Functional and morphological characteristics of isolated pancreatic islets in tissue culture.

Islets were isolated from the pancreas of female rats by using the collagenase technique. After culturing for 4 days, the islets were taken for measurement of insulin release biosynthesis as well as glucose utilization in subsequent short-time incubations. A low glucose concentration was insufficient to maintain a glucose-stimulated insulin release in vitro. A high glucose concentration had a protecting and restoring effect on the insulin release: ultrastructurally, such islets showed signs of an active biosynthesis in the electron micrograph. The enhancement of Mg++ in the culture medium resulted in an improvement of insulin storage in the islets, accompanied by a well-preserved action of glucose in a subsequent incubation.

Influence of culture conditions on B-cell functions in vitro.

Isolated pancreatic islets from Wistar rats were maintained in vitro at 37 degrees C for 12 days in TCM 199 containing 10 mmol/l glucose and supplemented with 10% serum. 10 mmol/l glucose completely prevented the diminution of insulin content independent of the type of serum (fetal calf serum; serum from newborn calfs; horse serum), whereas the insulin biosynthesis as well as the insulin release were modulated by the kind of serum. Fetal calf serum can be replaced by horse serum.

Effect of glucose on human fetal pancreatic tissue in vitro.

For investigating insulin secretion in vitro human fetal pancreatic slices prepared from women with a normal carbohydrate tolerance at a mean gestational age of 12 +/- 1 weeks were incubated in the presence of various secretagogues. Glucose concentration up to 20 mmol/l failed to enhance insulin secretion, whereas an increase of glucose up to 40 mmol/l, the addition of the phosphodiesterase inhibitor IBMX or a priming period of 30 min in the presence of 20 mmol/l glucose resulted in an enhancement of hormone release, calculated per microgram dry weight. For the first time the results demonstrates an intact secretory machinery of the B-cells under controlled conditions in an early stage of gestational development.

The influence of cultivation on (pro-)insulin biosynthesis and secretion of isolated pancreatic islets of C57BL-mice, long-term treated with glibenclamide in vivo.

Long-term treatment of C57Bl/KsJ and C57Bl/6J mice with glibenclamide in vivo caused a diminished response of insulin to high glucose concentrations (20 mmol/l). Islets of those mice were investigated to answer the question whether it is possible to overcome this diminution of glucose sensitivity by cultivation in presence of high glucose (20 mmol/l). The insulin release of islets of glibenclamide treated mice was significantly lowered also in the first 48 h of cultivation. In the following short-term incubation (2 h) no differences in the glucose stimulated insulin secretion could be seen. The second cultivation period (48-96 h) confirmed these results. Both groups of islets (controls and glibenclamide treated) reached comparable values in insulin release. Cultivation of islets of glibenclamide treated mice in presence of 20 mmol/l glucose for at least 2 days led to a restoration of the glucose sensitivity of insulin release. Insulin biosynthesis, judged by measuring 3H-leucine incorporation into (pro-)insulin, was largely unaffected. In all experimental conditions the insulin content was comparable to that of controls.