Spatial Analysis of Slowly Oscillating Electric Activity in the Gut of Mice Using Low Impedance Arrayed Microelectrodes

Smooth and elaborate gut motility is based on cellular cooperation, including smooth muscle, enteric neurons and special interstitial cells acting as pacemaker cells. Therefore, spatial characterization of electric activity in tissues containing these electric excitable cells is required for a precise understanding of gut motility. Furthermore, tools to evaluate spatial electric activity in a small area would be useful for the investigation of model animals. We thus employed a microelectrode array (MEA) system to simultaneously measure a set of 8×8 field potentials in a square area of ∼1 mm². The size of each recording electrode was 50×50 µm², however the surface area was increased by fixing platinum black particles. The impedance of microelectrode was sufficiently low to apply a high-pass filter of 0.1 Hz. Mapping of spectral power, and auto-correlation and cross-correlation parameters characterized the spatial properties of spontaneous electric activity in the ileum of wild-type (WT) and W/W^v mice, the latter serving as a model of impaired network of pacemaking interstitial cells. Namely, electric activities measured varied in both size and cooperativity in W/W^v mice, despite the small area. In the ileum of WT mice, procedures suppressing the excitability of smooth muscle and neurons altered the propagation of spontaneous electric activity, but had little change in the period of oscillations. In conclusion, MEA with low impedance electrodes enables to measure slowly oscillating electric activity, and is useful to evaluate both histological and functional changes in the spatio-temporal property of gut electric activity.     

Relationship between interstitial cells of Cajal, fibroblast-like cells and inhibitory motor nerves in the internal anal sphincter

Interstitial cells of Cajal (ICC) have been shown to participate in nitrergic neurotransmission in various regions of the gastrointestinal (GI) tract. Recently, fibroblast-like cells, which are positive for platelet-derived growth factor receptor ?? (PDGFR??⁺), have been suggested to participate additionally in inhibitory neurotransmission in the GI tract. The distribution of ICC and PDGFR??⁺ cell populations and their relationship to inhibitory nerves within the mouse internal anal sphincter (IAS) are unknown. Immunohistochemical techniques and confocal microscopy were therefore used to examine the density and arrangement of ICC, PDGFR??⁺ cells and neuronal nitric-oxide-synthase-positive (nNOS⁺) nerve fibers in the IAS of wild-type (WT) and W/W ^v mice. Of the total tissue volume within the IAS circular muscle layer, 18% consisted in highly branched PDGFR??⁺ cells (PDGFR??⁺-IM). Other populations of PDGFR??⁺ cells were observed within the submucosa and along the serosal and myenteric surfaces. Spindle-shaped intramuscular ICC (ICC-IM) were present in the WT mouse IAS but were largely absent from the W/W ^v IAS. The ICC-IM volume (5% of tissue volume) in the WT mouse IAS was significantly smaller than that of PDGFR??⁺-IM. Stellate-shaped submucosal ICC (ICC-SM) were observed in the WT and W/W ^v IAS. Minimum surface distance analysis revealed that nNOS⁺ nerve fibers were closely aligned with both ICC-IM and PDGFR??⁺-IM. An even closer association was seen between ICC-IM and PDGFR??⁺-IM. Thus, a close morphological arrangement exists between inhibitory motor neurons, ICC-IM and PDGFR??⁺-IM suggesting that some functional interaction occurs between them contributing to inhibitory neurotransmission in the IAS.     

Roles of mast cells in the pathogenesis of inflammatory myopathy

Introduction

In addition to the pivotal roles of mast cells in allergic diseases, recent data suggest that mast cells play crucial roles in a variety of autoimmune responses. However, their roles in the pathogenesis of autoimmune skeletal muscle diseases have not been clarified despite their distribution in skeletal muscle. Therefore, the objective of this study is to determine the roles of mast cells in the development of autoimmune skeletal muscle diseases.

Methods

The number of mast cells in the affected muscle was examined in patients with dermatomyositis (DM) or polymyositis (PM). The susceptibility of mast cell-deficient WBB6F1-Kit^W/Kit^Wv mice (W/W^v mice) to a murine model of polymyositis, C protein-induced myositis (CIM), was compared with that of wild-type (WT) mice. The effect of mast cell reconstitution with bone marrow-derived mast cells (BMMCs) on the susceptibility of W/W^v mice to CIM was also evaluated.

Results

The number of mast cells in the affected muscle increased in patients with PM as compared with patients with DM. W/W^v mice exhibited significantly reduced disease incidence and histological scores of CIM as compared with WT mice. The number of CD8⁺ T cells and macrophages in the skeletal muscles of CIM decreased in W/W^v mice compared with WT mice. Engraftment of BMMCs restored the incidence and histological scores of CIM in W/W^v mice. Vascular permeability in the skeletal muscle was elevated in WT mice but not in W/W^v mice upon CIM induction.

Conclusion

Mast cells are involved in the pathogenesis of inflammatory myopathy.     

Nav1.7 sodium channel-induced Ca influx decreases tau phosphorylation via glycogen synthase kinase-3β in adrenal chromaffin cells

In cultured bovine adrenal chromaffin cells expressing Na_v1.7 sodium channel isoform, veratridine increased Ser⁴⁷³-phosphorylation of Akt and Ser⁹-phosphorylation of glycogen synthase kinase-3β by 217 and 195%, while decreasing Ser³⁹⁶-phosphorylation of tau by 36% in a concentration (EC₅₀ = 2.1 μM)- and time (t_1/2 = 2.7 min)-dependent manner. These effects of veratridine were abolished by tetrodotoxin or extracellular Ca²⁺ removal. Veratridine (10 μM for 5 min) increased translocation of Ca²⁺-dependent conventional protein kinase C-α from cytoplasm to membranes by 47%; it was abolished by tetrodotoxin, extracellular Ca²⁺ removal, or Gö6976 (an inhibitor of protein kinase C-α), and partially attenuated by LY294002 (an inhibitor of phosphatidylinositol 3-kinase). LY294002 (but not Gö6976) abrogated veratridine-induced Akt phosphorylation. In contrast, either LY294002 or Gö6976 alone attenuated veratridine-induced glycogen synthase kinase-3β phosphorylation by 65 or 42%; however, LY294002 plus Gö6976 completely blocked it. Veratridine (10 μM for 5 min)-induced decrease of tau phosphorylation was partially attenuated by LY294002 or Gö6976, but completely blocked by LY294002 plus Gö6976; okadaic acid or cyclosporin A (inhibitors of protein phosphatases 1, 2A, and 2B) failed to alter tau phosphorylation. These results suggest that Na⁺ influx via Na_v1.7 sodium channel and the subsequent Ca²⁺ influx via voltage-dependent calcium channel activated (1) Ca²⁺/protein kinase C-α pathway, as well as (2) Ca²⁺/phosphatidylinositol 3-kinase/Akt and (3) Ca²⁺/phosphatidylinositol 3-kinase/protein kinase C-α pathways; these parallel pathways converged on inhibitory phosphorylation of glycogen synthase kinase-3β, decreasing tau phosphorylation.     

Western equatorial Pacific planktic foraminiferal fluxes and assemblages during a La Niña year (1999)

A correlation between foraminiferal community dynamics and environmental conditions may provide a basis for establishing paleoclimatic proxies. We studied planktic foraminiferal shell fluxes and assemblages in samples collected in three time-series sediment trap deployments in the western equatorial Pacific under La Niña conditions from January to November 1999. Eleven species contributed about 90% of the total flux in all traps. Two sites (MT1, MT3) in the Western Pacific Warm Pool region (WPWP) were characterized by common occurrences of the species Globigerinoides ruber, Globigerinoides sacculifer, Globigerinoides tenellus, and Neogloboquadrina dutertrei. Site MT5 farther to the east in the equatorial upwelling region had common occurrences of Globigerina bulloides, Globigerinita glutinata, and Pulleniatina obliquiloculata. Very high abundances of G. bulloides and G. glutinata at MT5 indicate that equatorial upwelling (EU) occurred during the 1999 La Niña. The two western sites have similar assemblage compositions, but MT1 ( 135°E) has the highest fluxes (up to  3800 tests m^− 2 day^− 1), whereas MT3 ( 145° E) has fluxes below  2200 tests m^− 2 day^− 1. Relatively high fluxes (up to  3000 tests m^− 2 day^− 1) occur at site MT5 ( 176° E), where upwelling occurred.The differences in faunal composition in the WPWP and EU might be attributable to differences in the way in which nutrients are supplied to the phytoplankton: large amounts of suspended material are supplied to the WPWP by advection of waters passing through the coastal region of an archipelago, whereas upwelling of nutrient-rich waters enhances primary production in the EU. At the westernmost site in the WPWP, a peak in the G. bulloides flux coincided with southward flow of the New Guinea Coastal Current (NGCC) in late February, but the highest G. ruber flux coincided with northward flow of this current in late May. Thus, the differences in species dominance at this location may be caused by monsoon-driven variability in the flow direction of the NGGC.     

Anti-arrhythmic effects of cyclopiazonic acid in Langendorff-perfused murine hearts

We investigated the effects of reducing sarcoplasmic reticular (SR) Ca²⁺ stores using the Ca²⁺-ATPase inhibitor cyclopiazonic acid (CPA) in Langendorff-perfused mouse hearts exposed to different pro-arrhythmic agents all known to produce Ca²⁺-mediated arrhythmogenesis. CPA (100 and 150 nM) produced progressive (beginning over 1 min) and significant (P < 0.0001) reductions in peak amplitudes of Ca²⁺ transients evoked by regular stimulation in isolated Fluo-3 loaded myocytes from F/F₀ = 3.2 ± 0.16 (n = 12 cells) to 1.62 ± 0.012 (n = 6 cells) and 1.53 ± 0.06 (n = 12 cells), respectively, consistent with previous reports describing reductions of store Ca²⁺ in other cell systems. The corresponding effects of CPA were then examined in intact hearts exposed to isoproterenol (100 nM), elevated extracellular [Ca²⁺] (5 mM) and caffeine (1 mM). All three agents produced ventricular tachycardia either when added alone or simultaneously with CPA during programmed electrical stimulation. However, arrhythmogenicity was not observed when such agents were added 10 min after introduction of CPA. CPA thus antagonized this Ca²⁺-mediated arrhythmogenesis but only under circumstances of SR Ca²⁺ depletion. These alterations in arrhythmogenic tendency took place despite an absence of alterations in electrogram and monophasic action potential characteristics. This was in sharp contrast to previous observations in murine, ΔKPQ-Scn5a (LQT3) and KCNE1^−/− (LQT5), systems where re-entry has been implicated in arrhythmogenesis.     

Decomposition of four macrophytes in wetland sediments: Organic matter and nutrient decay and associated benthic processes

Decomposition rates of Phragmites australis, Carex riparia, Nuphar luteum and Salvinia natans and benthic processes were measured from December 2003 to December 2004 in a shallow wetland (Paludi di Ostiglia, Northern Italy) by means of litter bags and intact cores incubations. Decay rate was highest for N. luteum (k = 0.0152 d⁻¹), intermediate for S. natans (k = 0.0041 d⁻¹) and similar for P. australis (k = 0.0027 d⁻¹) and C. riparia (k = 0.0028 d⁻¹).Benthic metabolism followed a seasonal pattern with summer peaks of O₂ demand and TCO₂, CH₄ and NH₄⁺ efflux whilst soluble reactive phosphorus (SRP) fluxes were negligible also under hypoxic conditions, indicating that P was mainly retained by sediment. The initial C:P ratio was similar in N. luteum and S. natans (170) and significantly lower than that of P. australis and C. riparia (360). During the detritus decay P was progressively lost by N. luteum and S. natans tissues, whereas, after an initial leaching, it was probably re-used during the microbial decomposition of the more refractory P. australis and C. riparia detritus. Nuphar luteum, P. australis and S. natans had comparable initial C:N mass ratio (15), significantly lower than that of C. riparia (26). The C:N ratio was rather constant for N. luteum (12.9 ± 1.5) and S. natans (14.6 ± 0.9), decreased slightly to below 20 for C. riparia and increased up to 30 for P. australis. Overall, differences among species were likely due to the recalcitrance of decomposing detritus, whilst process rates were controlled by limitation of microbial processes by nutrients and electron acceptor availability.     

Graphitized macroporous carbon microarray with hierarchical mesopores as host for the fabrication of electrochemical biosensor

A novel graphitized ordered macroporous carbon (GMC, pore size 380 nm) with hierarchical mesopores (2–30 nm) and high graphitization degree was prepared by nickel-catalyzed graphitization of polystyrene arrays. The obtained GMC possessed high specific surface area, large pore volume, and good electrical conductivity, which was explored for the enzyme entrapment and biosensor fabrication by a facile method. With advantages of novel nanostructure and good electrical conductivity, direct electrochemistry of hemoglobin (a model protein) was observed on the GMC-based biocomposite with a formal potential of −0.36 V (vs. Ag/AgCl) and an apparent heterogeneous electron transfer rate constant (k_s) of 1.2 s⁻¹ in pH 7.0 buffer. Comparative studies revealed that GMC offered significant advantages over carbon nanotubes (CNTs) in facilitating direct electron transfer of entrapped Hb. The fabricated biosensor exhibited good sensitivity (101.6 mA cm⁻² M⁻¹) and reproducibility, wide linear range (1–267 μM), low detection limit (0.1 μM), and good long-term stability for H₂O₂ detection. GMC proved to be a promising matrix for enzyme entrapment and biosensor fabrication, and may find wide potential applications in biomedical detection and environmental analyses.     

Growth phase-dependant enzyme profile of pyruvate catabolism and end-product formation in Clostridium thermocellum ATCC 27405

End-product synthesis and enzyme activities involved in pyruvate catabolism, H₂ synthesis, and ethanol production in mid-log (OD₆₀₀  0.25), early stationary (OD₆₀₀  0.5), and stationary phase (OD₆₀₀  0.7) cell extracts were determined in Clostridium thermocellum ATCC 27405 grown in batch cultures on cellobiose. Carbon dioxide, hydrogen, ethanol, acetate and formate were major end-products and their production paralleled growth and cellobiose consumption. Lactate dehydrogenase, pyruvate:formate lyase, pyruvate:ferredoxin oxidoreductase, methyl viologen-dependant hydrogenase, ferredoxin-dependant hydrogenase, NADH-dependant hydrogenase, NADPH-dependant hydrogenase, NADH-dependant acetaldehyde dehydrogenase, NADH-dependant alcohol dehydogenase, and NADPH-dependant alcohol dehydrogenase activities were detected in all extracts, while pyruate dehydrogenase and formate dehydrogenase activities were not detected. All hydrogenase activities decreased (2–12-fold) as growth progressed from early exponential to stationary phase. Alcohol dehydrogenase activities fluctuated only marginally (<45%), while lactate dehydrogenase, pyruvate:formate lyase, and pyruvate:ferredoxin oxidoreductase remained constant in all cell extracts. We have proposed a pathway involved in pyruvate catabolism and end-product formation based on enzyme activity profiles in conjunction with bioinformatics analysis.     

Histamine from Brain Resident MAST Cells Promotes Wakefulness and Modulates Behavioral States

Mast cell activation and degranulation can result in the release of various chemical mediators, such as histamine and cytokines, which significantly affect sleep. Mast cells also exist in the central nervous system (CNS). Since up to 50% of histamine contents in the brain are from brain mast cells, mediators from brain mast cells may significantly influence sleep and other behaviors. In this study, we examined potential involvement of brain mast cells in sleep/wake regulations, focusing especially on the histaminergic system, using mast cell deficient (W/W^v) mice. No significant difference was found in the basal amount of sleep/wake between W/W^v mice and their wild-type littermates (WT), although W/W^v mice showed increased EEG delta power and attenuated rebound response after sleep deprivation. Intracerebroventricular injection of compound 48/80, a histamine releaser from mast cells, significantly increased histamine levels in the ventricular region and enhanced wakefulness in WT mice, while it had no effect in W/W^v mice. Injection of H1 antagonists (triprolidine and mepyramine) significantly increased the amounts of slow-wave sleep in WT mice, but not in W/W^v mice. Most strikingly, the food-seeking behavior observed in WT mice during food deprivation was completely abolished in W/W^v mice. W/W^v mice also exhibited higher anxiety and depression levels compared to WT mice. Our findings suggest that histamine released from brain mast cells is wake-promoting, and emphasizes the physiological and pharmacological importance of brain mast cells in the regulation of sleep and fundamental neurobehavior.     

Initiation of motility by steelhead (Oncorhynchus mykiss) sperm: membrane ion exchangers and pH sensitivity

Initiation of motility in salmonid sperm is sensitive to the pH of the extracellular medium, however, the basis of this sensitivity is not clear. Sperm incubated in an immobilization buffer (SI) at low pH ( 7.1–7.2) become motile when diluted with activating medium (AM) at high ( 8.5) but not low pH. Based on this observation, various agents were tested to determine whether the onset of steelhead sperm motility upon activation with high pH AM, following incubation with low pH SI, could be blocked by inhibiting membrane exchangers postulated to be important in intracellular pH (pHi) regulation. Amiloride (inhibitor of proton:sodium exchange), SITS and DIDS (inhibitors of anion exchange) and bafilomycin A 1 (inhibitor of H⁺-ATPase activity) were not effective in this experimental design. However, regardless of SI pH, DIDS was effective in blocking motility as was replacing chloride with thiocyanate or including the chloride channel blocker, niflumic acid, in SI suggesting that chloride efflux plays a key role in motility initiation. Nonetheless, the results of this study suggest that the rapid onset of sperm motility with activation at high pH following incubation at low pH is probably not based on rapid adjustment of pHi via membrane exchangers/transporters but rather due to an effect of pH on motility-associated processes at the extracellular surface of the sperm.     

The control of transmembrane helix transverse position in membranes by hydrophilic residues

The ability of hydrophilic residues to shift the transverse position of transmembrane (TM) helices within bilayers was studied in model membrane vesicles. Transverse shifts were detected by fluorescence measurements of the membrane depth of a Trp residue at the center of a hydrophobic sequence. They were also estimated from the effective length of the TM-spanning sequence, derived from the stability of the TM configuration under conditions of negative hydrophobic mismatch. Hydrophilic residues (at the fifth position in a 21-residue hydrophobic sequence composed of alternating Leu and Ala residues and flanked on both ends by two Lys) induced transverse shifts that moved the hydrophilic residue closer to the membrane surface. At pH 7, the dependence of the extent of shift upon the identity of the hydrophilic residue increased in the order: L < GYT < RH < S < P < K < EQ < N < D. By varying pH, shifts with ionizable residues fully charged or uncharged were measured, and the extent of shift increased in the order: L < GYH^oT < E^oR < S < P < K⁺< QD^oH⁺ < NE^– < D^–. The dependence of transverse shifts upon hydrophilic residue identity was consistent with the hypothesis that shift magnitude is largely controlled by the combination of side chain hydrophilicity, ionization state, and ability to position polar groups near the bilayer surface (snorkeling). Additional experiments showed that shift was also modulated by the position of the hydrophilic residue in the sequence and the hydrophobicity of the sequence moved out of the bilayer core upon shifting. Combined, these studies show that the insertion boundaries of TM helices are very sensitive to sequence, and can be altered even by weakly hydrophilic residues. Thus, many TM helices may have the capacity to exist in more than one transverse position. Knowledge of the magnitudes of transverse shifts induced by different hydrophilic residues should be useful for design of mutagenesis studies measuring the effect of transverse TM helix position upon function.     

Tissue-Specific Mathematical Models of Slow Wave Entrainment in Wild-Type and 5-HT2B Knockout Mice with Altered Interstitial Cells of Cajal Networks

Gastrointestinal slow waves are generated within networks of interstitial cells of Cajal (ICCs). In the intact tissue, slow waves are entrained to neighboring ICCs with higher intrinsic frequencies, leading to active propagation of slow waves. Degradation of ICC networks in humans is associated with motility disorders; however, the pathophysiological mechanisms of this relationship are uncertain. A recently developed biophysically based mathematical model of ICC was adopted and updated to simulate entrainment of slow waves. Simulated slow wave propagation was successfully entrained in a one-dimensional model, which contained a gradient of intrinsic frequencies. Slow wave propagation was then simulated in tissue models which contained a realistic two-dimensional microstructure of the myenteric ICC networks translated from wild-type (WT) and 5-HT_2B knockout (degraded) mouse jejunum. The results showed that the peak current density in the WT model was 0.49 μA mm⁻² higher than the 5-HT_2B knockout model, and the intracellular Ca²⁺ density after 400 ms was 0.26 mM mm⁻² higher in the WT model. In conclusion, tissue-specific models of slow waves are presented, and simulations quantitatively demonstrated physiological differences between WT and 5-HT_2B knockout models. This study provides a framework for evaluating how ICC network degradation may impair slow wave propagation and ultimately motility and transit.     

Quantitative analysis of DNA hybridization in a flowthrough microarray for molecular testing

Quantitative information about the nucleic acids hybridization reaction on microarrays is fundamental to designing optimized assays for molecular diagnostics. This study presents the kinetic, equilibrium, and thermodynamic analyses of DNA hybridization in a microarray system designed for fast molecular testing of pathogenic bacteria. Our microarray setup uses a porous, nylon membrane for probe immobilization and flowthrough incubation. The Langmuir model was used to determine the reaction rate constants of hybridization with antisense targets specific to Staphylococcus epidermidis and Staphylococcus aureus strains. The kinetic analysis revealed a sequence-dependent reaction rate, with association rate constants on the order of 10⁵ M⁻¹ s⁻¹ and dissociation rate constants of 10⁻⁴ s⁻¹. We found that by increasing the probe surface density from 10¹¹ to 10¹² molecules/cm², the hybridization rate and efficiency are suppressed while the melting temperature of the DNA duplex increases. The maximum fraction of hybridized capture probes at equilibrium did not exceed 50% for hybridization with antisense sequences and was below 6% for hybridization with long targets obtained from PCR. The van’t Hoff analysis of the temperature denaturation data showed that the DNA hybridization in our porous, flowthrough microarray is thermodynamically less favorable than the hybridization of the same sequences in solution.     

c-Kit-negative fibroblast-like cells express platelet-derived growth factor receptor α in the murine gastrointestinal musculature

Platelet-derived growth factor receptors (PDGFRs) belong to the same kinase group as c-Kit receptor tyrosine kinase that is specifically expressed in the interstitial cells of Cajal (ICC) in the gastrointestinal tract. In this study, we examined PDGFRα immunoreactivity in the murine gastrointestinal tract. PDGFRα-immunopositive (PDGFRα-ip) cells were observed in the musculature in all parts of the gastrointestinal tract. Although PDGFRα-ip cells were distinct from ICC and neurons, these cells were closely associated with intramuscular ICC and enteric nerve fibers. In the myenteric layer, PDGFRα-ip cells formed a cellular network with their ramified processes and encompassed myenteric ganglia. Numerous PDGFRα-ip cells were observed in the subserosal plane and showed a multipolar shape. The distribution pattern of the PDGFRα-ip cells in the ICC-deficient W ^v /W ^v mutant mice was the same as that in normal mice. PDGFRα-ip cells that showed intense immunoreactivity of SK3 potassium channel were considered to correspond to fibroblast-like cells or non-Cajal interstitial cells. Our observations suggest that PDGFRα-ip cells are basic cellular elements throughout the gastrointestinal musculature and are involved in the gastrointestinal functions.     

Structural elucidation of an exopolysaccharide from mycelial fermentation of a Tolypocladium sp. fungus isolated from wild Cordyceps sinensis

A novel polysaccharide designated EPS-1A with an average molecular weight around 40 kDa was fractionated and purified by anion-exchange and gel-filtration chromatography from the crude exopolysaccharide (EPS) isolated from fermentation broth of Cs-HK1, a Tolypocladium sp. fungus isolated from wild Cordyceps sinensis. The structural characteristics of EPS-1A were determined with various methods (e.g. GC, GC–MS, FT-IR, ¹H NMR and ¹³C NMR) and through acid hydrolysis, methylation, periodate-oxidation and Smith degradation. The results suggested that EPS-1A was composed of glucose, mannose and galactose at 15.2:3.6:1.0 M ratio. EPS-1A was a slightly branched polysaccharide and its backbone was composed of (1 → 6)-α-d-glucose residues (77%) and (1 → 6)-α-d-mannose residues (23%). Branching occurred at O-3 position of (1 → 6)-α-d-mannose residues of the backbone with (1 → 6)-α-d-mannose residues and (1 → 6)-α-d-glucose residues, and terminated with β-d-galactose residues.     

The Holocene Pulleniatina Minimum Event revisited: Geochemical and faunal evidence from the Okinawa Trough and upper reaches of the Kuroshio current

The Holocene Pulleniatina Minimum Event (PME) is characterized by a very low abundance of the planktonic foraminifer Pulleniatina obliquiloculata between  4.5 and 3 ka. The PME occurs widely in the Okinawa Trough and the South China Sea, and can be correlated throughout this area; it has been related to variability in the Kuroshio current. To further explore the nature of the PME, we studied cores obtained from the southern Okinawa Trough and the upper reaches of the Kuroshio current. Faunal census data indicate that all cores record the PME between  4.5 and  3 ka. The relative abundance of Neogloboquadrina dutertrei is negatively correlated to that of P. obliquiloculata in the southern Okinawa Trough, but not in the sites at the upper reaches. Mg/Ca and δ¹⁸O measurements on Globigerinoides ruber shells from the southern Okinawa Trough indicate that there was no change in sea surface temperature or sea surface salinity during the PME. The vertical structure of the water column as reconstructed by multispecies δ¹⁸O and δ¹³C profiles shows no consistent anomalies in the southern Okinawa Trough and western Philippine Sea during the PME. These observations suggest that: (1) the PME was not restricted to marginal seas, but widespread in the western North Pacific. (2) The high abundance of N. dutertrei during the PME in the Okinawa Trough may be a result of higher food-availability in the absence of P. obliquiloculata. (3) No distinctive, consistent anomalies in the paleoceanographic proxies are associated with the PME, implying there were no changes in hydrography and productivity. The absence of a linkage between faunal variation and paleoceanographic proxies indicates that we do not yet understand what causes changes in planktonic foraminiferal assemblages. This lack of understanding implies that we cannot always trust fauna-based paleothermometry at millennial timescales.     

Biphasic kinetics in the reaction between amino acids or glutathione and the chromium acetate cluster, [Cr3O(OAc)6]

Kinetics for the breakdown of the trinuclear chromium acetate cluster with a series of monoprotic and diprotic amino acid ligands and with glutathione in aqueous media have been investigated spectrophotometrically at pH 3.5–5.5 and in a temperature range of 45–60 °C. Under pseudo-first-order conditions, reactions with these ligands exhibited biphasic kinetic behavior that can be accounted for by a consecutive two-step reaction, A → B → C, where A is assumed to be a forced ion pair, B an intermediate and C is the product; experimental data fit to a biexponential equation for the transformation. Rates for k_short, k_long, and k_obs were determined by manual extrapolation of absorbance data or curve-fitting routines; associated activation parameters for each step of the reaction were calculated using the Eyring equation. Rates for the first and second steps of the reaction are on the order of 10⁻⁴ and 10⁻⁵ s⁻¹, respectively. The large negative values of ΔS^‡ and smaller ΔH^‡ in the first step indicate an associative step, while high positive values of ΔS^‡ in the second step indicate dissociation. To account for the results mechanistically, the results are interpreted to be a first step of ligand exchange with a pseudo-axial aqua ligand, followed by a dissociative step involving acetate or oxo ligand displacement. The dissociative step is the rate determining step, with k_obs ≈ k_long.The results demonstrate reaction pathways that are available to the Cr(III) metal centers that may be physiologically relevant in the ligand-rich environment of biological systems. Under general conditions Cr(III) clusters may be expected to be broken down, unless some unique biological environment stabilizes the cluster. The present study has application to the processes related to Cr(III) transport and excretion, to potential mechanisms of Cr(III) action in a biological setting, and to the pharmacokinetics of Cr(III) supplements for animal and human consumption.