Isc In Tissues Research Articles

Effects of Enterococcus faecium NCIMB 10415 as probiotic supplement on intestinal transport and barrier function of piglets

Many studies report positive effects of probiotic supplementation on the performance and health of piglets. The intention of this study was to describe the effects of Enterococcus faecium NCIMB 10415 on the transport and barrier functions of pig small intestine to improve our understanding of the underlying mechanisms of this probiotic. Ussing chamber studies were conducted with isolated jejunal epithelia of piglets at the age of 14, 28, 35 and 56 days. Jejunal tissues of the control group were compared with epithelia of piglets that had received a diet supplemented with the probiotic Enterococcus faecium NCIMB 10415. Transport properties (absorption and secretion) of the epithelia were examined by mucosal addition of glucose or L-glutamine or by serosal addition of PGE2. Electrophysiology of the epithelia was continuously recorded and the change in short circuit current (Isc) was determined. Paracellular permeability was measured by measuring the flux rates of mannitol. The increase of Isc caused by mucosal addition of glucose was, at all glucose concentrations, higher in the probiotic group compared with the control group. However, the difference (up to 100% of the control) was not significant. The increase of Isc after the mucosal addition of L-glutamine (12 mmol/l) was higher in the tissues of the probiotic group but did not reach significance. Serosal PGE2 induced a significantly higher increase of Isc in tissues of the probiotic group at the age of 28 days. No consistent differences were observed in mannitol transport rates between the feeding groups. Significant age-dependent alterations of absorptive and secretory properties of the jejunal epithelium were observed; these were independent of the treatment. A probiotic supplementation seems to influence transport properties of small intestine epithelium. The increased absorption of glucose could be interpreted as a positive effect for the animal.

Dextran sodium sulphate-induced colitis perturbs muscarinic cholinergic control of colonic epithelial ion transport.

1. Neuronal cholinergic input is an important regulator of epithelial electrolyte transport and hence water movement in the gut. 2. In this study, colitis was induced by treating mice with 4% (w v(-1)) dextran sodium-sulphate (DSS)-water for 5 days followed by 3 days of normal water. Mid-colonic segments were mounted in Ussing chambers and short-circuit current (Isc, indicates net ion movement) responses to the cholinergic agonist, carbachol (CCh; 10(-4) M)+/-tetrodotoxin, atropine (ATR), hexamethonium (HEX), naloxone or phenoxybenzamine were assessed. 3. Tissues from mice with DSS-induced colitis displayed a drop in Isc in response to CCh (-11.3+/-3.3 microA/cm(2)), while those from control mice showed a transient increase in Isc (76.3+/-13.0 microA/cm(2)). The DeltaIsc in colon from DSS-treated mice was tetrodotoxin-sensitive, atropine-insensitive and was reversed by hexamethonium (HEX+CCh=16.7+/-7.8 microA/cm(2)), indicating involvement of a nicotinic receptor. 4. CCh induced a drop in Isc in tissues from controls only when they were pretreated with the cholinergic muscarinic receptor blocker, atropine: ATR+CCh=-21.3+/-7.0 microA/cm(2). Nicotine elicited a drop in Isc in Ussing-chambered colon from both control and DSS-treated mice that was TTX-sensitive. 5. The drop in Isc evoked by CCh challenge of colonic tissue from DSS-treated mice or ATR+CCh challenge of control tissue was not significantly affected by blockade of opiate or alpha-adrenergic receptors by naloxone or phenoxybenzamine, respectively. 6. The data indicate that DSS-colitis reveals a nicotinic receptor that becomes important in cholinergic regulation of ion transport.

Modulation of Cl − secretion in rat distal colon by genistein, a protein tyrosine kinase inhibitor

The protein tyrosine kinase inhibitor, genistein, caused an increase of short-circuit current ( Isc) across the rat distal colon in forskolin-pretreated tissues, suggesting a synergistic interaction of the drug with cAMP-dependent secretion. In the absence of forskolin, genistein had a dual effect on Isc, it increased Isc in tissues with a low baseline, but decreased Isc in tissues with a high baseline Isc. The secretory effect of genistein was dependent on the presence of Cl − and was blocked by inhibitors of Cl − secretion like bumetanide, an inhibitor of the Na +-K +-Cl − cotransporter, or 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB), a Cl − channel blocker. Unidirectional flux measurements revealed that genistein inhibited Na + and Cl − absorption and induced net Cl − secretion. The protein tyrosine phosphatase inhibitor vanadate suppressed the secretory effect of genistein. In contrast, genistein caused an inhibition of carbachol-induced, i.e. Ca 2+-mediated secretion. Whole-cell patch-clamp experiments confirmed the synergistic effect of genistein on cAMP-induced Cl − currents. In the presence of forskolin, genistein caused a depolarization concomitant with an increase in membrane inward current. In addition, genistein caused an inhibition of a basal K + conductance and inhibited the Ca 2+-dependent K + conductance stimulated by carbachol. These results suggest a complex role of the protein tyrosine kinase pathway in the control of colonic Cl − secretion, an antagonistic action on the cAMP pathway and a synergistic action on the Ca 2+ pathway as revealed by the opposing effects of genistein. The physiological importance of this regulation remains to be clarified.

Potassium dependence of Na-Cl cotransport in dog tracheal epithelium

In confluent primary cultures of dog tracheal epithelium, we tested whether Cl entry across the basolateral membrane is by cotransport with K. Two approaches were taken. First, we measured the inhibition of short-circuit current (Isc) by the K channel inhibitor, Ba2+. Consistent with Na-K-2Cl cotransport, maximal doses of Ba2+ inhibited five-sixths of Isc in tissues previously stimulated to secrete Cl; only two-thirds of Isc should be sensitive to Ba2+ if NaCl cotransport is the entry mechanism. Second, we measured basolateral 86Rb uptake and demonstrated inhibition by bumetanide, an inhibitor of Na-K-2Cl cotransport in other tissues. The degree of inhibition by bumetanide was consistent with the levels of Cl secretion measured as Isc. Uptake of 86Rb was also reduced by removal of Na or Cl, and under these conditions Rb uptake was not further inhibited by bumetanide. These results suggest that the process responsible for Cl entry across the basolateral membrane of tracheal epithelium during Cl secretion is Na-K-2Cl rather than Na-Cl cotransport.

CAMP-mediated regulation of chloride secretion by the opercular epithelium.

Catecholamine regulation of the Cl- secretion rate (short-circuit current, Isc) and adenosine 3',5'-cyclic monophosphate (cAMP) levels of the opercular epithelium was investigated by using 3-isobutyl-1-methylxanthine (IBMX), forskolin, and adrenergic agonists. In this tissue alpha-adrenergic agonists inhibit, and beta-adrenergic agonists stimulate, the Isc (J. Physiol. London 294: 483-495, 1979). IBMX and forskolin stimulated the Isc 125 and 85%, respectively, and simultaneously produced 2.5- and 70.0-fold elevations in the cAMP levels, respectively. These findings confirm previous observations demonstrating that stimulation of the Isc in this tissue is mediated by elevations in cAMP (J. Comp. Physiol. B 145: 29-35, 1981). Isoproterenol, a beta-agonist, had no effect on the Isc of either IBMX- or forskolin-stimulated tissues but increased the cAMP level an additional 5.8-fold in IBMX-stimulated tissues. Clonidine, an alpha-agonist, inhibited the Isc in IBMX-stimulated tissues only and had no effect on cAMP levels in either IBMX- or forskolin-stimulated tissues. These findings demonstrate that catecholamine-induced inhibition of the Isc can occur while the cAMP level remains elevated, indicating that this effect is not mediated by lowering cAMP levels. This observation is strong evidence for a cAMP-independent mechanism for catecholamine-induced inhibition of Cl- secretion in the opercular and similar epithelia.