The Influence of Lipopolysaccharides and Glucans from Two Rhizobium leguminosarum bv. viciae Strains on the Formation and Efficiency of Their Symbioses with Pea Plants

The influence of lipopolysaccharides (LPS), glucans, and their unseparated complexes on nodulation activity of rhizobia and efficiency of their symbioses with pea plants was studied in vegetation tests. Two Rhizobium leguminosarum bv. viciae strains which differed in their symbiotic properties were used: strain 31 (fix⁺, efficient, moderately virulent, and moderately competitive) and strain 248b (fix^–, inefficient, highly virulent, and highly competitive). Preparations of LPS–glucan complex and the respective LPS from the highly virulent strain 248b increased the nodulation activity of both strains by 10–26%. Analogous preparations from a less virulent strain 31 did not have this ability. Unseparated LPS–glucan complexes from these strains increased the productivity of plants infected with the efficient strain by 18–23% but did not change it in plants inoculated with the other, inefficient strain. No significant influence of LPS preparations on the symbiosis productivity was observed. Glucans from both strains enhanced the nodulation ability of the highly virulent strain by 36–56%. In addition, treatment of pea plants with glucan from strain 248b increased nitrogen fixation by root nodules by 27% in plants inoculated with strain 31. In general, the formation and efficiency of the symbiosis of R. leguminosarum bv. viciae with pea plants was more influenced by preparations from strain 248b, highly virulent but deficient in nitrogen fixation, than by preparations from the nitrogen fixation–proficient but less virulent strain 31.     

Binding and biological properties of lipopolysaccharide Proteus vulgaris O25 (48/57)–chitosan complexes

In this study the negatively charged Proteus vulgaris O25 LPS was chosen for studying interaction with polycationic chitosan. The complex formation of LPS with chitosan was demonstrated using gradient centrifugation and laser interferometry method. The presented results have shown that laser interferometry method is sensitive enough for LPS–chitosan interaction studies. The changing in the ultra structure of LPS during binding with chitosan was observed by electronic microscope. The interaction of P. vulgaris O25 LPS with chitosan was shown to modulate significantly the biological activities of LPS. The toxicity of P. vulgaris O25 LPS decreased 10-fold after forming complexes with chitosan at injection to mice in the similar concentration of endotoxin. The complex LPS–chitosan was less effective than LPS alone in Limulus amabocyte lysate assay. Induction of TNF biosynthesis by LPS–chitosan complex was found to be 65% lower than that by parent LPS at concentration of 100 ng/ml.     

Preparation and investigation of 99m technetium-labeled low-density lipoproteins in rabbits with experimentally induced hypercholesterolemia

Low-density lipoproteins (LDL) were radiolabeled in atherosclerosis studies. The aim was to investigate the biodistribution and uptake of ^99mTc-labeled LDL by atherosclerotic plaques in experimentally induced hyperlipidemia. Rabbits were fed a diet containing 2% cholesterol for 60 days to develop hyperlipidemia and atheromatous aortic plaques. A combination of preparative and analytical ultracentrifugation was used to investigate human LDL aliquots, to prepare radioactive-labeled lipoproteins and in rabbits with induced hyperlipidemia. Preparative density gradient centrifugation was applied for the simultaneous isolation of the major lipoprotein density classes, which form discrete bands of lipoproteins in the preparative tubes. The cholesterol and protein levels in the lipoprotein fractions were determined. LDL was subsequently dialysed against physiological solution and sterilized and apolipoprotein fragments and aggregates were eliminated by passage through a 0.22-micron filter. LDL was radiolabeled with ^99mTc by using sodium dithionite as a reducing agent. Radiochemical purity and in vitro stability were controlled by paper chromatography in acetone. The labelling efficiency was 85–90% for human LDL. Two months after the start of cholesterol feeding, the total cholesterol in the blood serum had increased approximately 33-fold in comparison with the basal cholesterol content of hypercholesterolemic rabbits. Investigation of LDL was performed by Schlieren analysis after adjustment of the density of serum and underlayering by salt solution in a spinning ultracentrifugation capillary band-forming cell. Quantitative results were obtained by measuring the Schlieren areas between the sample curves and the reference baseline curve by means of computerized numerical and graphic techniques. In this manner we measured the concentrations of human LDL and analyzed rabbit LDL levels in induced hyperlipidemia. Gamma scintillation camera scanning of the rabbits was performed. Overnight fasted rabbits were injected in the marginal ear vein with ^99mTc-labeled human LDL (4–10 mCi, 0.5–1.5 mg protein). The initial scintigram showing a typical blood-pool scan, gradually changing with time to an image of specific organ uptake of radioactivity by the liver, kidneys and brain and in the bladder. Gamma camera in vivo scintigraphy on rabbits revealed visible signals corresponding to atherosclerotic plaques in the aorta and carotid arteries. Our results show that ^99mTc-LDL can be used to assess the organ distribution pattern of LDL in the rabbit, and to detect and localize areas of arterial atherosclerotic lesions.     

Lipoproteins from normolipidemic and dyslipidemic subjects modify the thromboxane A2 generation by platelets in clotting human blood

The study was performed to investigate the influence of lipoproteins (LP) on the thromboxane (TX) A₂ formation capacity of platelets in clotting whole blood in vitro. The different lipoprotein fractions VLDL, LDL, HDL₂ and HDL₃ were isolated from blood of normo- or dyslipidemic volunteers by ultracentrifugation. These lipoproteins were incubated in blood with different levels of serum total cholesterol (TC) taken from normolipidemics (TC < 200 mg/dl), moderate hypercholesterolemics (TC: 200–250 mg/dl) or subjects with high cholesterol level (TC > 250 mg/dl), respectively. The amount of serum TXA₂ formed within 60 min at 37°C was measured by enzyme immunoassay. The results obtained show that the efficacy of separate LP fractions to influence the TXA₂ production depends not only on the type of LP fraction but also on the source of plasma used for isolation of LP and on the cholesterol level in the blood for incubation: LDL taken from normolipidemics or moderate hyperlipidemics inhibited the TXA₂ formation in blood from normolipidemics (P < 0.02, respectively), but enhanced it in blood from persons with moderate hypercholesterolemia (P < 0.05). LDL from hyperlipidemics enhanced TXA₂ production in blood from hyperlipidemics (P < 0.05). The HDL₂ fractions inhibited the TXA₂ formation in blood from normo- and hypercholesterolemics (P < 0.02, resp.), but there was no effect of HDL₂ in clotting blood from persons with moderate hypercholesterolemia. All HDL₃ fractions tested inhibited the TXA₂ formation in all types of blood used for clotting (P < 0.02, resp.), probably due to their great cholesterol accepting capacity.     

Chemiluminescence response of the human polymorphonuclear neutrophil to lipopolysaccharides

Polymorphonuclear neutrophils (PMN) respond to a variety of stimuli with a sequence of reactions that lead to the production of “active oxygen” species, including H₂O₂, free radicals, such as superoxide (O₂ ⁻·) and hydroxyl (HO·), and singlet molecular oxygen (¹O₂). Some of these can oxidize (5-amino-2,3-dihydrophthalazine 1,4-dione) (luminol) to the ground state aminophthalate ion; this reaction sequence is accompanied by the generation of a photon and forms the basis for the chemiluminescence (CL) response. In this work we used a dedicated photon counting instrument to record CL from PMN incubated with bacterial lipopolysaccharide (LPS). We have studied the CL response to the LPS fromEscherichia coli strains 026:B6 and 055:B5, as well asSalmonella minnesota RE 595 and have determined that CL requires heat-labile serum factors, these most likely being intact components of the complement system.     

Distribution of lecithin-cholesterol acyltransferase (LCAT) in human plasma lipoprotein fractions. Evidence for the association of active LCAT with low density lipoproteins

The distribution of lecithin-cholesterol acyltransferase (LCAT) in human plasma was assessed by measuring both LCAT mass and activity in plasma fractions separated by sequential flotation ultracentrifugation, single-spin gradient ultracentrifugation, dextran sulfate-Mg²⁺ precipitation or agarose gel filtration. Although most of the LCAT was found to be associated with the high density lipoprotein fraction, a small amount of active LCAT (approximately 1% of the plasma LCAT mass and activity) was consistently associated with the low density lipoprotein fraction. LCAT was not found in the very low density lipoprotein fraction. The LDL-associated LCAT may play an important role in the acylation of lysolecithin by lysolecithin acyltransferase activity of LCAT.     

Isolation of lipopolysaccharides and the effect of Polymyxin B on the outer membrane of Corynebacterium autotrophicum

Lipopolysaccharides (LPS) from Corynebacterium autotrophicum were isolated and analyzed. Autotrophically grown cells contained 2–5 mg of partly purified LPS per g dry weight of lyophilized cells. Serological cross reaction with Lipid A antigen of Salmonella minnesota confirmed the presence of LPS in C. autotrophicum. Electron microscopy of negatively stained Polymyxin B-treated cells showed formation of blebs on the Outer Membrane indicating an interaction of Polymyxin B specifically with LPS. Up to now, no Gram-positive organisms are known which contain any LPS. Thus, C. autotrophicum, though giving opposite results when the Gram-staining reaction was applied by several authors, has to be classified into the group of Gram-negative bacteria.Non-Common Abbreviations LPS lipopolysaccharide - KDO 2-keto-3-deoxyoctonate     

Serum lipoproteins and albumin in the lecithin: Cholesterol acyltransferase reaction

The unique features of pig ovarian follicular fluids, i.e., presence of high density lipoprotein (HDL) only and lecithin: cholesterol acyltransferase (EC 2.3.1.43; LCAT) activity, provides a good model to study the effect of serum lipoproteins and serum albumin on the LCAT reaction. $\text{In}\text{vitro}$ cholesterol esterification is enhanced when very low density lipoprotein (VLDL) and low density lipoprotein (LDL) fractions are added, but is inhibited when one or the other of these lipoproteins is absent. High concentrations of HDL₂ result in decreased activation which can be compensated for by the addition of the VLDL-LDL mixture. These findings suggest that the rate of cholesterol esterification in ovarian follicular fluid may be enhanced by providing the exogenous VLDL and LDL as the recipients of HDL-cholesteryl ester. The inhibition of LCAT activity caused by free fatty acid and lysophosphatidylcholine can be partially reversed by the addition of serum albumin, suggesting that serum albumin may regulate the LCAT reaction.     

Serum lipoproteins in elderly patients with cerebrovascular pathology

Contents of 7 serum lipoprotein (LP) fractions both native and after their incubation under 37 degrees C during 24 h in the patients with atherosclerosis and hypertension discirculatory encephalopathy were investigated by a gradient gel-electrophoresis method. Two groups of patients in the age of 30-59 in the first group and 61-73 in the second one were examined. It was found that for the age interval examined in native serum of the patients with atherosclerosis and hypertension similar changes of LP contents were observed. These changes were expressed a increasing apoA- and decreasing apoB-containing LP fractions as compared with the control. After the temperature incubation of healthy persons serum the amount of LDL increases and amount of HDL3 declines. The latter is a result of LP lypolysis and etherification normal processes. On the base of the data obtained we may suppose that in the patients with hypertension these processes also have some aged peculiarities, although less intensive as compared with the healthy persons. In contrast to the latter after the temperature incubation of serum (in vitro) in the patients with atherosclerosis there were no any statistically significant changes of any LP fractions contents that may evidence about depression of LP transformation in the isolated serum in such patients.     

Participation of lipopolysaccharide in hyperplasic adipose expansion: Involvement of NADPH oxidase/ROS/p42/p44 MAPK‐dependent Cyclooxygenase‐2

Obesity is a world‐wide problem, especially the child obesity, with the complication of various metabolic diseases. Child obesity can be developed as early as the age between 2 and 6. The expansion of fat mass in child age includes both hyperplasia and hypertrophy of adipose tissue, suggesting the importance of proliferation and adipogenesis of preadipocytes. The changed composition of gut microbiota is associated with obesity, revealing the roles of lipopolysaccharide (LPS) on manipulating adipose tissue development. Studies suggest that LPS enters the circulation and acts as a pro‐inflammatory regulator to facilitate pathologies. Nevertheless, the underlying mechanisms behind LPS‐modulated obesity are yet clearly elucidated. This study showed that LPS enhanced the expression of cyclooxygenase‐2 (COX‐2), an inflammatory regulator of obesity, in preadipocytes. Pretreating preadipocytes with the scavenger of reactive oxygen species (ROS) or the inhibitors of NADPH oxidase or p42/p44 MAPK markedly decreased LPS‐stimulated gene expression of COX‐2 together with the phosphorylation of p47^phox and p42/p44 MAPK, separately. LPS activated p42/p44 MAPK via NADPH oxidase‐dependent ROS accumulation in preadipocytes. Reduction of intracellular ROS or attenuation of p42/p44 MAPK activation both reduced LPS‐mediated COX‐2 expression and preadipocyte proliferation. Moreover, LPS‐induced preadipocyte proliferation and adipogenesis were abolished by the inhibition of COX‐2 or PEG₂ receptors. Taken together, our results suggested that LPS enhanced the proliferation and adipogenesis of preadipocytes via NADPH oxidase/ROS/p42/p44 MAPK‐dependent COX‐2 expression.     

Chronic oral infection with Porphyromonas gingivalis accelerates atheroma formation by shifting the lipid profile

Background

Recent studies have suggested that periodontal disease increases the risk of atherothrombotic disease. Atherosclerosis has been characterized as a chronic inflammatory response to cholesterol deposition in the arteries. Although several studies have suggested that certain periodontopathic bacteria accelerate atherogenesis in apolipoprotein E-deficient mice, the mechanistic link between cholesterol accumulation and periodontal infection-induced inflammation is largely unknown.

Methodology/Principal Findings

We orally infected C57BL/6 and C57BL/6.KOR-Apoe^shl (B6.Apoeshl) mice with Porphyromonas gingivalis, which is a representative periodontopathic bacterium, and evaluated atherogenesis, gene expression in the aorta and liver and systemic inflammatory and lipid profiles in the blood. Furthermore, the effect of lipopolysaccharide (LPS) from P. gingivalis on cholesterol transport and the related gene expression was examined in peritoneal macrophages. Alveolar bone resorption and elevation of systemic inflammatory responses were induced in both strains. Despite early changes in the expression of key genes involved in cholesterol turnover, such as liver X receptor and ATP-binding cassette A1, serum lipid profiles did not change with short-term infection. Long-term infection was associated with a reduction in serum high-density lipoprotein (HDL) cholesterol but not with the development of atherosclerotic lesions in wild-type mice. In B6.Apoeshl mice, long-term infection resulted in the elevation of very low-density lipoprotein (VLDL), LDL and total cholesterols in addition to the reduction of HDL cholesterol. This shift in the lipid profile was concomitant with a significant increase in atherosclerotic lesions. Stimulation with P. gingivalis LPS induced the change of cholesterol transport via targeting the expression of LDL receptor-related genes and resulted in the disturbance of regulatory mechanisms of the cholesterol level in macrophages.

Conclusions/Significance

Periodontal infection itself does not cause atherosclerosis, but it accelerates it by inducing systemic inflammation and deteriorating lipid metabolism, particularly when underlying hyperlidemia or susceptibility to hyperlipidemia exists, and it may contribute to the development of coronary heart disease.     

Induction of antitumor activities in spleen cells from mice and rats activated with lipopolysaccharide immobilized on beads

Summary Lipopolysaccharides (LPS) were coupled to polystyrene beads in order to apply the LPS without toxicity. The antitumor activity of the LPS-immobilizing beads was studied in experiments in vitro and in vivo. In vitro studies showed that spleen cells from C3H/HeN mice stimulated by beads immobilizing LPS fromEscherichia coli produced cytolytic activity as strong as that of lymphokine-activated killer (LAK) cells. Spleen cells from Sprague-Dawley rats stimulated by beads immobilizing LPS fromSalmonella minnesota produced cytolytic activity stronger than that of LAK cells. However, spleen cells stimulated by beads immobilizing each component of the LPS separately could not induce cytolysis. Contact stimulation, even for a brief period, sufficed for cytolytic activity, and was enhanced by culture for 48–72 h. Through in vivo studies, the suppression of tumor growth and a prolongation of the survival time were observed in tumorbearing mice injected with spleen cells activated by beads immobilizing LPS fromE.coli, and in mice injected with LAK cells. The effect of the activated spleen cells was stronger than that of the LAK cells. In rats bearing metastatic tumors, spleen cells activated by beads immobilizing LPS fromS.minnesota suppressed lung metastases more strongly than did LAK cells. These findings indicate that LPS immobilized by beads induced killer cells more strongly than interleukin-2. Ex vivo immunomodulation with LPS-immobilizing beads can be applied usefully as an anticancer treatment.     

Effects of Apigenin on the Expression of LOX-1, Bcl-2, and Bax in Hyperlipidemia Rats

We aimed to investigate the impact of apigenin on LOX-1, Bcl-2, and Bax expression in hyperlipidemia rats and explore the possible molecular pathological mechanism of apigenin in improving hyperlipidemia and preventing atherosclerosis. In hyperlipidemia models, the levels of total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-c) and the LOX-1 protein expression were apparently increased (P<0.01), while the high-density lipoprotein cholesterol (HDL-c) levels and the ratio of Bcl-2/Bax were reduced significantly (P<0.01) in comparison with the standard control group. After the treatment of apigenin, the levels of TC, TG, LDL-c, and the LOX-1 protein expression were noticeably decreased (P<0.01), while the levels of HDL-c and the Bcl-2/Bax ratio were increased (P<0.01). The intima was thickened and had protrusions in the hyperlipidemia model group compared to the normal control group. In comparison with the atherosclerosis model group, the degree of aortic lesions in the low-dose, middle-dose, high-dose groups was alleviated. Apigenin can reduce the level of blood lipid, improve hyperlipidemia, and prevent atherosclerosis in hyperlipidemia rats. The molecular mechanism may be related to inhibiting LOX-1 gene expression and increasing the Bcl-2/Bax ratio.     

Effect of Sargassum polycystum (Phaeophyceae)-sulphated polysaccharide extract against acetaminophen-induced hyperlipidemia during toxic hepatitis in experimental rats

The effect of Sargassum polycystum crude extract on lipid metabolism was examined against acetaminophen-induced (800 mg/kg body wt., intraperitoneally) hyperlipidemia during toxic hepatitis in experimental rats. The animals intoxicated with acetaminophen showed significant elevation in the levels of cholesterol, triglycerides and free fatty acid in both serum and liver tissue. The levels of tissue total lipids and serum LDL-cholesterol were also elevated with depleted levels of serum HDL-cholesterol and tissue phospholipid. The acetaminophen-induced animals showed significant alterations in the activities of lipid metabolizing enzymes serum lecithin cholesterol acyl transferase (LCAT) and hepatic triglyceride lipase (HTGL). The levels of liver tissue fatty acids (saturated, mono and polyunsaturated) such as palmitic acid, stearic acid, oleic acid, linoleic acid, arachidonic acid and linolenic acid monitored by gas chromatography were considerably altered in acetaminophen intoxicated animals when compared with control animals. The prior oral administration of Sargassum polycystum (200 mg/kg body wt./day for a period of 15 days) crude extract showed considerable prevention in the severe disturbances of lipid profile and metabolizing enzymes triggered by acetaminophen during hepatic injury. Liver histology also showed convincing supportive evidence regarding their protective nature against fatty changes induced during acetaminophen intoxication. Thus the present study indicates that the protective nature of Sargassum polycystum extract may be due to the presence of active compounds possessing antilipemic property against acetaminophen challenge. (Mol Cell Biochem 276: 89–96, 2005)     

Carbohydrate restriction and dietary cholesterol distinctly affect plasma lipids and lipoprotein subfractions in adult guinea pigs

To evaluate the effects of carbohydrate restriction (CR) and dietary cholesterol on lipoprotein metabolism, adult male guinea pigs (10 guinea pigs/diet) were fed either low (0.04 g/100 g) or high (0.25 g/100 g) amounts of dietary cholesterol, in combination with either low (10% total energy) or high (54.2% total energy) dietary carbohydrate (control groups) for a total of four groups: high carbohydrate–low cholesterol (control-L), high carbohydrate–high cholesterol (control-H), low carbohydrate–low cholesterol (CR-L) and low carbohydrate–high cholesterol (CR-H). Plasma triglyceride concentrations were lower (P<.01%), while high-density lipoprotein cholesterol concentrations were higher (P<.05) in the CR groups compared to the control groups. In contrast, high dietary cholesterol (CR-H and control-H) resulted in higher concentrations of total and low-density lipoprotein (LDL) cholesterol compared to those guinea pigs fed the low-cholesterol diets (P<.01). Dietary cholesterol significantly increased the total number of LDL particles (P<.001) and the number of small LDL (P<.001), as determined by nuclear magnetic resonance. In contrast, carbohydrate restriction (CR-L and CR-H) resulted in lower concentrations of medium very-low-density lipoprotein and small LDL particles compared to the high-carbohydrate groups. Plasma lecithin:cholesterol acyltransferase (LCAT) activity was decreased and cholesterol ester transfer protein activity was increased by dietary cholesterol, whereas carbohydrate restriction increased LCAT activity (P<.05). These findings are similar to those observed in humans, thus validating the use of adult guinea pigs to study lipid responses to carbohydrate restriction. The results also indicate that the atherogenicity of lipoproteins induced by high dietary cholesterol is attenuated by carbohydrate restriction in guinea pigs.     

Interaction of Porin from Yersinia pseudotuberculosis with Different Structural Forms of Endogenous Lipopolysaccharide

The interaction of Yersinia pseudotuberculosis porin solubilized in deoxycholate with the S- and R-forms of endogenous lipopolysaccharide (LPS) was studied by the quenching of intrinsic protein fluorescence. The samples of S-LPS differed both in the length of O-specific polysaccharide (n = 1 and 4) and in the acylation degree of the 3-hydroxytetradecanoic acid residues of the lipid A moiety (12-66%). R-LPS (12%) binding to porin was found to occur with positive cooperativity on two integrated structural regions of the R-LPS macromolecule, namely, core oligosaccharide and lipid A. The mode of porin interaction with low-acylated S-LPSs (15 or 20%) coincided with a model involving three types of binding sites. The shape of Scatchard curves of binding indicates that a complex formation between porin and low-acylated S-LPS is cooperative at low and moderate ligand concentration, whereas at near-saturating LPS concentrations porin binds to LPS independently on two types of binding sites. The O-specific polysaccharide chain in the S-LPS macromolecule increases the affinity of its interaction with porin in comparison with R-LPS–porin binding. A significant increase (to 66%) in the degree of S-LPS acylation substantially changed its porin-binding character: the process becomes anti-cooperative with lowered affinity. Thus, the features of LPS–porin interaction significantly depend on the conformational changes in the LPS molecule due to expanding of its hydrophobic region.     

Effects of serum,its protein and lipid extracts,and commercial serum proteins and lipid on the isolated frog heart

Summary This study investigates the inotropic effects of serum, its protein and lipid extracts, and commercial serum proteins and lipid on the isolated, spontaneously-beating heart and superfused, hypodynamic ventricle of the frog. Serum taken from either man, horse, calf, frog, or rabbit evoked marked positive inotropic responses which were unaffected by cholinergic, serotonergic, and adrenergic receptor antagonists. Dialysed serum (dialisand) and void volume fractions from Sephadex G200–120 columns corresponding to large molecular weight constituents evoked marked positive inotropic responses. When serum was separated into fractions containing either proteins or lipids/lipoproteins by high-density ultracentrifugation or activated charcoal, both extracts evoked marked positive inotropic responses. Commerical serum globulins and serum containing a high proportion of immunoglobulins elicited large increases in contractile force, whereas serum albumin evoked a negative inotropic effect. Serum which was either boiled and/or treated with chymotrypsin to denature proteins also caused a marked increase in isometric twitch tension in the frog heart. Similar inotropic response was obtained with fractions of boiled serum eluted on columns of Sephadex G200–120. These fractions corresponded to molecular weight in the region of 60–70 kDa. However, the inotropic effect of boiled serum was abolished following pretreatment with lipase. Superfusion of frog hearts with commercial cardiolipin resulted in marked dose-dependent increases in contractile force. The results demonstrate the presence of at least two large molecular weight cardioactive principles in serum. These substances are comparable in size to constituents of serum proteins (e.g., globulins and immuno-globulins) and serum lipids/lipoproteins (e.g., cardiolipin) and may serve as physiological regulators of cardiac function.Abbreviations Ca ²⁺ Calcium - Da dalton - IgG immunoglobulins - Na ⁺ Sodium - K ⁺ potassium     

Branchial Cl transport, anion-stimulated ATPase and acid-base balance inAnguilla anguilla adapted to freshwater: Effects of hyperoxia

Summary Freshwater eel gills are notorious for their limited ability to pump chloride. As a result there is a considerable discrepancy between the Na⁺ and Cl^– plasma levels, and plasma HCO₃ ^– and blood pH are relatively high in this species.When eels are kept in tanks aerated with pure oxygen, significant alterations in blood acid-base balance, an increase in plasma pCO₂ and a decrease in blood pH, are observed. In fish studied after 3 weeks hyperoxia, the decrease in blood pH is compensated by an increase in plasma HCO₃ ^–. Such fish exhibit a Cl^– influx 5 times higher than that observed in normoxic fish. This Cl^– influx is readily inhibited by addition of SCN^– to the external medium.An anion-stimulated ATPase activated by HCO₃ ^– and by Cl^– and inhibited by SCN^– was recently described in membrane fractions of the gills ofCarassius auratus, a fish noted for its high Cl^– pumping rate. This enzyme is also found in the gills of the eel. While the maximal rates of enzyme activation by HCO₃ ^– and by Cl^– are similar inCarassius andAnguilla, the affinity of the enzyme for Cl^– is 25 times higher inCarassius. In the microsomal fraction of the hyperoxic eel gills, the maximal anionstimulated ATPase activity remains unchanged but HCO₃ ^– affinity decreases by 50%, while Cl^– affinity increases 5 times. Thus some characteristics of this ATPase seem to be closely related to the Cl^– pump activity exhibited by the gill in fresh water.     

Unaccustomed high mileage compared to intensity training-related neuromuscular excitability in distance runners

The influence of a 4-week unaccustomed average 103% mileage increase (ITV, increase in training volume;n = 8; average baseline mileage 85.9 km · week^–1, final mileage 174.6km · week^–1) on performance and neuromuscular excitability (NME) was tested in experienced distance runners and controlled 1 year later by a 4-week unaccustomed average 152% increase in tempo-pace and interval-runs (ITI, increase in training intensity;n = 9; baseline 9 km · week^–1, final 22.7 km · week^–1) with an average total mileage of 61.7 km · week^–1 (week 1) to 84.7 km · week^–1 (week 4). Seven athletes participated in ITV as well as in ITI. During incremental treadmill test performance at a lactate concentration of 2 mmol · l^–1 (2 LP) increased, and at 4 mmol · l^–1 (4 LP) performance did not change, whereas total running distance (TD) during the incremental test decreased in ITV compared to an increase in 2 LP, 4 LP and TD during ITI which may indicate that there was an ITV-related overtraining. The NME of the reference muscles vastus medialis and rectus femoris deteriorated in ITV (day 28 compared to 0) compared to constant values during ITI, reflecting an ITV-related overload of neuromuscular structures.     

Mechanism of action of cholera toxin: Effect of receptor density and multivalent binding on activation of adenylate cyclase

Summary Cl^– influx into cells ofChara corallina is shown to be stimulated by a factor of 2 to 4 by starvation of Cl^–. The time constant for the induction of this effect is about 4.0 ksec and that for its decay when Cl^– is reprovided, 1.7 ksec. Intracellular perfusion of tonoplast-free cells with solutions of varying Cl^– concentration shows that Cl^– influx can be controlled directly by the concentration of Cl^– at the inside of the plasma membrane. Both the time course for the initial stages of induction of the starvation-stimulated flux and its absolute magnitude can be accounted for by assuming cytoplasmic Cl^– concentration to be the only intracellular condition to change during Cl^– starvation. The existence of a feedback loop between cytoplasmic Cl^– and Cl^– influx provides an alternative explanation to observations previously used in support of a Cl^–/OH^– exchange hypothesis (F.A. Smith, 1972,New Phytol. 71:595).