255
554th MEETING, LONDON
Inhibitionby Hodoacetate of ‘Downhill’ Transport of Sodium Ions across Isolated Rat Small Intestine MICHAEL L. G. GARDNER Department of Biochemistry, University Medical School, Teviot Place, Edinburgh EH8 9AG, U.K. The net transport of Na+ ions across intestinal mucosa, from lumen to serosal pole, is accepted to be an activetransport process. A ouabain-sensitive sodium pump, thought to be in close association with (Na++K+)-stimulated ATPase* activity, is located in the serosal membrane of these epithelial cells and maintains a low concentration of Na+ and a high concentration of K+ inside the cells. Thus application of ouabain to the serosal surface of the epithelium results in a decreased net flux of Na+ from lumen to serosal pole (e.g. Schultz & Zalusky, 1964) and in an elevated intracellular concentration of Na+ (e.g. Schultz et al., 1966). In some way, probably connected with Na+ movements or concentrations, ouabain also inhibits the active absorption of glucose from the intestinal lumen (e.g. Fisher & Gardner, 1974). However, if the Na+ concentration in the tissue fluid bathing the serosal surface of these cells is kept low by arterial infusion then ouabain has no eKect on glucose absorption nor on the net rate of appearance of Na+ ions in the vascular (serosal) effluent (Fisher & Gardner, 1974). Thus ouabain inhibition of the sodium pump appears to have no effect on net Na+movement when the transport of these ions is down the electrochemical gradient. This communication describes the effect of a glycolytic inhibitor, iodoacetate, on such ‘downhill’ transport of Na+ across isolated perfused rat small intestine. The jejunum and ileum were perfused by the arteria1:infusion technique of Fisher & Gardner (1974); see Fig. 1. The medium infused into the superior mesenteric artery was the bicarbonate-saline solution of Fisher & Gardner (1974) in which the sodium salts had been replaced by salts of choline. Initially the luminal perfusate was the normal medium of Fisher & Gardner (1974) containing sodium (145mequiv./litre) and glucose (28m); after control measurements had been made this was replaced by one which also contained sodium iodoacetate (0.1 m).Serial measurements were made of the rate,of appearance of Na+ in the serosal (vascular) effluent over Smin periods. After the introduction of the iodoacetate the net rate of Na+ transport decreased
* Abbreviation : ATPase, adenosine triphosphatase.
(t inhibitor)
“a’]
4
(Na+-free)
= 115 mequiv./l
5
Mucoral cells
/ /
&. / / /
4
. 8 m
a/
L L
n/
= n 10 E
P/ 5+
Vol. 3
4
/
“a’]
I 19
mequiv./l
256
BIOCHEMICAL SOCIETY TRANSACTIONS
5
10
IS
20
25
30
35
40
45
Time (min) Fig. 2. Rate of secretion of Nu+ ions into serosal (vascular)effruentbefore and a f e r introduction of iodoacetate in the luminal perfusate Sodium was present (145mequivJitre) in the luminal perfusate but absent from the arterial infusate. After three 5min control periods sodium iodoacetate (0.1 mM) was introduced into the luminal perfusate. Values are means ~ s . E . M .of eight experiments.
(Fig. 2). The Na+ concentration in the intestinal lumen was 145mequivJitre and the mean concentration in the serosal effluent was 19mequiv./litre.Therefore Na+ movement was definitely down a concentration gradient. It also appeared to be down an electrochemical gradient since the equilibrium (Nernst) potential calculated for this distribution of Na+ activity (55mV) is much greater than the normal maximum potential difference across the intestine in the presence of glucose of about 12mV (Barry et al., 1964). The latter would itself be decreased by the iodoacetate. Preliminary experiments with an alternative metabolic inhibitor, 2,4dinitrophenol, have shown the same inhibition of 'downhill' Na+ movement. This is in contrast with the effects of ouabain which only inhibited Na+ movement when there was a relatively high Na+ concentration in the arterial infusate (Fisher & Gardner, 1974). Separate control experiments without any inhibitor showed that the rate of Na+ movement remained constant after a rise during the initial 5-l0min. Thus 'downhill' net movement of Na+ ions appears to require metabolicenergy. Either this requirement is at a stage other than the ouabain-sensitiveNa+ pump, or else the Na+ pump is inhibited by ouabain only in the presence of a relatively high concentration of Na+ (>72mequiv./litre) in the serosal extracellular fluid. Barry, R. J. C.. Dickstein, S., Matthews, J., Smyth, D. H. & Wright, E. M. (1964)J. Physiol. (London)171,316338 Fisher, R. B. & Gardner, M. L. G. (1974)J. Physiol. (London) 241,235-260 Schultz, S. G.& Zalusky, R. (1964)J. Gen. Physiol. 47,567-584 Schultz, S.G., Fuisz, R. E. & Curran, P.F. (1966)J. Gen. Physiol. 49,849-866
Characteristics of Intact Lactose Transport in the Rat Jejunum in v i m J. E. JOSfi OYESIKU, D. P. R. MULLER and J. T. HARRIES Institute of Child Health, 30 Guilford Street, London WC1N 1 EH, U.K.
The jejunal transport of glucose and galactose is carrier-mediated, sodium- and energydependent. The characteristics of intact disaccharide transport, however, have not been defined, This paper reports on the magnitude and kinetics of transmural transport of 1975
554th MEETING, LONDON
Inhibitionby Hodoacetate of ‘Downhill’ Transport of Sodium Ions across Isolated Rat Small Intestine MICHAEL L. G. GARDNER Department of Biochemistry, University Medical School, Teviot Place, Edinburgh EH8 9AG, U.K. The net transport of Na+ ions across intestinal mucosa, from lumen to serosal pole, is accepted to be an activetransport process. A ouabain-sensitive sodium pump, thought to be in close association with (Na++K+)-stimulated ATPase* activity, is located in the serosal membrane of these epithelial cells and maintains a low concentration of Na+ and a high concentration of K+ inside the cells. Thus application of ouabain to the serosal surface of the epithelium results in a decreased net flux of Na+ from lumen to serosal pole (e.g. Schultz & Zalusky, 1964) and in an elevated intracellular concentration of Na+ (e.g. Schultz et al., 1966). In some way, probably connected with Na+ movements or concentrations, ouabain also inhibits the active absorption of glucose from the intestinal lumen (e.g. Fisher & Gardner, 1974). However, if the Na+ concentration in the tissue fluid bathing the serosal surface of these cells is kept low by arterial infusion then ouabain has no eKect on glucose absorption nor on the net rate of appearance of Na+ ions in the vascular (serosal) effluent (Fisher & Gardner, 1974). Thus ouabain inhibition of the sodium pump appears to have no effect on net Na+movement when the transport of these ions is down the electrochemical gradient. This communication describes the effect of a glycolytic inhibitor, iodoacetate, on such ‘downhill’ transport of Na+ across isolated perfused rat small intestine. The jejunum and ileum were perfused by the arteria1:infusion technique of Fisher & Gardner (1974); see Fig. 1. The medium infused into the superior mesenteric artery was the bicarbonate-saline solution of Fisher & Gardner (1974) in which the sodium salts had been replaced by salts of choline. Initially the luminal perfusate was the normal medium of Fisher & Gardner (1974) containing sodium (145mequiv./litre) and glucose (28m); after control measurements had been made this was replaced by one which also contained sodium iodoacetate (0.1 m).Serial measurements were made of the rate,of appearance of Na+ in the serosal (vascular) effluent over Smin periods. After the introduction of the iodoacetate the net rate of Na+ transport decreased
* Abbreviation : ATPase, adenosine triphosphatase.
(t inhibitor)
“a’]
4
(Na+-free)
= 115 mequiv./l
5
Mucoral cells
/ /
&. / / /
4
. 8 m
a/
L L
n/
= n 10 E
P/ 5+
Vol. 3
4
/
“a’]
I 19
mequiv./l
256
BIOCHEMICAL SOCIETY TRANSACTIONS
5
10
IS
20
25
30
35
40
45
Time (min) Fig. 2. Rate of secretion of Nu+ ions into serosal (vascular)effruentbefore and a f e r introduction of iodoacetate in the luminal perfusate Sodium was present (145mequivJitre) in the luminal perfusate but absent from the arterial infusate. After three 5min control periods sodium iodoacetate (0.1 mM) was introduced into the luminal perfusate. Values are means ~ s . E . M .of eight experiments.
(Fig. 2). The Na+ concentration in the intestinal lumen was 145mequivJitre and the mean concentration in the serosal effluent was 19mequiv./litre.Therefore Na+ movement was definitely down a concentration gradient. It also appeared to be down an electrochemical gradient since the equilibrium (Nernst) potential calculated for this distribution of Na+ activity (55mV) is much greater than the normal maximum potential difference across the intestine in the presence of glucose of about 12mV (Barry et al., 1964). The latter would itself be decreased by the iodoacetate. Preliminary experiments with an alternative metabolic inhibitor, 2,4dinitrophenol, have shown the same inhibition of 'downhill' Na+ movement. This is in contrast with the effects of ouabain which only inhibited Na+ movement when there was a relatively high Na+ concentration in the arterial infusate (Fisher & Gardner, 1974). Separate control experiments without any inhibitor showed that the rate of Na+ movement remained constant after a rise during the initial 5-l0min. Thus 'downhill' net movement of Na+ ions appears to require metabolicenergy. Either this requirement is at a stage other than the ouabain-sensitiveNa+ pump, or else the Na+ pump is inhibited by ouabain only in the presence of a relatively high concentration of Na+ (>72mequiv./litre) in the serosal extracellular fluid. Barry, R. J. C.. Dickstein, S., Matthews, J., Smyth, D. H. & Wright, E. M. (1964)J. Physiol. (London)171,316338 Fisher, R. B. & Gardner, M. L. G. (1974)J. Physiol. (London) 241,235-260 Schultz, S. G.& Zalusky, R. (1964)J. Gen. Physiol. 47,567-584 Schultz, S.G., Fuisz, R. E. & Curran, P.F. (1966)J. Gen. Physiol. 49,849-866
Characteristics of Intact Lactose Transport in the Rat Jejunum in v i m J. E. JOSfi OYESIKU, D. P. R. MULLER and J. T. HARRIES Institute of Child Health, 30 Guilford Street, London WC1N 1 EH, U.K.
The jejunal transport of glucose and galactose is carrier-mediated, sodium- and energydependent. The characteristics of intact disaccharide transport, however, have not been defined, This paper reports on the magnitude and kinetics of transmural transport of 1975
