Cardiac contractility modulation by electric currents applied during the refractory period

Inotropic effects of electric currents applied during the refractory period have been reported in cardiac muscle in vitro using voltage-clamp techniques. We investigated how electric currents modulate cardiac contractility in normal canine hearts in vivo. Six dogs were instrumented to measure regional segment length, ventricular volume (sonomicrometry), and ventricular pressure. Cardiac contractility modulating (CCM) electric currents (biphasic square pulses, amplitude +/-20 mA, total duration 30 ms) were delivered during the refractory period between pairs of electrodes placed on anterior and posterior walls. CCM significantly increased index of global contractility (E(es)) from 5.9 +/- 2.9 to 8.3 +/- 4.6 mmHg/ml with anterior CCM, from 5.3 +/- 1.8 to 8.9 +/- 4.0 mmHg/ml with posterior CCM, and from 6.1 +/- 2.6 to 11.0 +/- 7.0 mmHg/ml with combined CCM (P < 0.01, no significant change in volume axis intercept). End-systolic pressure-segment length relations showed contractility enhancement near CCM delivery sites, but not remotely. Relaxation was not influenced. CCM increased mean aortic pressure, but did not change peripheral resistance. Locally applied electrical currents enhanced global cardiac contractility via regional changes in myocardial contractility without impairing relaxation in situ.     

Isolation and structure elucidation of Peronosporomycetes hyphal branching-inducing factors produced by Pseudomonas jessenii EC-S101

Pseudomonas jessenii EC-S101 produced hyphal branching-inducing and mitosis-accelerating factors active towards Peronosporomycetes, Aphanomyces cochlioides hyphae. In searching for the active substances, EtOAc-solubles extracted from EC-S101-cultured solid medium were fractionated under the guidance of a paper disc assay using an A. cochlioides mycelium. Two active substances were subsequently isolated and the structure was elucidated by spectroscopic analysis to be (+)-4,5-didehydroacaterin (1) and 3-[(1R)-hydroxyhexyl]-5-methylene-2(5H)-furanone (2), both of which accelerated the mitotic process of A. cochlioides hyphae along with excessive branching at 1.0 microg per disc. These compounds are likely to affect the morphophysiological development of certain eukaryotic organisms in the terrestrial ecosystem.     

Skewed reproductive success among male white-spotted charr land-locked by an erosion control dam: Implications for effective population size

In 1996 and 1997, the spawning behavior of fluvial white-spotted charr Salvelinus leucomaenis, was observed in the upstream area of an erosion control dam. A small number of males with relatively large body size mated successfully with females as a pair, while almost all satellite males did not sneak successfully, resulting in a non-random mating system. The low sneaking success of subordinate males, in addition to the monopolization of spawning opportunities by a few dominant males, is one of the most important causes of skewed reproductive success among males. The total number of adult fishes in the study area (N: approximately half of the whole tributary above a dam) was estimated as 148 and 102 in 1996 and 1997, respectively. Based on these findings and some further assumptions, the estimated effective population size (N_e) was low in both years. The N_e/N ratio ranged from 0.33 to 0.36 in both years. In addition to reduced population size by construction of an impassable dam, the above-dam population suffered low N_e due to skewed reproductive success among males. The low N_e may be one cause of extinction in above-dam populations of fluvial charr, especially just after the construction of impassable barriers.     

Activation of smooth muscle and myenteric plexus cells of jejunum via Toll-like receptor 4

The cell types of the gut expressing Toll-like receptor 4, which recognizes specifically bacterial lipopolysaccharides, as well as the functionality of this receptor, have remained controversial. We aimed to clarify these issues. Mouse and human intestinal specimens were stained immunohistochemically to detect Toll-like receptor 4 expression. Smooth muscle and myenteric plexus cells but not enterocytes revealed receptor expression. Murine intestinal smooth muscle and myenteric plexus cells but not enterocytes showed nuclear translocation of nuclear factor-kappaB after in vivo stimulation with lipopolysaccharide. Moreover, lipopolysaccharide added to human jejunum biopsies free of epithelial cells induced release of interleukin-8 (IL-8). We can conclude that Toll-like receptor 4 is not expressed in epithelial layer, but rather on smooth muscle and myenteric plexus cells and that expression is functional. The expression of Toll-like receptor 4 on smooth muscle and myenteric plexus cells is consistent with the possibility that these cells are involved in intestinal immune defense; the low or absent expression of Toll-like receptor 4 on enterocytes might explain the intestinal epithelium hyporesponsiveness to the abundance of LPS in the intestinal lumen.     

Intrasexual copulation and mate discrimination in a population of Nodilittorina radiata (Gastropoda: Littorinidae)

Intrasexual copulation and mate discrimination by Nodilittorina radiata (Gastropoda: Littorinidae) were studied on a concrete breakwater at Hakodate Bay, southern Hokkaido, Japan. Intrasexual (male–male) copulations were observed in 4.7–21.1% of copulating pairs on the shore. As females were relatively larger than males and males copulated with females larger than themselves, we hypothesized that males choose potential mates larger than themselves. However, two male mates showed no significant size preference in intrasexual copulations, suggesting that males do not choose relatively larger individuals as mates. In a laboratory mate-choice experiment, male N. radiata preferred to mate with females, indicating precopulatory sex identification. They copulated with males, however, at the frequency of 37%, perhaps because of sex misidentification.     

Functional domains essential for Gs activity in prostaglandin EP2 and EP3 receptors

The interaction of cell surface hormone receptors with heterotrimeric G proteins is crucial for hormonal actions. The domains of the receptor, which interact with and activate G protein, have been extensively studied. However, precise molecular mechanisms underlying regulation of the receptor-induced G protein activation are still poorly understood. Prostaglandin E(2) (PGE(2)) receptors comprise of four subtypes, EP1, EP2, EP3 and EP4. Among them, EP2 and EP4 couple to Gs and EP3 to Gi. To assess the functional domains essential for Gs activation in prostanoid receptors, EP2, EP3beta and each intracellular loop- (IC-) interchanged EP2/EP3 chimeras were tested for agonist binding and functional responses. In EP2 receptor, substitution of IC1 or IC3 resulted in loss of binding activity, while substitution of IC2, N- (IC2N) or C-terminal half region of IC2 (IC2C) had no effects on the binding activity. Wild-type EP2 and IC2C-substituted EP2 showed agonist-induced Gs activity, but IC2- and IC2N-substituted EP2 failed to elicit Gs activity upon agonist stimulation. On the other hand, in EP3 receptor substitution of IC1 resulted in loss of PGE(2) binding, while substitution of IC2, IC3, IC2N or IC2C had no effects on binding activity. Wild-type EP3beta, IC3- or IC2C-substituted EP3 failed to show Gs activity upon agonist stimulation, but IC2- or IC2N-substituted EP3 chimera showed agonist-dependent Gs activity. These results indicated that the second intracellular loop of the EP2 plays an essential role in activation of Gs.     

Decrease of hepatic delta-aminolevulinate dehydratase activity in an animal model of fatigue

Fatigue can be defined physiologically as inability to maintain the expected power output. At present, no standard of fatigue are yet available. In order to find biomarkers of fatigue, we investigated the level of delta-aminolevulinic acid (ALA), the first intermediate metabolite in the heme biosynthetic pathway, in the plasma and urine of an animal model of fatigue. To prepare fatigued animals, we kept rats for 5 days in a cage filled with water to a height of 1.5 cm. As a result, the plasma and urinary ALA levels were increased in the fatigued animals as compared with those in the control animals. One day after the rats had been returned to their normal cages, these increased levels were restored to the control ones. We also examined the activity of the enzyme ALA dehydratase (ALAD), which is the second enzyme in the heme biosynthetic pathway, and ALAD gene expression during the fatigue and its recovery sessions. The ALAD activity, as well as its gene expression, in the liver of the fatigued animals was decreased as compared with those of the control animals. Both activity and gene expression of ALAD were recovered to their respective control levels after the rats had been allowed to rest in their normal cages for 1 day. Furthermore, the activity of ALA synthase (ALAS), the rate-limiting enzyme in the heme biosynthesis, in the liver was increased after the fatigue session for 5 days. Although this level of increase in the plasma concentration of ALA may not induce fatigue, increase in plasma and urinary ALA levels can be biomarkers of fatigue.     

Fabrication of Biomimetic Bone Tissue Using Mesenchymal Stem Cell-Derived Three-Dimensional Constructs Incorporating Endothelial Cells

The development of technologies to promote vascularization of engineered tissue would drive major developments in tissue engineering and regenerative medicine. Recently, we succeeded in fabricating three-dimensional (3D) cell constructs composed of mesenchymal stem cells (MSCs). However, the majority of cells within the constructs underwent necrosis due to a lack of nutrients and oxygen. We hypothesized that incorporation of vascular endothelial cells would improve the cell survival rate and aid in the fabrication of biomimetic bone tissues in vitro. The purpose of this study was to assess the impact of endothelial cells combined with the MSC constructs (MSC/HUVEC constructs) during short- and long-term culture. When human umbilical vein endothelial cells (HUVECs) were incorporated into the cell constructs, cell viability and growth factor production were increased after 7 days. Furthermore, HUVECs were observed to proliferate and self-organize into reticulate porous structures by interacting with the MSCs. After long-term culture, MSC/HUVEC constructs formed abundant mineralized matrices compared with those composed of MSCs alone. Transmission electron microscopy and qualitative analysis revealed that the mineralized matrices comprised porous cancellous bone-like tissues. These results demonstrate that highly biomimetic bone tissue can be fabricated in vitro by 3D MSC constructs incorporated with HUVECs.