Effects of glucose, forskolin and tolbutamide on membrane potential and insulin secretion in the insulin-secreting cell line INS-1.
Membrane voltages (Vm) 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 Vm was possible because seal resistances were very high and because the membrane obviously had a sufficiently high conductance (probably via K+ channels). Resting Vm 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 potentials at 50 and 100 micromol/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 micromol/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/l and 16. 7 mmol/l glucose. At 3 mmol/l glucose, tolbutamide increased insulin release slightly. Forskolin elevated secretion twofold at 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 cells. 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.
Ullrich
S
S
Physiologisches Institut der Albert-Ludwigs Universität, Abteilung II, Hermann-Herder-Strasse 7, D-79104 Freiburg, Germany.
Abel
K B
KB
Lehr
S
S
Greger
R
R
D000200:Action Potentials / Q000187:drug effects; D000818:Animals; D002460:Cell Line; D005576:Colforsin / Q000008:administration & dosage / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D005947:Glucose / Q000008:administration & dosage / Q000494:pharmacology*; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000187:drug effects / Q000502:physiology*; D008564:Membrane Potentials / Q000187:drug effects; D051381:Rats; D014044:Tolbutamide / Q000008:administration & dosage / Q000494:pharmacology*
D007328:Insulin; D005576:Colforsin; D014044:Tolbutamide; D005947:Glucose