostly from the lateral tegmental noradrenergic system. The accumulating evidences show that the noradrenergic afferents to the medial preoptic area are involved in the induction of sleep. This hypnogenic mechanism operates through the postsynaptic alpha1 and alpha2-adrenergic receptors. Noradrenergic afferents are also involved in the thermoregulatory mechanisms, and the activation of these fibers brings about a fall in body temperature. Though the body temperature changes are brought about by the same receptor subtypes as those involved in hypnogenesis, observations suggest the possibility of separate sets of noradrenergic afferents in the medial preoptic area for sleep regulation and thermoregulation. In this review, we present the compelling evidences, which showed that the noradrenergic afferents of the medial preoptic area bring about a fall in body temperature and other thermoregulatory behavioral alterations associated with sleep.     

Purification and properties of alpha-galactosidase from white-rot fungus Pleurotus florida

alpha-Galactosidase was strongly induced in the white-rot fungus Pleurotus florida by arabinose than its natural substrates and was purified to homogeneity by acetone precipitation, ultrafiltration and DEAE-Sepharose chromatography. The enzyme was a monomeric protein with a molecular mass of approximately equal to 99 kDa, as revealed by native-PAGE and SDS-PAGE. alpha-Galactosidase was optimally active at 55 degrees C for the hydrolysis of p-nitrophenyl-alpha-galactopyranoside (PNPalphaG) and lost its 20% and 50% of original activity in 30 min at 60 degres C and 70 degrees C, respectively. The pH optimum of the enzyme was between 4.6 and 5.0. It was stable in a wide pH range (pH 4.0 to 9.0) at 55 degrees C for 2 h. The Ag+ and Hg2+ strongly inhibited the enzyme activity. Galactose, glucose, maltose and lactose also inhibited the enzyme activity, whereas N-bromosuccinimide treatment resulted in near total loss of acitivity. The Km and Vmax values of the enzyme for PNPalphaG were found to be 1.1 mM, and 77 micromol min(-1) mg(-1), respectively. alpha-Galactosidase immobilized in agar was more effective for the degradation of raffinose than in the sodium alginate. TLC results indicated its potential for the removal of raffinose and stachyose in soymilk.     

Dual electrochemical determination of glucose and insulin using enzyme and ferrocene microcapsules

Dual electrochemical determination of glucose and insulin has been developed, based on enzymatic reaction and immunoassay with utilization of ferrocene microcapsules, respectively. Glucose was determined through electrochemical oxidation of formed product, hydrogen peroxide, by the action of glucose oxidase (GOx). The layer-by-layer (LbL) films on the ferrocene microcrystal followed by anti-insulin antibody sensitization were employed for the biolabled ferrocene microcapsules production. The antibody sensitized ferrocene microcapsules worked as a probe in the proposed system. The microcapsules provided a higher signal generating molecule to antibody (S/P) ratio of 4.52x10(6) to 12.4x10(6). Microcapsules with different antibody loads (388-1070 antibody molecules per capsule) were subjected to a solid-phase immunoassay for the detection of insulin. The microcapsule having 1030 anti-insulin antibody molecules per capsule demonstrated good performance for insulin determination. The calibration curve for insulin had a linear range of 10(-10) to 10(-7) g mL(-1) with R(2)=0.990, 3.9% R.S.D. The limit of detection for insulin was 10 pg mL(-1) of 100 microL sample (equivalent to 10(-12)g of insulin). The determination range for the glucose was 0.5 and 40 mM with R(2)=0.996 and 4.1% R.S.D.     

DNA hybridization detection with organic thin film transistors: toward fast and disposable DNA microarray chips

We demonstrate a novel DNA hybridization detection method with organic thin film transistors. DNA molecules are immobilized directly on the surface of organic semiconductors, producing an unambiguous doping-induced threshold voltage shift upon hybridization. With these shifts, single-stranded DNA (ssDNA) and double-stranded DNA (dsDNA) are differentiated successfully. This method is expected to result in higher sensitivity than the main competitive technology, ISFET-based sensors because of the direct exposure of DNA molecules to sensitive layers. Factors that influence sensor sensitivity have been analyzed and optimum conditions have been determined using statistically designed experiments. Under the optimum conditions, the maximum difference between saturation current ratios caused by ssDNA and dsDNA reaches as high as 70%. In order to make DNA detection fast, we also demonstrate rapid on-chip electrically enhanced hybridization using the TFTs. These technologies together will enable the realization of disposable, rapid-turnaround tools for field-deployable genomic diagnosis.     

Cytotoxic effect of achatininH (lectin) from Achatina fulica against a human mammary carcinoma cell line (MCF7)

The hemolymph-derived achatinin_H (lectin) from Achatina fulica showed a marked cytotoxic effect on MCF7, a human mammary carcinoma cell line. IC₅₀ values as measured by the 3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide assay for achatinin_H ranged from 6 to 10 μg/ml in the MCF7 cells. MCF7 cells showed significant morphological changes leading to cell death. The above cell death was observed after 48 h of treatment with 8 μg/ml when compared to untreated cells. Alterations in the tumor marker enzymes, as well as in antioxidant enzymes, were observed after achatinin_H treatment. The specificity and purity of the achatinin_H was confirmed by the Western blot assay. Achatinin_H binding to MCF7 cells was detected by anti-achatinin_H, and visualization of the achatinin_H binding sites on confluent MCF7 cells was confirmed by flourescein isothiocyanate conjugated secondary antibody. MCF7-treated cells fluoresced, indicating the presence of achatinin_H binding sites. Fluorescence-activated cell sorting analysis of the cell cycle showed a significant increase in S-phase in MCF7 cells after 48 h of achatinin_H treatment. The cells were arrested in G₂/M phase of the cell cycle after 48 h with significant changes in cell viability. Cellular damage was confirmed by agarose gel electrophoresis with the characteristic appearance of a DNA streak in treated MCF7 cells indicating the ongoing apoptosis. An erratum to this article can be found at     

Ancient population genomics and the study of evolution

Recently, the study of ancient DNA (aDNA) has been greatly enhanced by the development of second-generation DNA sequencing technologies and targeted enrichment strategies. These developments have allowed the recovery of several complete ancient genomes, a result that would have been considered virtually impossible only a decade ago. Prior to these developments, aDNA research was largely focused on the recovery of short DNA sequences and their use in the study of phylogenetic relationships, molecular rates, species identification and population structure. However, it is now possible to sequence a large number of modern and ancient complete genomes from a single species and thereby study the genomic patterns of evolutionary change over time. Such a study would herald the beginnings of ancient population genomics and its use in the study of evolution. Species that are amenable to such large-scale studies warrant increased research effort. We report here progress on a population genomic study of the Adélie penguin (Pygoscelis adeliae). This species is ideally suited to ancient population genomic research because both modern and ancient samples are abundant in the permafrost conditions of Antarctica. This species will enable us to directly address many of the fundamental questions in ecology and evolution.