TY - JOUR
T1 - A novel micromethod for pancreatic acinar secretion - Time course of CCK stimulation and its inhibition by L-364,718
AU - Adrian, Thomas E.
AU - Zucker, Karl A.
AU - Bilchik, Anton J.
AU - Modlin, Irvin M.
PY - 1990/1/1
Y1 - 1990/1/1
N2 - Dispersed acini have proven to be particularly valuable in the study of pancreatic enzyme secretion. Complex time course studies or experiments requiring large numbers of replicates have proven difficult, however, with currently available techniques. Using a custom-designed incubation chamber, a miniaturized incubation method has been devised, which allows for continuous oxygenation of acini in 96-well microtiter plates and rapid separation of medium from acini by vacuum filtration. The filtrates from individual wells are collected into the wells of a second microtiter plate for pancreatic enzyme measurement. Using the above method, the dose response and time course of cholecystokinin (CCK-8)-stimulated amylase secretion was investigated. During a 1-h incubation, unstimulated amylase secretion was 4.1 ±0.3% of total acini content. Response to CCK was very sensitive, being detected at 10-13 M (p<0.05), half-maximal at 10-11 M CCK (14.0±0.6%, p<0.001) and maximal at 10-9M CCK (24.8 ± 1.0%, p<0.001). In the time course experiments, an increase in amylase secretion was detected by 2.5 min and continued to increase steadily to a plateau at 40 min, with both submaximal (10-11M) and maximal (10-9M) CCK concentrations. The potent and specific CCK-receptor antagonist, l-364,718, caused a dose-dependent decrease in CCK-stimulated amylase secretion, with a half-maximal effect at 10-10M. The receptor antagonist, l-364,718, at 10-8M completely abolished CCK-stimulated amylase secretion. This microtechnique provides a simple, reliable, and reproducible method for the study of dispersed pancreatic acini. With this new methodology, the simultaneous study of 400-500 replicates is possible from a single guinea pig pancreas. Cells can be readily harvested at the end of the incubation period for measurement of intracellular messengers. With this technique, it is possible to carry out complex time course or dose response studies that should prove useful in the study of acinar cell function.
AB - Dispersed acini have proven to be particularly valuable in the study of pancreatic enzyme secretion. Complex time course studies or experiments requiring large numbers of replicates have proven difficult, however, with currently available techniques. Using a custom-designed incubation chamber, a miniaturized incubation method has been devised, which allows for continuous oxygenation of acini in 96-well microtiter plates and rapid separation of medium from acini by vacuum filtration. The filtrates from individual wells are collected into the wells of a second microtiter plate for pancreatic enzyme measurement. Using the above method, the dose response and time course of cholecystokinin (CCK-8)-stimulated amylase secretion was investigated. During a 1-h incubation, unstimulated amylase secretion was 4.1 ±0.3% of total acini content. Response to CCK was very sensitive, being detected at 10-13 M (p<0.05), half-maximal at 10-11 M CCK (14.0±0.6%, p<0.001) and maximal at 10-9M CCK (24.8 ± 1.0%, p<0.001). In the time course experiments, an increase in amylase secretion was detected by 2.5 min and continued to increase steadily to a plateau at 40 min, with both submaximal (10-11M) and maximal (10-9M) CCK concentrations. The potent and specific CCK-receptor antagonist, l-364,718, caused a dose-dependent decrease in CCK-stimulated amylase secretion, with a half-maximal effect at 10-10M. The receptor antagonist, l-364,718, at 10-8M completely abolished CCK-stimulated amylase secretion. This microtechnique provides a simple, reliable, and reproducible method for the study of dispersed pancreatic acini. With this new methodology, the simultaneous study of 400-500 replicates is possible from a single guinea pig pancreas. Cells can be readily harvested at the end of the incubation period for measurement of intracellular messengers. With this technique, it is possible to carry out complex time course or dose response studies that should prove useful in the study of acinar cell function.
KW - CCK
KW - Pancreatic acini
KW - cholecystokinin receptor antagonist
UR - http://www.scopus.com/inward/record.url?scp=0025281973&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0025281973&partnerID=8YFLogxK
U2 - 10.1007/BF02924344
DO - 10.1007/BF02924344
M3 - Review article
C2 - 1700032
AN - SCOPUS:0025281973
SN - 0169-4197
VL - 6
SP - 61
EP - 69
JO - International Journal of Pancreatology
JF - International Journal of Pancreatology
IS - 1
ER -