Effect of synthetic trypsin inhibitor on plasma immunoreactive cholecystokinin in rats

Abstract

A sensitive and specific radioimmunoassay system for rat plasma cholecystokinin (CCK) was employed to study the effect of trypsin inhibitor on plasma CCK levels. Feeding the trypsin inhibitor, FOY-305 200 mg/kg, to rats for 10 days stimulated the pancreatic growth. However, there was no significant difference in fasting plasma concentrations and duodenal contents of CCK and secretin. On the other hand, acute ingestion of same dosage of FOY-305 caused a marked (20-fold) and sustained elevation of plasma CCK levels. The principal form of circulating CCK was CCK-22 like on Sephadex G-50, in contrast to that of the cerebrum and duodenum where CCK-8 like form, or CCK-8 and CCK-22 like forms were main components, respectively. A significant rise in plasma levels of immunoreactive secretin was also found. These results suggest that the pancreatic growth observed by feeding the trypsin inhibitor to rats is caused by an excessive amount of CCK in conjunction with secretin.

CCK-58 is the only detectable endocrine form of cholecystokinin in rat

“…A new method of collecting and processing blood has been developed that yields similar recovery (80%) for all molecular forms of cholecystokinin that have been identified in blood (8,11,13,14) or intestinal extracts (3,25) of rat. Furthermore, unlike previous methods used to determine the endocrine form of cholecystokinin in rat (8,11,13,14), this new optimized method prevented degradation of radiolabeled CCK-8 or CCK-58.…”

“…These labels were also used as internal markers of recovery in all determinations of the molecular forms of cholecystokinin in fasted and stimulated rat blood and as markers to decide which fractions from SepPak should be pooled. Additionally, 125 I-Tyr 20 , 125 I-Tyr 52 -labeled CCK-58 was used to compare degradation of peptides by the method developed in this report to a method that simulates prior studies on the molecular forms of cholecystokinin in rats (8,11,13,14).…”

“…Previous reports on the molecular forms of cholecystokinin in rat blood involved collecting blood (usually trunk blood) into heparinized or EDTA tubes and forming plasma (8,11,13,14). Freely fed Sprague-Dawley rats (275-325 g) were anesthetized with pentobarbital sodium (10 mg/kg).…”

“…One study in rats in which cholecystokinin release into blood was stimulated by diversion of bile and pancreatic juice reported that CCK-22 and CCK-8 were the major forms in blood, although CCK-33/39 was the major form stored in the small intestine (8). In another study, in which cholecystokinin release was stimulated with a trypsin inhibitor, the major circulating form reported was CCK-22 (the same predominant form observed in their small intestine extracts) (11). In another work (14), the blood of fasting rats contained nearly equal amounts of CCK-8 and CCK-33, whereas rats stimulated by ad libitum feeding or suckling contained a greater proportion of CCK-8.…”

“…We present a second method different from the acidic buffer method to suggest a possible explanation as to why other investigators see smaller molecular forms of cholecystokinin than we observed. This method formed plasma directly after collecting blood, as done in earlier studies (8,11,13,14) on the molecular forms of cholecystokinin in rat blood. Our sample handling (incubation on ice for 1 h before freezing plasma and thawing plasma for 1 h) may not exactly duplicate the way samples were processed in earlier studies.…”