L-lactate Accumulation Research Articles

Amperometric monitoring of lactate accumulation in rabbit ischemic myocardium

Fabrication and characterization of miniature, flexible, planar biosensors for monitoring l-lactate accumulation in an ischemic myocardium are described. Three configurations of Au-based electrodes were fabricated by a photolithographic technique on flexible polyimide Kapton ® foil. All sensors are based on an immobilized lactate oxidase with amperometric detection of the enzymatically produced hydrogen peroxide at a platinum-electroplated-gold base electrode polarized at 0.5 V versus Ag/AgCl. An inner electropolymeric layer is used to prevent electrode fouling and to reject the interference effects of easily oxidizable molecules. In addition, a diffusion controlling outer layer that greatly enhances the linear dynamic range of the sensor, is obtained by casting a polyurethane external film. The developed sensor was evaluated in vitro and proved to have high selectivity, good operational stability, good accuracy and precision (average recovery = 102.3 ± 0.4% for control sera), fast response time ( t 95 = 20 s) and high upper limit of the linear dynamic range (25–80 mM, with sensitivity of 1.7-0.4 nA mM −1 respectively at PO 2 = 15 mmHg). Subsequently, the sensor was brought into direct contact with the surface of the rabbit papillary muscle and used for continuous quantitative monitoring of extracellular lactate accumulation during no-flow ischemia.

Characterization of the Enhanced Transport of L- and D-Lactate into Human Red Blood Cells Infected with Plasmodium falciparum Suggests the Presence of a Novel Saturable Lactate Proton Cotransporter

Human erythrocytes parasitized with the malarial protozoan Plasmodium falciparum showed rates of L-lactate, D-lactate, and pyruvate uptake many fold greater than control cells. Thus it was necessary to work at 0 degrees C to resolve true initial rates of transport. Studies on the dependence of the rate of transport on substrate concentration implied the presence in parasitized cells of both a saturable mechanism blocked by alpha-cyano-4-hydroxycinnamate (CHC) and a nonsaturable mechanism insensitive to CHC. The former was dominant at physiological substrate concentrations with Km values for pyruvate and D-lactate of 2.3 and 5.2 mM, respectively, with no stereoselectivity for L- over D-lactate. CHC was significantly less effective as an inhibitor of lactate transport in parasitized erythrocytes than in uninfected cells, whereas p-chloromercuribenzenesulfonate, a potent inhibitor in control cells, gave little or no inhibition of lactate transport into parasitized erythrocytes. Inhibition of transport into infected cells was also observed with phloretin, furosemide, niflumic acid, stilbenedisulfonate derivatives, and 5-nitro-2-(3-phenylpropylamino)benzoic acid at concentrations similar to those that inhibit the lactate carrier of control erythrocytes. These compounds were more effective inhibitors of the rapid transport of chloride into infected cells than of lactate transport, whereas CHC was more effective against lactate transport. This implies that different pathways are involved in the parasite-induced transport pathways for lactate and chloride. The transport of L-lactate into infected erythrocytes was also inhibited by D-lactate, pyruvate, 2-oxobutyrate, and 2-hydroxybutyrate. The intracellular accumulation of L-lactate at equilibrium was dependent on the transmembrane pH gradient, suggesting a protogenic transport mechanism. Our data are consistent with lactate and pyruvate having direct access to the malarial parasite, perhaps via the proposed parasitophorous duct or some close contact between the host cell and parasite plasma membranes, with transport across the latter by both a proton-linked carrier (CHC-sensitive, saturable, and the major route) and free diffusion of the undissociated acid (CHC-insensitive, unsaturable, and a minor route).

Responses in ciliated protozoa and rumen fermentation in sheep supplemented with barley plus virginiamycin

An experiment was conducted to determine the changes in the population of ciliate protozoa and the pattern of rumen fermentation in sheep fed a supplement of barley grain on its own or containing virginiamycin. There were three treatment groups with a total of 25 sheep housed in individual pens. All animals were fed chaffed wheaten hay to a maximum of 1.4 kg head-1 day-1. One group received no grain supplement (n = 5), one barley grain (n = 10) and the third barley plus virginiamycin (40 g/t grain; n = 10). The amount of grain was gradually increased over a period of 5 days to a maximum of 700 g of barley or barley plus virginiamycin on day 5. Grain (700 g/head) was then fed twice weekly for a further 18 days. Samples of rumen fluid were taken on days 1, 2, 3, 5*, 6, 9*, 12*, 23* and 24 (* signifies days when 700 g barley was fed). In sheep supplemented with barley the population density of ciliate protozoa was increased (P < 0.01) from day 3 of the 5 day introduction to grain (approximately 170x104 protozoa/mL at day 5), but the increase was not sustained when grain was fed at intervals of 3 or 4 days. In sheep supplemented with barley plus virginiamycin, the population density of ciliate protozoa was increased significantly (P < 0.01) only at days 5 and 6 of the experiment (approximately 100x l04 protozoa/mL at day 5). Sheep supplemented with barley plus virginiamycin at day 5 had a higher concentration of L-lactate (P < 0.05) at 6 and 12 h and propionate (P < 0.05) at 6, 9, 12 and 24 h than sheep supplemented with barley alone. It was concluded that virginiamycin was inhibitory to protozoa during the introduction to grain, and L-lactate accumulation in the rumen of sheep supplemented with barley plus virginiamycin may be a result of altered fermentation associated with reduced numbers of protozoa.

Demonstration of organic anion transport in T lymphocytes. L-lactate and fluo-3 are target molecules.

In this paper, we describe for the first time the existence of organic anion transport in T lymphocytes, exemplified by the transmembrane transport of the anions L-lactate and the Ca2+ indicator fluo-3. The transport of either anion was found to be inhibitable by probenecid, a common blocker of organic anion transport. Transport of L-lactate was observed in long-term cultured T cell lines, as well as in freshly ex vivo isolated T cells, and occurred via a saturable, pH-dependent, and stereospecific process. L-Lactate uptake was dependent on the activation state of the T cells, because activation of T cells by Con A strongly enhanced accumulation of L-lactate from the medium. Because L-lactate may be transported bidirectionally through the T cell membrane in vivo, different physiologic roles of L-lactate transport are discussed. L-Lactate uptake may serve as an alternative source of energy in an inflamed, glucose-deficient tissue or may represent a prerequisite for the earlier-published immunoregulatory function of this molecule on T cells. On the other hand, release of L-lactate emerging from glycolysis could be necessary to avoid acidification of the cell. The fact that the Ca2+ indicator fluo-3 is also transported through the cellular membranes of long-term cultured T cells via organic anion transport has important implications for the determination of Ca2+ influx into T cells. Even though the transport of both molecules, L-lactate and fluo-3, represents organic anion transport, evidence is presented that confirms that the respective transport systems are different.

Changes within the digestive tract of sheep following engorgement with barley

The effect of feeding 1.4 kg of barley on the amount and composition of digesta in the rumen, small intestine, caecum, and colon was measured in sheep given either a gradual introduction to barley (over 8 days), no introduction, Yea Sacc (4 g/day for 9 days), virginiamycin (30 mg/day for 4 days) or rumen inoculum (600 mL/day for 4 days). The rumen pH was higher ( P < 0.05) and the number of sheep with high (> 5 mmol/L) levels of L-lactate and D-lactate significantly lower (P < 0.05) in sheep receiving no introduction compared to sheep receiving the gradual introduction to barley. Conversely the pH was lower (P < 0.05) and the molar proportion of L-lactate higher ( P < 0.001) in the caecum of the sheep receiving no introduction compared to sheep receiving the gradual introduction to barley. There was no difference (P > 0.05) in the incidence of ruminitis between treatment groups. Both virginiamycin and the transfer of rumen fluid from well adapted animals appeared to be as effective as the gradual introduction of barley in controlling L-lactate accumulation in the caecum and colon and maintaining the pH within these organs. The probiotic Yea Sacc did not appear to cause any changes in the pattern of fermentation and digestion when compared to the untreated control animals.

Adaptations to a terrestrial existence in the robber crab, Birgus latro L. — IV. l-lactate dehydrogenase function and l-lactate accumulation during exercise

1. 1. The activity of l-lactate dehydrogenase (LDH) and the generation of l-lactate in tissues of resting and exercised Birgus latro was investigated. 2. 2. Comparisons of efferent and afferent blood from the branchial and pulmonary circuits showed decreases in l-lactate content during transit of gills and lungs. This decrease was significant in the case of the lung and suggested that l-lactate oxidation occurred. 3. 3. Leg muscle had a very high specific activity of LDH (112 μmol/min/g FW) for the reduction of pyruvate which correlated well with the high concentration of l-lactate ( > 25 mmol/l ) measured in the haemolymph following exercise. Lung and gill tissue had lower LDH activity (1.74 and 0.09 μmol/min/g FW respectively). 4. 4. The lungs and especially the leg muscle had exhibited activity of LDH in the direction of l-lactate oxidation (0.03 and 0.12 μmol/min/g FW respectively). 5. 5. Birgus possessed good anaerobic support for sustained (30 min) activity but no specific site of lactate oxidation was definitely identified. 6. 6. LDH in the lung exhibited the greatest l-lactate oxidation activity relative to that for l-lactate production. Some Cori cycle activity was therefore possible, especially when circulating lactate levels were high, but the extent of this must be small and thus repayment of the oxygen debt must occur substantially during the post exercise phase.

Transport of Methionine Hydroxy Analog across the Brush Border Membrane of Rat Jejunum

Brush border membrane vesicles and everted rings of rat jejunum were used to study the absorption of 2-hydroxy-4-methylthiobutyric acid, the α-hydroxy analog of methionine (DL-MHA). Uptake by membrane vesicles was found to be unaltered by an inwardly directed sodium gradient in contrast to that of L-methionine, D-methionine and L-lactate, which is sodium stimulated and exhibits a characteristic “overshoot” phenomenon. Initial uptake rates of both D- and D-methionine (0.77 mM) were respectively 6.8 and 1.5 times higher than that of DL-MHA (0.80 mM) under Na+-gradient conditions. However, the hydroxy analog significantly inhibited L-lactate accumulation into membrane vesicles. Uptake of DL-MHA into rat jejunal rings was the sum of a saturable Michaelian component [KappM = 4.4 mM and Vappmax = 587 nmol/(g·min)] and a diffusive component [Kd = 47 µL/(g·min)]. Furthermore, L-lactate completely inhibited the carrier-mediated uptake of DL-MHA. All this demonstrates that jejunal uptake of DL-MHA in the rat is mediated by an Na+-Independent carrier system associated with L-lactate transport and thus is quite distinct from the L-methionine pathway. This difference could help explain numerous findings with respect to a lower biological utilization of the hydroxy analog in some species when fed as the sole source of methionine in purified crystalline amino acid diets.

Effects of hypoxia and l-lactate on the haemocyanin-oxygen affinity of the lobster, Homarus vulgaris

1. 1.Dialysis of lobster haemolymph against a physiological saline entailed a decrease of the haemocyanin-oxygen affinity. 2. 2. L-Lactate added to dialysed haemolymph increased the oxygen affinity of the haemocyanin without changing the Bohr effect. The effect of adding D-lactate was much less. 3. 3.In lobsters exposed to ambient hypoxia ( P O 2 = 35 Torr at 15°C), whole haemolymph oxygen affinity increased irrespective of pH. This in vivo increase cannot be accounted for by L-lactate accumulation alone, but must involve another, as yet unidentified dialysable factor.

Sodium ion/L-lactate co-transport in rabbit small-intestinal brush-border-membrane vesicles.

Uptake of L-lactate into rabbit jejunal brush-border-membrane vesicles prepared by a Ca2+-precipitation procedure was studied by a rapid filtration technique with L-[14C]-lactate as tracer. Transport of L-lactate into an intravesicular (osmotically reactive) space could be established. An inwardly directed NaCl gradient (outside 21 mM/inside 0mM) stimulated the uptake of L-lactate at 15 s 2-4-fold compared with that observed with an equal KCl gradient. A transient accumulation of L-lactate inside the vesicles (overshoot) was observed in the presence of an NaCl gradient. Gradients of LiCl, RbCl, CsCl or choline chloride were not able to replace NaCl in the stimulation of L-lactate uptake. L-Lactate uptake was saturable only in the presence of Na+. D-Lactate, DL-thiolactate (2-DL-mercaptopropionate), pyruvate and propionate inhibited the Na+-stimulated L-lactate uptake; D-lactate, thiolactate and pyruvate provoked trans-stimulation of L-lactate uptake. Artificially imposed diffusion potentials (inside negative) did not exert any effect on the Na+-dependent L-lactate uptake. The results are consistent with the existence of an electroneutral Na+/L-lactate co-transport system in the brush border of rabbit small intestine.