Effects of D2O on permeation and gating in the Ca2+-activated potassium channel fromChara

We studied the effects of H₂O/D₂O substitution on the permeation and gating of the large conductance Ca²⁺-activated K⁺ channels inChara gymnophylla droplet membrane using the patchclamp technique. The selectivity sequence of the channel was: K⁺>Rb⁺≫Li⁺, Na⁺, Cs⁺ and Cl⁻. The conductance of this channel in symmetric 100mm KCl was found to be 130 pS. The single channel conductance was decreased by 15% in D₂O as compared to H₂O. The blockade of channel conductance by cytosolic Ca²⁺ weakened in D₂O as a result of a decrease in zero voltage Ca²⁺ binding affinity by a factor of 1.4. Voltage-dependent channel gating was affected by D₂O primarily due to the change in Ca²⁺ binding to the channel during the activation step. The Hill coefficient for Ca²⁺ binding was 3 in D₂O and around 1 in H₂O. The values of the Ca²⁺ binding constant in the open channel conformation were 0.6 and 6 μm in H₂O and D₂O, respectively, while the binding in the closed conformation was much less affected by D₂O. The H₂O/D₂O substitution did not produce a significant change in the slope of channel voltage dependence but caused a shift as large as 60 mV with 1mm internal Ca²⁺.     

Orthologous gene-expression profiling in multi-species models: search for candidate genes

Microarray-driven gene-expression profiles are generally produced and analyzed for a single specific experimental model. We have assessed an analytical approach that simultaneously evaluates multi-species experimental models within a particular biological condition using orthologous genes as linkers for the various Affymetrix microarray platforms on multi-species models of ventilator-associated lung injury. The results suggest that this approach may be a useful tool in the evaluation of biological processes of interest and selection of process-related candidate genes.     

Physiological Heterothallism and Sexuality in Euascomycetes: A Partial History

Copyright 1998 Academic Press.     

Expression of Toll-like receptor 2 is up-regulated in monocytes from patients with chronic obstructive pulmonary disease

Background

Chronic obstructive pulmonary disease (COPD) is characterised by pulmonary and systemic inflammation which flare-up during episodes of acute exacerbation (AECOPD). Given the role of Toll-like receptors (TLRs) in the induction of inflammatory responses we investigated the involvement of TLRs in COPD pathogenesis.

Methods

The expression of TLR-2, TLR-4 and CD14 in monocytes was analyzed by flow cytometry. To study the functional responses of these receptors, monocytes were stimulated with peptidoglycan or lipopolysaccharide and the amounts of TNFα and IL-6 secreted were determined by ELISA.

Results

We found that the expression of TLR-2 was up-regulated in peripheral blood monocytes from COPD patients, either clinically stable or during AECOPD, as compared to never smokers or smokers with normal lung function. Upon stimulation with TLR-2 ligand monocytes from COPD patients secreted increased amounts of cytokines than similarly stimulated monocytes from never smokers and smokers. In contrast, the expressions of TLR-4 and CD14 were not significantly different between groups, and the response to lipopolysaccharide (a TLR-4 ligand) stimulation was not significantly different either. At discharge from hospital TLR-2 expression was down-regulated in peripheral blood monocytes from AECOPD patients. This could be due to the treatment with systemic steroids because, in vitro, steroids down-regulated TLR-2 expression in a dose-dependent manner. Finally, we demonstrated that IL-6, whose plasma levels are elevated in patients, up-regulated in vitro TLR-2 expression in monocytes from never smokers.

Conclusion

Our results reveal abnormalities in TLRs expression in COPD patients and highlight its potential relationship with systemic inflammation in these patients.

     

Big game hunting in New Zealand: per capita effort,harvest and expenditure in 2011–2012

Recreational big game hunters make a significant contribution to conservation through kills of deer, pigs, chamois and tahr. New opportunities for managing recreational hunting through the proposed Game Animal Council underscore the need to understand the implications of potential changes in recreational hunting participation and harvests. Based on a survey of hunters' recall over a year, hunters averaged 15.63 (SEM = 0.58) big game hunts per year, spending 30.53 (SEM = 0.85) days hunting and killing 8.92 (SEM = 0.69) big game animals. Hunters commonly targeted several species on a single hunt, with highly skewed distributions for hunter effort and kills. Mean monthly expenditure on big game hunting items was $296.78 (SEM = $8.95). Results demonstrate that big game hunting is a significant activity in New Zealand, but this varies considerably among hunters with a small number responsible for the vast majority of kills. These are important considerations for future big game hunting management.     

Expression of AMPA receptor subunits at synapses in laminae I–III of the rodent spinal dorsal horn

Neuropathic pain may arise following peripheral nerve injury though the molecular mechanisms associated with this are unclear. We used proteomic profiling to examine changes in protein expression associated with the formation of hyper-excitable neuromas derived from rodent saphenous nerves. A two-dimensional difference gel electrophoresis (2D-DIGE) profiling strategy was employed to examine protein expression changes between developing neuromas and normal nerves in whole tissue lysates. We found around 200 proteins which displayed a >1.75-fold change in expression between neuroma and normal nerve and identified 55 of these proteins using mass spectrometry. We also used immunoblotting to examine the expression of low-abundance ion channels Nav1.3, Nav1.8 and calcium channel α2δ-1 subunit in this model, since they have previously been implicated in neuronal hyperexcitability associated with neuropathic pain. Finally, S³⁵methionine in vitro labelling of neuroma and control samples was used to demonstrate local protein synthesis of neuron-specific genes. A number of cytoskeletal proteins, enzymes and proteins associated with oxidative stress were up-regulated in neuromas, whilst overall levels of voltage-gated ion channel proteins were unaffected. We conclude that altered mRNA levels reported in the somata of damaged DRG neurons do not necessarily reflect levels of altered proteins in hyper-excitable damaged nerve endings. An altered repertoire of protein expression, local protein synthesis and topological re-arrangements of ion channels may all play important roles in neuroma hyper-excitability.     

Trichogynes and Fertilization in Uni- and Bimating Type Colonies of Neurospora tetrasperma

Microscopic examination of living, protoperithecium-bearing colonies of a, A, and a + A that have been challenged by macroconidia from the same three colony types has shown that active trichogynes, i.e., those that grow to and fuse with a conidium, are to be found only in the first two types. Thus, in the a colony the trichogynes respond to conidia from the A and a + A colonies while in the A colony they respond to conidia from a and a + A colonies. In contrast to this ability of conidia from a + A colonies to function as fertilizing elements, the trichogynes of these colonies, if indeed they are formed at all, do not so respond. This nonresponse in a + A colonies may be due to the perithecia that are developing at the time of the challenge. Evidence for this conclusion comes from unimating type colonies in which the two halves of each colony were challenged at different times, 48 h apart. Trichogynes and perithecia developed in the first half; neither developed in the second. This inhibition of trichogyne development and response in the presence of developing perithecia may be only one manifestation of a more general inhibitory action by these structures.     

An Evaluation of Methods for Inferring Boolean Networks from Time-Series Data

Regulatory networks play a central role in cellular behavior and decision making. Learning these regulatory networks is a major task in biology, and devising computational methods and mathematical models for this task is a major endeavor in bioinformatics. Boolean networks have been used extensively for modeling regulatory networks. In this model, the state of each gene can be either ‘on’ or ‘off’ and that next-state of a gene is updated, synchronously or asynchronously, according to a Boolean rule that is applied to the current-state of the entire system. Inferring a Boolean network from a set of experimental data entails two main steps: first, the experimental time-series data are discretized into Boolean trajectories, and then, a Boolean network is learned from these Boolean trajectories. In this paper, we consider three methods for data discretization, including a new one we propose, and three methods for learning Boolean networks, and study the performance of all possible nine combinations on four regulatory systems of varying dynamics complexities. We find that employing the right combination of methods for data discretization and network learning results in Boolean networks that capture the dynamics well and provide predictive power. Our findings are in contrast to a recent survey that placed Boolean networks on the low end of the “faithfulness to biological reality” and “ability to model dynamics” spectra. Further, contrary to the common argument in favor of Boolean networks, we find that a relatively large number of time points in the time-series data is required to learn good Boolean networks for certain data sets. Last but not least, while methods have been proposed for inferring Boolean networks, as discussed above, missing still are publicly available implementations thereof. Here, we make our implementation of the methods available publicly in open source at http://bioinfo.cs.rice.edu/.     

Calcium‐permeable AMPA receptors and silent synapses in cocaine‐conditioned place preference

Exposure to cocaine generates silent synapses in the nucleus accumbens (NAc), whose eventual unsilencing/maturation by recruitment of calcium‐permeable AMPA‐type glutamate receptors (CP‐AMPARs) after drug withdrawal results in profound remodeling of NAc neuro‐circuits. Silent synapse‐based NAc remodeling was shown to be critical for several drug‐induced behaviors, but its role in acquisition and retention of the association between drug rewarding effects and drug‐associated contexts has remained unclear. Here, we find that the postsynaptic proteins PSD‐93, PSD‐95, and SAP102 differentially regulate excitatory synapse properties in the NAc. Mice deficient for either of these scaffold proteins exhibit distinct maturation patterns of silent synapses and thus provided instructive animal models to examine the role of NAc silent synapse maturation in cocaine‐conditioned place preference (CPP). Wild‐type and knockout mice alike all acquired cocaine‐CPP and exhibited increased levels of silent synapses after drug‐context conditioning. However, the mice differed in CPP retention and CP‐AMPAR incorporation. Collectively, our results indicate that CP‐AMPAR‐mediated maturation of silent synapses in the NAc is a signature of drug–context association, but this maturation is not required for establishing or retaining cocaine‐CPP.