A Creatine Transporter Is Operative at the Brush Border Level of the Rat Jejunal Enterocyte

Abstract Although ergogenic effects and health benefits have been reported for creatine used as nutritional supplement, to date little is known about the mechanism of creatine absorption in the small intestine. Thus the current study was undertaken to elucidate the mechanism of creatine intake in rat jejunum with the use of well-purified brush border membrane vesicles, isolated from jejunal enterocyte. Creatine uptake was found markedly stimulated by inwardly directed Na(+) and Cl(- )gradients, potential-sensitive, strongly reduced by the substitution of Na(+) and Cl(-) with various cations and anions and positively affected by intravesicular K(+). Moreover, creatine uptake is: 1) significantly inhibited by creatine structural analogs, 2) abolished by low concentrations of 2-aminoethyl methanethiosulfonate hydrobromide (MTSEA), 3) saturable as a function of creatine concentration with an apparent Michaelis-Menten constant of 24.08 +/- 0.80 muM and a maximal velocity of 391.30 +/- 6.19 pmoles mg protein(-1) 30 s(-1). The transport is electrogenic since at least two Na(+) and one Cl(-) are required to transport one creatine molecule. Western blot analysis showed the same amount of creatine transport protein in the jejunal apical membrane when compared to ileum. Thus, these data demonstrate the existence of a Na(+)- and Cl(-)-dependent, membrane potential-sensitive, electrogenic carrier-mediated mechanism for creatine absorption in rat jejunal apical membrane vesicles, which is biochemically and pharmacologically similar to those observed in other tissues. However, in other cell types the stimulatory effect of intravesicular K(+) was never detected

Action observation combined with conventional training improves the rugby lineout throwing performance: A pilot study

Combining action observation (AO) and physical practice contributes to motor skill learning, and a number of studies pointed out the beneficial role of AO training in improving the motor performance and the athletes' movement kinematics. The aim of this study was to investigate if AO combined with immediate conventional training was able to improve motor performance and kinematic parameters of a complex motor skill such as the lineout throw, a gesture that represents a key aspect of rugby, that is unique to this sport. Twenty elite rugby players were divided into 2 groups. The AO group watched a 5-min video-clip of an expert model performing the lineout throw towards a target at 7 m distance and, immediately after the AO, this group executed the conventional training, consisting of 6 repetitions x 5 blocks of throws. The CONTROL group performed only the conventional lineout training. Intervention period lasted 4 weeks, 3 sessions/week. The AO group showed significant improvements in throwing accuracy (i.e. number of throws hitting the target), whilst no significant changes were observed in the CONTROL group. As concerns kinematic parameters, hooker's arm mean velocity significantly increased in both groups, but the increase was higher in AO group compared to CONTROL group. Ball velocity significantly increased only in the AO group, whereas ball angle release and ball spinning significantly decreased in both groups, with no differences between groups. Finally, no significant changes in knee and elbow angles were observed. Our results showed that the combination of AO and conventional training was more effective than a conventional training alone in improving the performance of elite rugby players, in executing a complex motor skill, such as the lineout. This combined training led to significant improvements in throwing accuracy and in hooker's and ball's kinematic parameters. Since AO can be easily implemented in combination with conventional training, the results of this study can encourage coaches in designing specific lineout training programs, which include AO cognitive training

Near term perspectives for fusion research and new contributions by the Ignitor program

The main advances made within the Ignitor program, that is aimed at investigating the physics of fusion burning plasmas near ignition, are described. In particular, the operation of the machine in the H and I regimes at the 10 MA plasma current levels has been considered and analyzed. The unique properties of the plasmas that can be generated by operating the machine with reduced parameters (lower magnetic fields and plasma currents) relative to those needed to achieve ignition are identified. A key feature of this operation is the relatively fast duty cycle that can be maintained. The Ideal Ignition Conditions, under which the density barrier due to bremsstrahlung emission in high density plasmas is removed, can be attained in this case. The plasma heating cycles are identified for which the contribution of ICRH is used both to enter the H-regime and to optimize the time needed for ignition. The on going effort to set up a test ICRH facility is described. The initial results (2 km/sec) of the high speed pellet injection system developed for Ignitor and operated at Oak Ridge are reported. The combined structural analysis and integration of the entire machine core (Load Assembly) is discussed. The adopted control system for both the machine and the plasma column has been designed and is described. The design solutions of the vertical field coils made of MgB2 and operating at 10 K have been identified and the relevant R&D program is underway. The analysis of the Caorso site and of its facility for the operation of the Ignitor with approved safety standards is completed. The relevant results are being made available for the operation of Ignitor at the Triniti site within the framework of the Italy-Russia agreement on the joint construction and operation of the Ignitor facility. A development effort concerning the advanced diagnostic systems that is being carried out for fusion burning plasma regimes is described. An initial analysis of the characteristics of a neutron source based on a system of Ignitor-like machines is reported

Effects of semistarvation on transintestinal d-glucose transport and d-glucose uptake in brush border and basolateral membranes of rat enterocytes

The present work shows that semistarvation (8-10 g of food for 10 days) increases net d-glucose, Na+ and water transport in the everted and perfused rat jejunum. A lincar and positive correlation between cell sugar concentration and transport was found in control and semistarved rats, but the phenomenon was more relevant only in semistarved animals. Membrane vesicle experiments showed that semistarvation increases sugar overshoot only in brush border membrane vesicles, while this situation does not occur in basolateral membrane vesicles. The effect of partial food deprivation seems to enhance net sugar transport by mereasing sugar entry across the apical membrane of enterocytes

An endogenous monocarboxylate transport in Xenopus laevis oocytes

We investigated the existence of an endogenous system for lactate transport in Xenopus laevis oocytes. Cl-36-uptake studies excluded the involvement of a DIDS-sensitive anion antiporter as a possible pathway for lactate movement. L-[C-14]lactate uptake was unaffected by superimposed pH gradients, stimulated by the presence of Na+ in the incubating solution, and severely reduced by the monocarboxylate transporter inhibitor p-chloromercuribenzenesulphonate (pCMBS). Transport exhibited a broad cation specificity and was cis inhibited by other monocarboxylates, mostly by pyruvate. These results suggest that lactate uptake is mediated mainly by a transporter and that the preferred anion is pyruvate. [C-14]pyruvate uptake exhibited the same pattern of functional properties evidenced for L-lactate. Kinetic parameters were calculated for both monocarboxylates, and a higher affinity for pyruvate was revealed. Various inhibitors of monocarboxylate transporters reduced significantly pyruvate uptake. These studies demonstrate that Xenopus laevis oocytes possess a monocarboxylate transport system that shares some functional features with the members of the mammalian monocarboxylate cotransporters family, but, in the meanwhile, exhibits some particular properties, mainly concerning cation specificity

Calmodulin-mediated regulation of bicarbonate and lactate transports in rat jejunum

BACKGROUND/AIMS: Ca(2+)/CaM is known to modulate the activity of several transport systems and its regulation can be accomplished either directly or via the involvement of specific protein kinases. Aim of this study was to investigate the possible role of Ca(2+)/CaM on bicarbonate and lactate transports in rat jejunal enterocyte. METHODS: Enzymatic assays in isolated plasma membranes were performed. Moreover membrane vesicles, transiently opened and resealed, were loaded with Ca(2+) and calmodulin, both in the absence and in the presence of ATP, and were used after that to perform uptake studies. RESULTS: Enzymatic assays gave evidence for the presence of Ca(2+)/CaM-dependent protein kinase II (CaMKII) in plasma membranes from rat jejunum. However, uptake experiments suggest that Ca(2+)/CaM, and not CaMKII, inhibits both basolateral Cl(-)/HCO(3)(-) exchange and H(+)-lactate symport, whilst HCO(3)(-) and Cl(-) conductances are unaffected. Neither Ca(2+)/CaM nor CaMKII seem to regulate brush border Na(+)/H(+) exchanger activity. CONCLUSION: These data are consistent with a Ca(2+)/CaM-mediated reduction of bicarbonate and lactate exit from jejunal enterocyte

PKA regulation of bicarbonate and lactate movements across rat jejunal plasma membranes

Background/Aims: Evidence was previously given that the mechanisms involved in bicarbonate and lactate movements across rat jejunal enterocyte are modulated by PKC and Ca2+/CaM. Aim of this study was to investigate the possible role of PKA on bicarbonate and lactate transports. Methods: Enzymatic assays in isolated plasma membranes were performed. Moreover membrane vesicles, transiently opened and resealed, were treated with a phosphorylating solution (leading to PKA activation) and were used after that to perform uptake studies. Results: Enzymatic assays give evidence for the presence of PKA in plasma membranes from rat jejunum. Uptake experiments suggest that PKA stimulates the two systems that accomplish basolateral HCO3- efflux from the enterocyte, namely Cl-/ HCO3- exchanger and HCO3- conductance, without affecting HCO3- influx from the lumen mediated by Na+/H+ exchanger activity. Moreover basolateral H+/lactate symporter is stimulated by PKA, as well as the brush border isoform of Na+-glucose cotransporter SGLT1. Conclusion: PKA activation evokes individual responses that could be coordinated through cellular metabolism

PKC regulation of rat jejunal transport systems : mechanisms involved in lactate movement

We examined whether PKC modulates the transport systems involved in lactate movements across the plasma membranes of rat jejunum. \u201cIn vitro\u201d phosphorylated membrane vesicles were used to perform uptake studies, whose results suggest that PKC activation exerts an inhibitory effect on basolateral H+-lactate symport, as well as on apical Na+-glucose cotransport. The specificity of the response to PKC was confirmed by using staurosporine, chelerythrine or 4-\uf061-PMA. Experiments performed using the whole tissue incubated "in vitro" confirmed the reduction of lactate transport elicited by PKC and gave evidence for a contemporaneous inhibition of fluid transport. (Na+, K+)-ATPase activity seems to be unaffected by the kinase and inhibited by Ca2+. Taken together, our results suggest that the overall action of PKC results from the contemporaneous modulation of multiple pathways, targeted to a reduction of both lactate and bicarbonate transports without altering cell pH homeostasis