Circulating Ouabain Modulates Expression of Claudins in Rat Intestine and Cerebral Blood Vessels

The ability of exogenous low ouabain concentrations to affect claudin expression and therefore epithelial barrier properties was demonstrated previously in cultured cell studies. We hypothesized that chronic elevation of circulating ouabain in vivo can affect the expression of claudins and tight junction permeability in different tissues. We tested this hypothesis in rats intraperitoneally injected with ouabain (1 μg/kg) for 4 days. Rat jejunum, colon and brain frontal lobes, which are variable in the expressed claudins and tight junction permeability, were examined. Moreover, the porcine jejunum cell line IPEC-J2 was studied. In IPEC-J2-cells, ouabain (10 nM, 19 days of incubation) stimulated epithelial barrier formation, increased transepithelial resistance and the level of cSrc-kinase activation by phosphorylation, accompanied with an increased expression of claudin-1, -5 and down-regulation of claudin-12; the expression of claudin-3, -4, -8 and tricellulin was not changed. In the jejunum, chronic ouabain increased the expression of claudin-1, -3 and -5 without an effect on claudin-2 and -4 expression. In the colon, only down-regulation of claudin-3 was observed. Chronic ouabain protected the intestine transepithelial resistance against functional injury induced by lipopolysaccharide treatment or by modeled acute microgravity; this regulation was most pronounced in the jejunum. Claudin-1 was also up-regulated in cerebral blood vessels. This was associated with reduction of claudin-3 expression while the expression of claudin-5 and occludin was not affected. Altogether, our results confirm that circulating ouabain can functionally and tissue-specifically affect barrier properties of epithelial and endothelial tissues via Na,K-ATPase-mediated modulation of claudins expression

P5A-Type ATPase Cta4p Is Essential for Ca2+ Transport in the Endoplasmic Reticulum of Schizosaccharomyces pombe

This study establishes the role of P5A-type Cta4 ATPase in Ca2+ sequestration in the endoplasmic reticulum by detecting an ATP-dependent, vanadate-sensitive and FCCP insensitive 45Ca2+-transport in fission yeast membranes isolated by cellular fractionation. Specifically, the Ca2+-ATPase transport activity was decreased in ER membranes isolated from cells lacking a cta4+ gene. Furthermore, a disruption of cta4+ resulted in 6-fold increase of intracellular Ca2+ levels, sensitivity towards accumulation of misfolded proteins in ER and ER stress, stimulation of the calcineurin phosphatase activity and vacuolar Ca2+ pumping. These data provide compelling biochemical evidence for a P5A-type Cta4 ATPase as an essential component of Ca2+ transport system and signaling network which regulate, in conjunction with calcineurin, the ER functionality in fission yeast

Microstimulation of human somatosensory cortex evokes task-dependent, spatially patterned responses in motor cortex

The primary motor (M1) and somatosensory (S1) cortices play critical roles in motor control but the signaling between these structures is poorly understood. To fill this gap, we recorded – in three participants in an ongoing human clinical trial (NCT01894802) for people with paralyzed hands – the responses evoked in the hand and arm representations of M1 during intracortical microstimulation (ICMS) in the hand representation of S1. We found that ICMS of S1 activated some M1 neurons at short, fixed latencies consistent with monosynaptic activation. Additionally, most of the ICMS-evoked responses in M1 were more variable in time, suggesting indirect effects of stimulation. The spatial pattern of M1 activation varied systematically: S1 electrodes that elicited percepts in a finger preferentially activated M1 neurons excited during that finger’s movement. Moreover, the indirect effects of S1 ICMS on M1 were context dependent, such that the magnitude and even sign relative to baseline varied across tasks. We tested the implications of these effects for brain-control of a virtual hand, in which ICMS conveyed tactile feedback. While ICMS-evoked activation of M1 disrupted decoder performance, this disruption was minimized using biomimetic stimulation, which emphasizes contact transients at the onset and offset of grasp, and reduces sustained stimulation

EFFECTS OF PLACENTAL FACTORS UPON DEVELOPMENT OF TUBULAR STRUCTURES BY ENDOTHELIAL CELLS IN PRESENCE OF TROPHOBLASTIC CELLS

Trophoblast cells actively interact with endothelial cells participating in the process of vasculogenesis in the uterus/placenta contact area and remodeling of uterine spiral arteries. Cytokine production by the placental cells is subject to gradual changes from the 1st to 3rd trimester of physiological pregnancy. It is also changed in cases of obstetric disorders, e.g., in pre-eclampsia. At present time, there are lacking data on effects of cytokines and placenta-derived factors upon local interactions between endothelium and trophoblast cells. Hence, the aim of our study was to assess the influence of placental factors upon formation of tube-like structures by endothelial cells in presence of trophoblastic cells. We performed co-cultures of Ea.Hy926 endothelial cell line and Jeg-3 trophoblastic cells in a 3-D collagen matrix («Matrigel», BD, USA) with secretable factors from placentas of healthy pregnant women at 9-11 weeks of gestation (n = 15), healthy pregnant women at 38-39 weeks of gestation (n = 15), or the women with preeclampsia at 38-39 weeks of gestation (n = 14). We have shown that the trophoblastic cells may modify the ability of endothelial cells to form tube-like structures only with placental factors from health pregnant women. In pre-eclampsia condition, the trophoblast cells are not able to correct the behavior of endothelial cells, and to promote physiological growth of blood vessels

Extracellular glucose increases the coupling capacity of the yeast V H+-ATPase and the resistance of its H+ transport activity to nitrate inhibition.

V H(+)-ATPase has an important role in a variety of key physiological processes. This enzyme is reversibly activated/partly inactivated by the addition/exhaustion of extracellular glucose. The current model of its regulation assumes the reversible disassembly/reassembly of ∼60-70% of the V1 and V0 membrane complexes, which are responsible for ATP hydrolysis and H(+) conductance, respectively. The number of assembled complexes determines the pump activity because disassembled complexes are inactive. The model predicts the identical catalytic properties for the activated and semi-active enzymes molecules. To verify the model predictions we have isolated total membranes from yeast spheroplasts that were pre-incubated either with or without glucose. Nitrate treatment of membranes revealed the similar ATPase inhibition for two enzyme states, suggesting that they have identical structures that are essential for ATP hydrolysis. However, H(+) transport was inhibited more than the ATPase activities, indicating a nitrate uncoupling action, which was significantly higher for the nonactivated enzyme. This finding suggests that the structure of the non-activated enzyme, which is essential for H(+) transport, is less stable than that of the activated enzyme. Moreover, the glucose activation of the pump increases i) its coupling capacity; ii) its K(M) for ATP hydrolysis and ATP affinity for H(+) transport; iii) the Vmax for H(+) transport in comparison with the Vmax for ATP hydrolysis and iv) the immune reactivity of catalytic subunit A and regulatory subunit B by 9.3 and 2.4 times, respectively. The protein content of subunits A and B was not changed by extracellular glucose. We propose that instead of the dissociation/reassociation of complexes V1 and V0, changes in the extracellular glucose concentration cause reversible and asymmetrical modulations in the immune reactivity of subunits A and B by their putative biochemical modifications. This response asymmetrically modulates H(+)-transport and ATP hydrolysis, exhibiting distinct properties for the activated versus non-activated enzymes

A model for the hypothetical regulation of V H⁺-ATPase activity by extracellular glucose.

<p>Step A: the dissociation and reassociation of the catalytic complex V₁, according to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0049580#pone.0049580-Sumner1" target="_blank">[17]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0049580#pone.0049580-Kane1" target="_blank">[18]</a>; alternative step B: a putative biochemical modification and conformational change of V₁ subunits (this report).</p

The coupling capacity of the sub-populations of the V H⁺-ATPase molecules inactivated by growing concentrations of nitrate.

<p>For each nitrate concentration, the coupling ratio was calculated by dividing the difference of the initial velocities of H⁺ transport (V₀) for neighbouring concentrations by the similar difference in ATP hydrolysis. The enzymes exhibiting the highest coupling capacity of activated and non-activated states were inhibited by 5 mM nitrate in presence of 50 mM KCl. An example of the calculations: (V₀ at 0 mM nitrate – V₀ at 5 mM):(ATPase at 0 mM – ATPase at 5 mM).</p