Effects of glucose, forskolin and tolbutamide on membrane potential and insulin secretion in the insulin-secreting cell line INS-1

Membrane voltages (V-m) of INS-1 cells, an insulin-secreting cell line. were measured mostly using the cell-attached mode of the patch-clamp method. The cell-attached configuration allowed the cell to be kept intact. Measurement of V-m was possible because seal resistances were very high and because the membrane obviously had a sufficiently high conductance (probably via K+ channels). Resting V-m was -80 +/- 1 mV (n = 42) and was mainly determined by sulphonylurea-sensitive K-ATP(+) channels since tolbutamide depolarized the plasma membrane in a concentration-dependent manner and generated action potential at 50 and 100 mu mol/l. D-Glucose, tested between 0.5 and 16.7 mmol/l, also depolarized the plasma membrane in a concentration-dependent manner and induced action potentials at concentrations higher than 5.6 mmol/l. Similarly, forskolin (5 mmol/l) depolarized the cells and increased the frequency of Ca2+-mediated action potentials. Insulin secretion was measured from cells growing in culture dishes, by radioimmunoassay. Glucose doubled secretion in INS-1 cells, whereas tolbutamide had no significant effect on secretion in the presence of 0.5 mmol/I and 16.7 mmol/l glucose At 3 mmol/l glucose, tolbutamide increased insulin release slightly. Forskolin elevated secretion twofold al a low glucose concentration. In contrast, when glucose or tolbutamide were added together with forskolin secretion was potentiated five- to tenfold. These results show that glucose induces membrane activation in INS-1 eels. Furthermore, the potent effect of tolbutamide, i.e. to depolarize the plasma membrane without inducing insulin release. leads to the conclusion that effects distal to depolarization are pivotal for secretion in INS-1 cells.