Ionotropic GABA receptor expression in the lung during development

Cl(-) transport is essential for lung development. Because gamma-aminobutyric acid (GABA) receptors allow the flow of negatively-charged Cl(-) ions across the cell membrane, we hypothesized that the expression of ionotropic GABA receptors are regulated in the lungs during development. We identified 17 GABA receptor subunits in the lungs by real-time PCR. These subunits were categorized into four groups: Group 1 had high mRNA expression during fetal stages and low in adults; Group 2 had steady expression to adult stages with a slight up-regulation at birth; Group 3 showed an increasing expression from fetal to adult lungs; and Group 4 displayed irregular mRNA fluctuations. The protein levels of selected subunits were also determined by Western blots and some subunits had protein levels that corresponded to mRNA levels. Further studied subunits were primarily localized in epithelial cells in the developing lung with differential mRNA expression between isolated cells and whole lung tissues. Our results add to the knowledge of GABA receptor expression in the lung during development.     

Development of Microplitis croceipes as a biological sensor

Classical conditioning, a form of associative learning, was first described in the vertebrate literature by Pavlov, but has since been documented for a wide variety of insects. Our knowledge of associative learning by insects began with Karl vonFrisch explaining communication among honeybees, Apis mellifera L. (Hymenoptera: Apidae). Since then, the honey bee has provided us with much of what we understand about associative learning in insects and how we relate the theories of learning in vertebrates to insects. Fruit flies, moths, and parasitic wasps are just a few examples of other insects that have been documented with the ability to learn. A novel direction in research on this topic attempts to harness the ability of insects to learn for the development of biological sensors. Parasitic wasps, especially Microplitis croceipes (Cresson) (Hymenoptera: Braconidae), have been conditioned to detect the odors associated with explosives, food toxins, and cadavers. Honeybees and moths have also been associatively conditioned to several volatiles of interest in forensics and national security. In some cases, handheld devices have been developed to harness the insects and observe conditioned behavioral responses to air samples in an attempt to detect target volatiles. Current research on the development of biological sensors with insects is focusing on factors that influence the learning and memory ability of arthropods as well as potential mathematical techniques for improving the interpretation of the behavioral responses to conditioned stimuli. Chemical detection devices using arthropod‐based sensing could be used in situations where trained canines cannot be used (such as toxic environments) or are unavailable, electronic devices are too expensive and/or not of sufficient sensitivity, and when conditioning to target chemicals must be done within minutes of detection. The purpose of this article is to provide a brief review of the development of M. croceipes as a model system for exploring associative learning for the development of biological sensors.     

Using insect sniffing devices for detection

Emerging information about the ability of insects to detect and associatively learn has revealed that they could be used within chemical detection systems. Such systems have been developed around free-moving insects, such as honey bees. Alternatively, behavioral changes of contained insects can be interpreted by sampling air pumped over their olfactory organs. These organisms are highly sensitive, flexible, portable and cheap to reproduce, and it is easy to condition them to detect target odorants. However, insect-sensing systems are not widely studied or accepted as proven biological sensors. Further studies are needed to examine additional insect species and to develop better methods of using their olfactory system for detecting odorants of interest.     

Ras is active throughout the cell cycle,but is able to induce cyclin D1 only during G2 phase

The control of cell cycle progression has been studied in asynchronous cultures using image analysis and time lapse techniques. This approach allows determination of the cycle phase and signaling properties of individual cells, and avoids the need for synchronization. In past studies this approach demonstrated that continuous cell cycle progression requires the induction of cyclin D1 levels by Ras, and that this induction takes place during G2 phase. These studies were designed to understand how Ras could induce cyclin D1 levels only during G2 phase. First, in studies with a Ras-specific promoter and cellular migration we find that endogenous Ras is active in all cell cycle phases of actively cycling NIH3T3 cells. This suggests that cyclin D1 induction during G2 phase is not the result of Ras activation specifically during this cell cycle period. To confirm this suggestion oncogenic Ras, which is expected to be active in all cell cycle phases, was microinjected into asynchronous cells. The injected protein induced cyclin D1 levels rapidly, but only in G2 phase cells. We conclude that in the continuously cycling cell the targets of Ras activity are controlled by cell cycle phase, and that this phenomenon is vital to cell cycle progression.     

Parasitic wasps learn and report diverse chemicals with unique conditionable behaviors

Parasitoids exploit numerous chemical cues to locate hosts and food. Whether they detect and learn chemicals foreign to their natural history has not been explored. We show that the parasitoid Microplitis croceipes can associate, with food or hosts, widely different chemicals outside their natural foraging encounters. When learned chemicals are subsequently detected, this parasitoid manifests distinct behaviors characteristic with expectations of food or host, commensurate with prior training. This flexibility of parasitoids to rapidly link diverse chemicals to resource needs and subsequently report them with recognizable behaviors offers new insights into their foraging adaptability, and provides a model for further dissection of olfactory learning related processes.     

Behavioral management of recurrent headache: three decades of experience and empiricism

In the past three decades, behavioral interventions (chiefly relaxation, biofeedback, and stress-management) have become standard components of the armamentarium for management of migraine and tension-type headaches. Meta-analytic literature reviews of these behavioral interventions have consistently identified clinically significant reductions in recurrent headache. Across studies, behavioral interventions have yielded approximately 35–50% reduction in migraine and tension-type headache activity. Although we have only recently begun to directly compare standard drug and nondrug treatments for headache, the available evidence suggests that the level of headache improvement with behavioral interventions may rival those obtained with widely used pharmacologic therapies in representative patient samples. In recent years, some attempts have been made to increase the availability and cost effectiveness of behavioral interventions through alternative delivery formats and mass communications. Recent developments within diagnosis and classification are summarized, pointing out implications for behavioral researchers. Select future directions are discussed, which include impact of the triptans, cost and cost effectiveness, and integration of behavioral treatments into primary care settings, the place where the great majority of headache sufferers receive treatment.     

Gene structure, purification and characterization of DNA polymerase beta from Xiphophorus maculatus

Cloning of the Xiphophorus maculatus Polbeta gene and overexpression of the recombinant Polbeta protein has been performed. The organization of the XiphPolbeta introns and exons, including intron-exon boundaries, have been assigned and were found to be similar to that for human Polbeta with identical exon sizes except for exon XII coding for an additional two amino acid residues in Xiphophorus. The cDNA sequence encoding the 337-amino acid X. maculatus DNA polymerase beta (Polbeta) protein was subcloned into the Escherichia coli expression plasmid pET. Induction of transformed E. coli cells resulted in the high-level expression of soluble recombinant Polbeta, which catalyzed DNA synthesis on template-primer substrates. The steady-state Michaelis constants (Km) and catalytic efficiencies (kcat/Km) of the recombinant XiphPolbeta for nucleotide insertion opposite single-nucleotide gap DNA substrates were measured and compared with previously published values for recombinant human Polbeta. Steady-state in vitro Km and kcat/Km values for correct nucleotide insertion by XiphPolbeta and human Polbeta were similar, although the recombinant Xiphophorus protein exhibited 2.5-7-fold higher catalytic efficiencies for dGTP and dCTP insertion versus human Polbeta. In contrast, the recombinant XiphPolbeta displayed significantly lower fidelities than human Polbeta for dNTP insertion opposite a single-nucleotide gap at 37 degrees C.     

DNA polymerase beta mRNA and protein expression in Xiphophorus fish

Herein we report Xiphophorus DNA polymerase beta (XiphPolbeta) mRNA and protein expression levels in brain, liver, gill, and testes tissues from Xiphophorus maculatus, Xiphophorus helleri, and Xiphophorus couchianus parental line fish and two different tumor-bearing Xiphophorus interspecies hybrids. Polymerase beta protein levels in the Xiphophorus tissues were measured by Western blot, and mRNA was measured with a quantitative real time RT-PCR method which employed cRNA construction to produce accurate calibration curves. We found significant differences in both mRNA and protein levels between the tumor-bearing hybrid animals and the three parental species. However, there were no significant differences in either mRNA levels or protein expression observed between the parental species. Thus, interspecies hybridization results in dysregulation of Polbeta expression and this may manifest a modulation in DNA repair capability and susceptibility to latent tumorigenesis.     

A ciliopathy complex builds distal appendages to initiate ciliogenesis

Cells inherit two centrioles, the older of which is uniquely capable of generating a cilium. Using proteomics and superresolved imaging, we identify a module that we term DISCO (distal centriole complex). The DISCO components CEP90, MNR, and OFD1 underlie human ciliopathies. This complex localizes to both distal centrioles and centriolar satellites, proteinaceous granules surrounding centrioles. Cells and mice lacking CEP90 or MNR do not generate cilia, fail to assemble distal appendages, and do not transduce Hedgehog signals. Disrupting the satellite pools does not affect distal appendage assembly, indicating that it is the centriolar populations of MNR and CEP90 that are critical for ciliogenesis. CEP90 recruits the most proximal known distal appendage component, CEP83, to root distal appendage formation, an early step in ciliogenesis. In addition, MNR, but not CEP90, restricts centriolar length by recruiting OFD1. We conclude that DISCO acts at the distal centriole to support ciliogenesis by restraining centriole length and assembling distal appendages, defects in which cause human ciliopathies.