Download CCK - 8 Sulphate Inhibits Intestinal Glucose Transport

592s Biochemical SocietyTransactions ( 1 995) 23
CCK - 8 Subhate Inhibits Intestinal Glucose Transport
M. Ekhesty Tabar and K. Burdett
Division of Biochemistry, The School of Biological
Sciences, The University of Manchester, MI3 9PT, UK
Gastrointestinal hormones and neurotransmitters regulate
food intake, digestion and distribution of nutrients. We are
interested in the acute regulation of non-electrolyte
absorption, in particular by events early in the digestive
process. Conditions in the gastric lumen as the meal is
terminating stimulate the release of a novel fundic peptide
that decreases intestinal glucose absorption [I]. Well
characterised peptides also act at this time. Cholecystokinin
(CCK) released from the proximal small intestine early
after ingestion [2] has been implicated in the regulation of
food intake and in the modulation of postprandial blood
glucose concentrations. CCK acts by delaying gastric
emptying and hence the supply of absorbable carbohydrate
to the proximal intestine [3]. It could do so by classic
endocrine action or by paracrine activity of peptide released
from the pyloric region of the stomach [4]. In man CCK
infusion delayed gastric glucose emptying but had no effect
on plasma glucose concentrations when glucose was
instilled directly into the duodenum [3]. There is no
published evidence that CCK directly affects glucose
transport from the intestine by paracrine or endocrine
action, though it may modulate distribution thereafter by
inducing insulin release [5].
Studying the effect of CCK on glucose transport through
mucosal sheets in vitro has the advantage that any other
important physiological effects such as changes in motility
and insulin release do not impinge on the data.
The methods used were those described previously [l].
5 mm diameter mucosal sheets from guinea pig jejunum or
ileum were equilibrated in glucose-free buffer for 30
minutes and then incubated for a further 45 minutes at
37°C with 5 mM glucose in the luminal buffer and 1 )LM
CCK-8 sulphate in 0.1% albumin in the serosal buffer.
The paired control sheets were exposed to 0.1% albumin.
'H polyethylene glycol 900 was added to the luminal
buffer; if more than 0.5% of the luminal dose appeared on
the serosal side the sheet was judged to be damaged. The
difference in serosal glucose gain between each sheet and
its paired control was measured. Replicate measurement of
glucose transport in control sheets showed that ileal sheets
absorbed approximately twice as much glucose as jejunal
sheets and that the intrinsic variation between sheets taken
from any one animal had a standard deviation of
3% of
the mean.
CCK-8 sulphate in the serosal compartment decreased the
transport of glucose by 40.8 f 13.6% (n=22 pairs of
sheets from 5 guinea pigs). The inhibition was consistent
and highly significant when the serosal CCK-8 sulphate
concentration was 1 M. Inhibition was not a non-specific
peptide effect, 3 mg ml of a low molecular weight peptide
mixture extracted from pig antral mucosa had no effect.
r
Preliminary experiments with the non-sulphated CCK
octapeptide showed that it too inhibits glucose transport
(41.9 f 18%, n = 11 pairs of sheets from 2 guinea pigs).
The CCK-8 effect appears to be more variable and to
require a higher serosal concentration (4 pM) of peptide.
Whether CCK-8 sulphate is genuinely a better agonist will
be investigated in future work. The sulphated octapeptide.
but not CCK-8, raised plasma insulin concentrations in
mice [6], so some biological systems do discriminate
between the two peptides. 1 pM is a high dose of CCK-8
sulphate and may be needed in vitro to survive proteolysis
and to produce a big enough gradient to drive the hormone
on to mucosal receptors within the sheet. CCK-8 sulphate
infused in vivo at a rate of 15 pmole/min/kg in guinea pigs
[7] significantly reduces gastrointestinal glucose absorption.
The inhibitory action of CCK-8 non-sulphated is being
used in the design of experiments to determine the
minimum amino acid sequence needed to inhibit intestinal
glucose transport, and any relationship between CCK-8 and
the novel inhibitory peptide from the gastric fundic mucosa
ill.
We gratefully acknowledge the financial support of The
Islamic Republic of Iran for M.B.T.
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Beccaria, L., Carter, J. D., & Goldfine, I. D.
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Reagan, J. E., Robinson, J. L., Lotti, V. J. &
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Mottaghitalab, M., & Burdett. K. (1995)
Unpublished results