Conductance and selectivity fluctuations in D127 mutants of the bacterial porin OmpF

A recent molecular dynamics study questioned the protonation state and physiological role of aspartate 127 (D127) of E. coli porin OmpF. To address that question we isolated two OmpF mutants with D127 either neutralized (D127N) or replaced by a positively charged lysine (D127K). The charge state of the residue at position 127 has clear effects on both conductance and selectivity. The D127K but not the D127N mutant expresses resilient conductance and selectivity fluctuations. These fluctuations reflect, we think, either changes in the ionization state of K127 and/or transitions between unstable subconformations as induced by the electrostatic repulsion between two positively charged residues, K127 and the nearby R167. Our results slightly favor the view that in WT OmpF residue D127 is deprotonated. As for the role of D127 in OmpF functionality, the gating of both mutants shows very similar sensitivity toward voltage as WT OmpF. Moreover, the current fluctuations of the D127K mutant were observed also in the absence of an applied electric field. We therefore dismiss D127 as a key residue in the control mechanism of the voltage-dependent gating of OmpF.     

Spawning habitat selection and suitability for Japanese dace, Tribolodon hakonensis

We have investigated the effects of three current velocities and three substrate sizes (gravel to cobble) on the spawning behavior of Japanese dace, Tribolodon hakonensis, in spawning-induction and habitat-selection experiments. In the spawning-induction experiment under laboratory conditions the number of females induced to spawn was significantly fewer at low current velocity (ca. 5 cm s^–1) than at medium (ca. 30 cm s^–1) or high (ca. 50 cm s^–1) current velocity. Females spawned independently of substrate size in the experimental tank, and did not bury their eggs in the substrate bed. In the habitat-selection experiment under field conditions, females selected the substrate microhabitat and spawned more frequently at a site with medium substrate size (very coarse gravel; major axis ca. 40 mm) than at sites with small (fine gravel; ca. 8 mm) or large (cobble; ca. 70 mm) substrate size. We conclude that Japanese dace select spawning sites at least partly on the basis of current velocity and substrate size, which affects the survival rate of eggs.     

Allosteric modulation of Ca2+ flux in ligand-gated cation channel (P2X4) by actions on lateral portals

Human P2X receptors are a family of seven ATP-gated ion channels that transport Na(+), K(+), and Ca(2+) across cell surface membranes. The P2X4 receptor is unique among family members in its sensitivity to the macrocyclic lactone, ivermectin, which allosterically modulates both ion conduction and channel gating. In this paper we show that removing the fixed negative charge of a single acidic amino acid (Glu(51)) in the lateral entrance to the transmembrane pore markedly attenuates the effect of ivermectin on Ca(2+) current and channel gating. Ca(2+) entry through P2X4 receptors is known to trigger downstream signaling pathways in microglia. Our experiments show that the lateral portals could present a novel target for drugs in the treatment of microglia-associated disease including neuropathic pain.     

Pelagic marine refugia and climatically sensitive areas in an eastern boundary current upwelling system

Refugia are areas relatively buffered from contemporary climate change that enable the persistence of valued physical, ecological, or sociocultural resources. Spatially identifying refugia is important for conservation and applied management. Yet the concept of refugia has not been broadly extended to marine ecosystems. Here, we analyze data from a unique and long‐term (1999–2015) standardized survey of pelagic marine and anadromous species off Oregon and Washington in the northern California Current to identify such refugia. We use quantitative approaches to assess locations with high species richness and community persistence relative to local and basin‐scale environmental fluctuations. We have identified a potential climate change refugial zone along the continental shelf of Washington State in the Northeastern Pacific Ocean, characterized by a species‐rich community with low interannual temporal community change. This region contrasts with adjacent areas to the south and offshore that have lower species richness, and higher temporal species community change. Also, using spatially variant generalized additive mixed models, we identify areas with species compositions that are more influenced by basin‐scale climatic fluctuations than others. We propose that upwelling regions with retentive topographic features, such as wide continental shelves, can function as marine refugia for pelagic fauna, whereas offshore locations are potentially more climatically sensitive and experience high temporal change in species composition. Further identification of these marine refugia using in situ data for pelagic biodiversity and climatically sensitive areas can help guide management in the face of inevitable climatically driven change.     

Ion permeation of pores in model membranes: selectivity,fluctuations and the role of surface charge

Fluctuation of surface charge on pore walls provides a realistic, additional mechanism for generating fluctuation of ionic current and ionic selectivity in narrow pores.     

Ciliary neurotrophic factor promotes inactivation of muscle Ca2+ channels via PKC

The actions of the ciliary neurotrophic factor (CNTF) were assessed on adult mouse skeletal muscle L-type Ca2+ currents and on Ca2+ release from sarcoplasmic reticulum. Currents were measured with the whole cell patch clamp technique. Ca2+ signals in response to single action potentials were recorded with Fluo3-AM. CNTF (20 ng/ml) reversibly reduced the amplitude of Ca2+ channel currents by 50% within 15 min. In addition, CNTF greatly increased the rate of inactivation during depolarizing pulses and shifted the steady state inactivation curve by -12 mV. The effects of CNTF were mimicked by the PKC activator PMA and prevented by the PKC-inhibitor chelerythrine. In contrast to the effects on the Ca2+ conductance, charge movement and Ca2+ signals remained unaffected by CNTF. These results suggest that CNTF can rapidly decrease muscle Ca2+ channel currents by promoting inactivation, probably through an intracellular PKC-dependent mechanism.     

Serum indole-3-acetic acid in control subjects and newly abstinent alcoholics after an oral loading with L-tryptophan: a preliminary study using liquid chromatography with amperometric detection

Described in this paper is a rapid, isocratic assay for serum indole-3-acetic acid (IAA). The sample preparation involves only protein precipitation using sulfosalicylic acid, and the sensitivity of amperometric detection is in the picogram range. The chromatographic analysis time is approximately 4 min. The devised method was used for a longitudinal study of IAA levels in serum samples from control subjects and newly abstinent alcoholics. Dietary variations were eliminated by administering a 2.0-g loading dose of L-Trp to all subjects investigated. The results are presented in the form of cumulative frequency polygons. Preliminary data indicate no differences in IAA levels between newly abstinent alcoholics and control subjects.     

Species-specific genetic markers for identification of early life-history stages of Cape hakes, Merluccius capensis and Merluccius paradoxus in the southern Benguela Current

The correct identification of fish egg and larval stages is crucial for inferring spawning areas and subsequent dispersal routes for marine fishes. The authors use species-specific mtDNA polymerase chain reaction to estimate proportions of Merluccius capensis and Merluccius paradoxus eggs and larvae and to elucidate early life stage distribution patterns in the southern Benguela system.