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Cannabinoid receptors (CBRs) play a key role in various physiological processes, including neurogenesis, synaptic plasticity, immune modulation, cell apoptosis, metabolism regulation, cardiovascular and reproductive systems activity. Since activation of CBRs suppresses hyperexcitation and protects cells from death, their modulation may have therapeutic prospects in the treatment of such pathologies of the nervous system as mental disorders, epilepsy, Parkinson's and Huntington's disease, multiple sclerosis, spinal cord and brain injuries. This paper presents experimental data on the effects of the cannabinoid receptor agonist WIN 55,212-2 on the induced oscillations of intracellular Ca2+ concentration ([Ca2+]i) in two in vitro models of epileptiform activity. To study the neuroprotective properties of WIN 55,212-2, hyperexcitation was induced by the application of a GABA(A) receptor antagonist, bicuculline, or depolarizing doses of ammonium chloride. As experiments have shown, WIN 55,212-2 at a concentration of 100 nM and above significantly suppresses the [Ca2+]i oscillations frequency and reduces the basal [Ca2+]i level. At the same time, the amplitude of calcium oscillations also decreased in the presence of the agonist. WIN 55,212-2 at a concentration of 2 μM suppressed NH4Cl-induced [Ca2+]i oscillations in all neurons but caused a transient biphasic increase in the basal [Ca2+]i level in 20% of astrocytes. Thus, in this work, using various models of hyperexcitation of neuronal networks, we have demonstrated a potential antiepileptic effect of the cannabinoid receptor agonist WIN 55,212-2. [ABSTRACT FROM AUTHOR]

Key points: Alternaria aeroallergens induce the release of ATP from human bronchial epithelial (HBE) cells by activating a conductive pathway involving voltage‐dependent anion channel‐1 (VDAC‐1) and by exocytosis of ATP localized within membrane vesicles.Inhibition of VDAC‐1 blocked Alternaria‐evoked Ca2+ uptake across the plasma membrane of HBE cells and interleukin (IL)‐33 release into the extracellular media.Reducing cholesterol content with a cholesterol scavenger (β‐methylcyclodextrin) or statin compound (simvastatin) blocked ATP and IL‐33 release by lowering the expression of VDAC‐1 in the plasma membrane.Pretreatment with simvastatin for 24 h also inhibited the increase in tight junction macromolecule permeability that occurs following Alternaria exposure.These results establish a novel role for VDAC‐1 as a mechanism underlying ATP release induced by fungal allergens and suggests a possible therapeutic use for cholesterol lowering compounds in reducing Alternaria‐stimulated allergic inflammation. Human bronchial epithelial (HBE) cells exposed to allergens derived from the common saprophytic fungus, Alternaria alternata release ATP, which in turn stimulates P2X7 receptor‐mediated Ca2+ uptake across the plasma membrane. The subsequent increase in intracellular calcium concentration induces proteolytic processing and secretion of interleukin (IL)‐33, a critical cytokine involved in the initiation of allergic airway inflammation. A major objective of the present study was to identify the mechanism responsible for conductive ATP release. The results show that pretreatment of HBE cells with inhibitors of the voltage‐dependent anion channel‐1 (VDAC‐1) or treatment with a VDAC‐1 selective blocking antibody or silencing mRNA expression of the channel by RNA interference, inhibit Alternaria‐evoked ATP release. Moreover, inhibition of VDAC‐1 channel activity or reducing protein expression blocked the secretion of IL‐33. Similarly, reducing the cholesterol content of HBE cells with simvastatin or the cholesterol scavenger β‐methylcyclodextrin also blocked ATP release and IL‐33 secretion by decreasing the level of VDAC‐1 expression in the plasma membrane. In addition, simvastatin inhibited the increase in tight junction macromolecule permeability that was previously observed after Alternaria exposure. These results demonstrate a novel function for VDAC‐1 as the conductive mechanism responsible for Alternaria‐induced ATP release, an essential early step in the processing, mobilization and secretion of IL‐33 by the airway epithelium. Furthermore, the simvastatin‐evoked reduction of VDAC‐1 expression in the plasma membrane, suggests the possibility that cholesterol lowering compounds may be beneficial in alleviating allergic airway inflammation induced by fungal allergens. Key points: Alternaria aeroallergens induce the release of ATP from human bronchial epithelial (HBE) cells by activating a conductive pathway involving voltage‐dependent anion channel‐1 (VDAC‐1) and by exocytosis of ATP localized within membrane vesicles.Inhibition of VDAC‐1 blocked Alternaria‐evoked Ca2+ uptake across the plasma membrane of HBE cells and interleukin (IL)‐33 release into the extracellular media.Reducing cholesterol content with a cholesterol scavenger (β‐methylcyclodextrin) or statin compound (simvastatin) blocked ATP and IL‐33 release by lowering the expression of VDAC‐1 in the plasma membrane.Pretreatment with simvastatin for 24 h also inhibited the increase in tight junction macromolecule permeability that occurs following Alternaria exposure.These results establish a novel role for VDAC‐1 as a mechanism underlying ATP release induced by fungal allergens and suggests a possible therapeutic use for cholesterol lowering compounds in reducing Alternaria‐stimulated allergic inflammation. [ABSTRACT FROM AUTHOR]

Silver nanoparticles (Ag NPs) are the world's most important nanomaterial and nanotoxicant. The aim of this study was to determine the early stages of interactions between Ag NPs and plant cells, and to investigate their physiological roles. We have shown that the addition of Ag NPs to cultivation medium, at levels above 300 mg L−1, inhibited Arabidopsis thaliana root elongation and leaf expansion. This also resulted in decreased photosynthetic efficiency and the extreme accumulation of Ag in tissues. Acute application of Ag NPs induced a transient elevation of [Ca2+]cyt and the accumulation of reactive oxygen species ( ROS; partially generated by NADPH oxidase). Whole-cell patch-clamp measurements on root cell protoplasts demonstrated that Ag NPs slightly inhibited plasma membrane K+ efflux and Ca2+ influx currents, or caused membrane breakdown; however, in excised outside-out patches, Ag NPs activated Gd3+-sensitive Ca2+ influx channels with unitary conductance of approximately 56 pS. Bulk particles did not modify the plasma membrane currents. Tests with electron paramagnetic resonance spectroscopy showed that Ag NPs were not able to catalyse hydroxyl radical generation, but that they directly oxidized the major plant antioxidant, l-ascorbic acid. Overall, the data presented shed light on mechanisms of the impact of nanosilver on plant cells, and show that these include the induction of classical stress signalling reactions (mediated by [Ca2+]cyt and ROS) and a specific effect on the plasma membrane conductance and the reduced ascorbate. [ABSTRACT FROM AUTHOR]

Abstract: Elevated [Ca2+]i in human red blood cells (RBCs) activates IK1 K+ channels leading to cell dehydration. NS309, a powerful IK1 agonist, has been shown to activate IK1 channels even at sub-physiological [Ca2+]i levels. An intriguing feature of this response is its all-or-none nature, with responsive cells dehydrating fully and refractory cells retaining normal volume. We investigated the mechanism of this response suspecting cell-age involvement. We expected the younger cells, with the more vigorous plasma membrane Ca2+ pumps (PMCA), to be the refractory cells because of their lower [Ca2+]i. Osmotic fragility measurements and density separation through phthalate oil were used to monitor red cell dehydration. The fraction of glycosilated haemoglobin (Hb A1c) was used to estimate the mean age of density fractionated cells. The results showed that inhibition of the PMCA by vanadate abolished the all-or-none response, that the mean age of the responsive cells was young, contrary to expectations, and that pump inhibition subsequent to an all-or-none response caused the refractory cells to dehydrate fully, suggesting that the all-or-none response resulted from a reduced efficiency of NS309 to increase the Ca2+ sensitivity of IK1 channels in aged RBCs. [Copyright &y& Elsevier]

Abstract: Vertebrate left–right (LR) body axis is manifested as an asymmetrical alignment of the internal organs such as the heart and the gut. It has been proposed that the process of LR determination commonly involves a cilia-driven leftward flow in the mammalian node and its equivalents (Kupffer’s vesicle in zebrafish and the gastrocoel roof plate in Xenopus). Recently, it was reported that Ca2+ flux regulates Kupffer’s vesicle development and is required for LR determination. As a basis of Ca2+ flux in many cell types, inositol 1,4,5-trisphosphate (IP3) receptor-mediated calcium release from the endoplasmic reticulum (ER) plays important roles. However, its involvement in LR determination is poorly understood. We investigated the role of IP3 signaling in LR determination in Xenopus embryos. Microinjection of an IP3 receptor-function blocking antibody that can inhibit IP3 calcium channel activity randomized the LR axis in terms of left-sided Pitx2 expression and organ laterality. In addition, an IP3 sponge that could inhibit IP3 signaling by binding IP3 more strongly than the IP3 receptor impaired LR determination. Examination of the gastrocoel roof plate revealed that the number of cilia was significantly reduced by IP3 signal blocking. These results provide evidence that IP3 signaling is involved in LR asymmetry formation in vertebrates. [Copyright &y& Elsevier]

Newborn rat distal cells express an apical Ca2+ channel activated by dihydropyridine drugs. Similarly, in Madin-Darby canine kidney (MDCK) cells, nifedipine increased Ca2+i in a concentration-dependent manner (IC50 = 4 μM) in fura-2-loaded cells. Response to nifedipine was abolished by EGTA, suggesting that it depends on extracellular calcium. Ca2+ channel antagonist isradipine and agonist BayK8644 increased Ca2+i indicating that this effect is related to the dihydropyridine group. Diltiazem (20 μM) and gadolinium (200 μM) decreased the nifedipine effect (62 and 43%, respectively). Lanthanum (100 μM) did not change the response. Valinomycin clamping of the membrane potential did not modify nifedipine-induced increment, indicating that it was unrelated to potassium fluxes. We performed whole cell clamp experiments in MDCK cells maintained at -50 mV with perfusion solution containing 10 mM CaCl2. Nifedipine (20 μM) induced an increase in current (1.2 ± 0.3 nA), which was partially inhibited by Gd3+. No significant current was induced by nifedipine in the presence of 0.5 mM EGTA. To determine the effects of nifedipine on the membrane potential, we performed oxonol fluorescence experiments. The addition of nifedipine or Bay K8644 induced depolarization, highly dependent on external sodium. Nifedipine (20 μM) induced depolarization of 6.9 ± 0.8 mV (n = 21). EC50 to nifedipine was in the 10 μM range. We conclude that MDCK cells exhibit a dihydropyridine-activated cationic channel. Copyright © 2011 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

We previously found that polycystin-1 accelerated the decay of ligand-activated cytoplasmic calcium transients through enhanced reuptake of calcium into the endoplasmic reticulum (ER; Hooper KM, Boletta A, Germino GO, Hu Q, Ziegelstein RC, Sutters M. Am J Physiol Renal Physiol 289: F521-F530, 2005). Calcium flux across the ER membrane is determined by the balance of active uptake and passive leak. In the present study, we show that polycystin- I inhibited calcium leak across the ER membrane, an effect that would explain the capacity of this protein to accelerate clearance of calcium from the cytoplasm following a calcium release response. Calcium leak was detected by measurement of the accumulation of calcium in the cytoplasm following treatment with thapsigargin. Heterologous polycystin-1, stably expressed in Madin-Darby canine kidney cells, attenuated the thapsigargin-induced calcium peak with no effect on basal calcium stores, mitochondrial calcium uptake, or extrusion of calcium across the plasma membrane. The capacity of polycystin-1 to limit the rate of decay of ER luminal calcium following inhibition of the pump was shown indirectly using the calcium ionophore ionomycin, and directly by loading the ER with a low-affinity calcium indicator. We conclude that disruption of ER luminal calcium homeostasis may contribute to the cyst phenotype in autosomal dominant polycystic kidney disease. [ABSTRACT FROM AUTHOR]

Abstract: Much of what is known of the activities of polycystin-1 has been inferred from the effects of the isolated cytoplasmic COOH-terminal domain, but it is not clear whether the truncation acts like polycystin-1, as a dominant negative, or in unrelated pathways. To address this question, we have examined functional interactions between the intact and truncated forms of polycystin-1 in one cell system. In cells expressing only native polycystin-1, introduction of the truncation replicated the activity of the full-length protein. Conversely, when background levels of polycystin-1 were modestly elevated, the truncation acted as a dominant negative. Hence, the truncation acts in the polycystin pathway, but with effects that depend upon the background level of polycystin-1 expression. Our data raise the possibility that the cytoplasmic carboxyl terminus, either through cleavage products or intramolecular interactions, might feed back to modulate the activity of parent or intact polycystin-1. [Copyright &y& Elsevier]

Abstract: Olive oil is a major component of the Mediterranean diet, and its role in human health is being actively debated. This study aimed to clarify the mechanism of pancreatic adaptation to dietary fat. For this purpose, we examined whether dietary-induced modification of pancreatic membranes affects acinar cell function in response to the secretagogue acetylcholine (ACh). Weaning male Wistar rats were assigned to one of two experimental groups and fed for 8 weeks with a commercial chow (C) or a semisynthetic diet containing virgin olive oil as dietary fat (OO). The fatty acid composition of pancreatic plasma membranes was determined by gas–liquid chromatography. For assessment of secretory function, viable acini were incubated with ACh and amylase of supernatant was further assayed with a substrate reagent. Changes in cytosolic Ca2+ concentration in response to ACh were measured by fura-2 AM fluorimetry. Compared to C rats, pancreatic cell membranes of OO rats had a higher level of monounsaturated fatty acids and a lower level of both saturated and polyunsaturated fatty acids, thus, reflecting the type of dietary fat given. Net amylase secretion in response to ACh was greatly enhanced after OO feeding, although this was not paralleled by enhancement of ACh-evoked Ca2+ peak increases. In conclusion, chronic intake of diets that differ in the fat type influences not only the fatty acid composition of rat pancreatic membranes but also the responsiveness of acinar cells to ACh. This mechanism may be, at least in part, responsible for the adaptation of the exocrine pancreas to the type of fat available. [Copyright &y& Elsevier]

Recent studies indicate that chemoafferent nerve fiber excitation in the rat carotid body is mediated by acetylcholine and ATP, acting at nicotinic chofinergic receptors and P2X2 purinoceptors, respectively. We previously demonstrated that, after a 10- to 14-day exposure to chronic hypoxia (CH), the nicotinic cholinergic receptor blocker mecamylamine no longer inhibits rat carotid sinus nerve (CSN) activity evoked by an acute hypoxic challenge. The present experiments examined the effects of CH (9–16 days at 380 Torr) on the expression of P2X2 purinoceptors in carotid body and chemoafferent neurons, as well as the effectiveness of P2X2 receptor blocking drugs on CSN activity evoked by hypoxia. In the normal carotid body, immunocytochemical studies demonstrated a dense plexus of P2X2-positive nerve fibers penetrating lobules of type I cells. In addition, type I cells were lightly stained, indicating P2X2 receptor expression. After CH, the intensity of P2X2 receptor immunostaining was maintained in chemosensory type I cells and in the soma of chemoafferent neurons. P2 receptor expression on type I cells was confirmed by demonstrations of ATP-evoked increased intracellular Ca2+; this response was modulated by simultaneous exposure to hypoxia. In normal preparations, CSN activity evoked by hypoxia in vitro was 65% inhibited in the presence of specific P2X2 receptor antagonists. However, unlike the absence of mecamylamine action after CH, P2X2 antagonists remained effective against hypoxia-evoked activity after CH. Our findings indicate that ATP acting at P2X2 receptors contributes to adjusted chemoreceptor activity after CH, indicating a possible role for purinergic mechanisms in the adaptation of the carotid body in a chronic low-O2 environment. [ABSTRACT FROM AUTHOR]

Membrane potential in oxygen-sensitive type 1 cells in carotid body is controlled by diverse sets of voltage-dependent and -independent K+ channels. Coupling of Po2 to the open-closed state of channels may involve production of reactive oxygen species (ROS) by NADPH oxidase. One hypothesis suggests that ROS are produced in proportion to the prevailing Po2 and a subset of K+ channels closes as ROS levels decrease. We evaluated ROS levels in normal and p47phox gene-deleted [NADPH oxidase knockout (KO)] type I cells using the ROS-sensitive dye dihydroethidium (DHE). In normal cells, hypoxia elicited an increase in ROS, which was blocked by the specific NADPH oxidase inhibitor 4-(2-amino-ethyl)-benzenesulfonyl fluoride (AEBSF, 3 mM). KO type I cells did not respond to hypoxia, but the mitochondrial uncoupler azide (5 µM) elicited increased fluorescence in both normal and KO cells. Hypoxia had no effect on ROS production in sensory and sympathetic neurons. Methodological control experiments showed that stimulation of neutrophils with a cocktail containing the chemotactic peptide N-formyl- Met-Leu-Phe (1 µM), arachidonic acid (10 µM), and cytochalasin B (5 µg/mI) elicited a rapid increase in DHE fluorescence. This response was blocked by the NADPH oxidase inhibitor diphenyleneiodonium (10 µM). KO neutrophils did not respond; however, azide (5 µM) elicited a rapid increase in fluorescence. Physiological studies in type I cells demonstrated that hypoxia evoked an enhanced depression of K+ current and increased intracellular Ca2+ levels in KO vs. normal cells. Moreover, AEBSF potentiated hypoxia-induced increases in intracellular Ca2+ and enhanced the depression of K+ current in low O2. Our findings suggest that local compartmental increases in oxidase activity and ROS production inhibit the activity of type I cells by facilitating K+ channel activity in hypoxia. [ABSTRACT FROM AUTHOR]

Abstract: Galanin is a neurotransmitter peptide that suppresses insulin secretion. The present study aimed at investigating how a non-peptide galanin receptor agonist, galnon, affects insulin secretion from isolated pancreatic islets of healthy Wistar and diabetic Goto–Kakizaki (GK) rats. Galnon stimulated insulin release potently in isolated Wistar rat islets; 100μM of the compound increased the release 8.5 times (p<0.001) at 3.3mM and 3.7 times (p<0.001) at 16.7mM glucose. Also in islet perifusions, galnon augmented several-fold both acute and late phases of insulin response to glucose. Furthermore, galnon stimulated insulin release in GK rat islets. These effects were not inhibited by the presence of galanin or the galanin receptor antagonist M35. The stimulatory effects of galnon were partly inhibited by the PKA and PKC inhibitors, H-89 and calphostin C, respectively, at 16.7 but not 3.3mM glucose. In both Wistar and GK rat islets, insulin release was stimulated by depolarization of 30mM KCl, and 100μM galnon further enhanced insulin release 1.5–2 times (p<0.05). Cytosolic calcium levels, determined by fura-2, were increased in parallel with insulin release, and the L-type Ca2+-channel blocker nimodipine suppressed insulin response to glucose and galnon. In conclusion, galnon stimulates insulin release in islets of healthy rats and diabetic GK rats. The mechanism of this stimulatory effect does not involve galanin receptors. Galnon-induced insulin release is not glucose-dependent and appears to involve opening of L-type Ca2+-channels, but the main effect of galnon seems to be exerted at a step distal to these channels, i.e., at B-cell exocytosis. [Copyright &y& Elsevier]

Oscillation in calcium levels in the cytoplasm of individual cells has been observed experimentally to consist of a series of spikes and plateaux of differing amplitudes and inter-peak intervals. On the other hand, mathematical models based on known biochemical reaction kinetic behaviours predict, in the main, limit cycle behaviour. Chaotic solutions do not mimic the observed variability, and so another solution was sought by the introduction of filtered noise into some of the kinetic coefficients. Some of the variability can be predicted from this mechanism, but it is likely that other sources contribute to this as well. [ABSTRACT FROM AUTHOR]

Abstract: Early contractile dysfunction and the later death of cardiomyocytes are two major problems that can follow myocardial infarction or major cardiovascular surgery that demands ischemic arrest of the heart. Here, we found that 24 h of hypoxia and 1 h of reoxygenation induced the expression of the chaperone ORP150 in cultured rat cardiomyocytes. Inhibition of its induction using an adenovirus to express anti-sense ORP150 significantly enhanced the hypoxia-reoxygenation-induced cardiomyocyte death; cell death was reduced by overexpressing ORP150. Decreased levels of ORP150 expression also enhanced caspase-3 and -8 activation, cytochrome-c release, and DNA fragmentation, suggesting that this chaperone regulates apoptotic cell death. In contrast, increasing the expression of ORP150 in the cardiomyocytes had the opposite effect on the expression of these molecules. Moreover, apoptotic cell death initiated by myocardial ischemia–reperfusion (I/R) was significantly inhibited in vivo by transfecting an ORP150 expression plasmid into whole rat heart using the hemagglutinating virus of Japan (HVJ)-liposome method. Interestingly, ORP150 seemed to preserve calcium homeostasis in cardiomyocytes that underwent ischemia-reoxygenation in vitro. Calpain activity in the cardiomyocytes was enhanced by anti-sense ORP150 and suppressed by sense ORP150. Finally, we examined the functional recovery of rat hearts that overexpressed ORP150 or GFP protein and were subjected to I/R; we found that ORP150 preserved early contractile function after transient ischemia. Our results indicated cytoprotective roles for ORP150 in rat heart and suggested a therapeutic role for the protein both in preventing cardiomyocyte death and in preserving contractile function after ischemic damage. [Copyright &y& Elsevier]

The frog brain survives hypoxia with a slow loss of energy charge and ion homeostasis. Because hypoxic death in most neurons is associated with increases in intracellular calcium ([Ca2+]i), we examined the relationship between [Ca2+]i and survival of a mixed population of isolated cells from the forebrain of North American bullfrog Rana catesbeiana tadpoles. Forebrain cells from stage V-XV tadpoles were isolated by enzymatic digestion and loaded with one of three different calcium indicators (Fura-2, Fura 2-FF and BTC) to provide estimates of [Ca2+]i accurate at low and high [Ca2+]i. Propidium iodide (PI) fluorescence was used as an indicator of cell viability. Cells were exposed to anoxia (100% N2) and measurements of [Ca2+]i and cell survival made from 1 h to 18 h. Intracellular [Ca2+] increased significantly after 3–6 h anoxia (P<0.05), regardless of the type of Ca2+ indicator used; however, there were substantial differences in the measurements of [Ca2+]i with the different indicators, reflecting their varying affinities for Ca2+. Resting [Ca2+]i was approximately 50 nmol l-1 and increased to about 9–30 μmol l-1 after 4–6 h anoxia. The significant increase in [Ca2+]i during anoxia was not associated with significant increases in cell death, with 85–95% survival over this time period. Cells exposed to anoxia for 18 h, or those made anoxic for 4–6 and reoxygenated for 12 h to 16 h, had survival rates greater than 70%, but survival was significantly less than normoxic controls. These results indicate that large increases in [Ca2+]i are not necessarily associated with hypoxic cell death in vertebrate brain cells. [ABSTRACT FROM AUTHOR]

Abstract: Phenols, present in the Mediterranean diet, have antioxidant properties and are free radical scavengers; however, the molecular mechanisms of their beneficial effects are not yet fully understood. The level of cytosolic calcium ([Ca2+]i) is an important signal also in nonexcitable cells, including immune cells, and regulates fundamental processes. In this paper, we determine [Ca2+]i in human lymphomonocytes incubated with two olive oil phenols: 3,4-(dihydroxyphenyl)ethanol and p-(hydroxyphenyl)ethanol. Both tested phenols increase [Ca2+]i in a dose-dependent way. This effect is antagonized by nifedipine and is noticeable both in the presence and in the absence of calcium in the extracellular medium. [Copyright &y& Elsevier]

A mathematical modeling of tight junction (TJ) dynamics was elaborated in a previous study (Kassab, F., Marques, R.P., Lacaz-Vieira, F. 2002. Modeling tight junction dynamics and oscillations. J. Gen. Physiol. 120:237–247) to better understand the dynamics of TJ opening and closing, as well as oscillations of TJ permeability that are observed in response to changes of extracellular Ca2+ levels. In this model, TJs were assumed to be specifically controlled by the Ca2+ concentration levels at the extracellular Ca2+ binding sites of zonula adhaerens. Despite the fact that the model predicts all aspects of TJ dynamics, we cannot rule out the likelihood that changes of intracellular Ca2+ concentration (Ca2+ cell), which might result from changes \ of extracellular Ca2+ concentration (Ca2+ extl), contribute to the observed results. In order to address this aspect of TJ regulation, fast Ca2+-switch experiments were performed in which changes of Ca2+ cell were induced using the Ca2+ ionophore A23187 or thapsigargin, a specific inhibitor of the sarco-endoplasmic reticulum Ca2+-ATPase. The results indicate that the ionophore or thapsigargin per se do not affect basal tissue electrical conductance (G), showing that the sealing of TJs is not affected by a rise in Ca2+ cell. When TJs were kept in a dynamic state, as partially open structures or in oscillation, conditions in which the junctions are very sensitive to disturbances that affect their regulation, a rise of Ca2+ cell never led to a decline of G, indicating that a rise of Ca2+ cell does not trigger per se TJ closure. On the contrary, always the first response to a rise of Ca2+ cell is an increase of G that, in most cases, is a transient response. Despite these observations we cannot assure that a rise of Ca2+ cell is without effect on the TJs, since an increase of Ca2+ cell not only causes a transient increase of G but, in addition, during oscillations a rise of Ca2+ cell induced by the Ca2+ ionophore transiently halted the oscillatory pattern of TJs. The main conclusion of this study is that TJ closure that is observed when basolateral Ca2+ concentration (Ca2+ bl) is increased after TJs were opened by Ca2+ bl removal cannot be ascribed to a rise of Ca2+ cell and might be a consequence of Ca2+ binding to extracellular Ca2+ sites. [ABSTRACT FROM AUTHOR]

SUMMARY 1. Increased sodium intake causes decreased formation of angiotensin (Ang) II and increased AngII causes increased Na+ retention. 2. Increased sodium intake and increased AngII causes cardiac hypertrophy, but decreased sodium intake regresses cardiac hypertrophy despite high AngII levels. Likewise, decreased Na+ and blockers of the renin–angiotensin system (RAS) in neonatal rats have similar effects on subsequent blood pressure development. 3. Cardiac hypertrophy due to renal hypertension does not regress when the RAS is blocked and rats are on a high salt intake. Likewise, sodium restriction alone does not cause regression; combination of reduced NaCl intake and RAS blockade is required. 4. High doses of perindopril and losartan in combination cause a syndrome in rats on 0.2% NaCl that leads to profound hypotension, polyuria, renal impairment and involution of the heart and death. This is reversed or prevented by a high (4%) NaCl intake, which also prevents the plasma angiotensinogen depletion that occurs with combined blockade on 0.2% NaCl intake. 5. Intake of NaCl and AngII interact at many levels. It is postulated that there is an important interaction at the cellular level that can explain the above events. [ABSTRACT FROM AUTHOR]

Increased lysophosphatidylcholine (LPC) production by the ischemic heart is associated with tissue damage. In vitro, LPC produces an increase in cytosolic [Ca2+], usually followed by cell contracture and lysis. Since ethanol reportedly protect cells during ischemia-reperfusion, we wished to determine whether ethanol could protect heart cells against LPC-induced Ca2+ overload. Newborn rat heart cells in culture were loaded with Fura-2 and [Ca2+]i recorded in individual cells. The presence of 22 or 44 mM ethanol increased the time required for 10 µM L-palmitoyl-LPC to produce maximal Ca2+ accumulation from 8.4±0.4 min (n=47) to 21.1±2.1 min (n=32; P<0.01) and 23.8±1.8 min (n=10; P<0.01) respectively. The onset of the [Ca2+]i increase could be reversed partially by the addition of ethanol (44 or 88 mM). After the addition of 22 mM ethanol, the cells retained the Fura-2 three times longer than under control conditions. Ethanol (88 mM) decreased the critical micelle concentration of LPC, thus decreasing the LPC monomer concentration in this solution. La3+ also protected the cells against LPC but no further protection was afforded by the addition of ethanol. Our results suggest that ethanol concentrations commonly found in the blood of social drinkers protect heart cells against the deleterious effect of LPC. [ABSTRACT FROM AUTHOR]

Intracellular Na+ directly modulates Na+,K+-ATPase gene expression in normal rat kidney epithelial cells. Background. In a wide variety of cell systems, increases in cell Na+ ([Na+]i) lead to an induction of N+,K+-ATPase mRNA expression. On the other hand, the increase in [Na+]i can also induce a rise in cell Ca2+ ([Ca2+]i) through a secondary inhibition of Na+/Ca2+ exchange and a decrease in cell pH (pHi) through a secondary inhibition of Na+/H+ exchange. It is not known whether [Na+]i, [Ca2+]i, and/or pHi directly modulate N+,K+-ATPase mRNA expression. Methods. We used normal rat kidney epithelial cells (NRK) to examine the effects of ouabain on N+,K+-ATPase α1- and β1-mRNA accumulation by Northern blot analysis and the relationship between the mRNA accumulation and [Na+]i, [Ca2+]i, or pHi. [Na+]i, [Ca2+]i, and pHi were measured using a Na+-sensitive fluorescent dye (SBFI), a Ca2+-sensitive fluorescent dye (Fura-2), and a pH-sensitive fluorescent dye (BCECF), respectively. Results. Ouabain (1 mmol/L) significantly increased [Na+]i. Upon addition of ouabain, α1-mRNA levels increased to 2.3 times the control level at three hours, with maximum 3.3-fold elevations at 12 hours. β1-mRNA levels also increased to 2.4 times the control level at 3 hours, with a maximum 3.3-fold increase at 12 hours. The ouabain-mediated α1- and β1-mRNA induction was inhibited by both the RNA transcription inhibitor (actinomycin D) and the protein synthesis inhibitor (cycloheximide). Ouabain at three hours caused an... [ABSTRACT FROM AUTHOR]

The intracellular calcium pool of adipose tissue was estimated by studying the kinetics ofCa efflux from the tissue of 9-week old spontaneously hypertensive (SHR) and normotensive Kyoto-Wistar (NKWR) rats subjected to peripheral immunosympathectomy (nerve growth factor antiserum treatment of newborn rats) and in intact (shamtreated) animals of both strains. The development of hypertension in SHR was prevented by immunosympathectomy, however, both groups, SHR-intact and immunosympathectomized rats showed an increase in the intracellular calcium content of adipocytes. These results indicate that the alteration of the membrane control over intracellular calcium distribution in adipose tissue of SHR cannot be attributed to an enhanced activity of the sympathetic nervous system. [ABSTRACT FROM AUTHOR]