A Kir6.2 mutation causing neonatal diabetes impairs electrical activity and insulin secretion from INS-1 beta-cells.
ATP-sensitive K(+) channels (K(ATP) channels) couple beta-cell metabolism to electrical activity and thereby play an essential role in the control of insulin secretion. Gain-of-function mutations in Kir6.2 (KCNJ11), the pore-forming subunit of this channel, cause neonatal diabetes. We investigated the effect of the most common neonatal diabetes mutation (R201H) on beta-cell electrical activity and insulin secretion by stable transfection in the INS-1 cell line. Expression was regulated by placing the gene under the control of a tetracycline promoter. Transfection with wild-type Kir6.2 had no effect on the ATP sensitivity of the K(ATP) channel, whole-cell K(ATP) current magnitude, or insulin secretion. However, induction of Kir6.2-R201H expression strongly reduced K(ATP) channel ATP sensitivity (the half-maximal inhibitory concentration increased from approximately 20 mumol/l to approximately 2 mmol/l), and the metabolic substrate methyl succinate failed to close K(ATP) channels or stimulate electrical activity and insulin secretion. Thus, these results directly demonstrate that Kir6.2 mutations prevent electrical activity and insulin release from INS-1 cells by increasing the K(ATP) current and hyperpolarizing the beta-cell membrane. This is consistent with the ability of the R201H mutation to cause neonatal diabetes in patients. The relationship between K(ATP) current and the membrane potential reveals that very small changes in current amplitude are sufficient to prevent hormone secretion.
Tarasov
Andrei I
AI
University Laboratory of Physiology, Parks Road, Oxford, OX1 3PT, UK.
Welters
Hannah J
HJ
Senkel
Sabine
S
Ryffel
Gerhart U
GU
Hattersley
Andrew T
AT
Morgan
Noel G
NG
Ashcroft
Frances M
FM
D000255:Adenosine Triphosphate / Q000378:metabolism / Q000494:pharmacology; D019943:Amino Acid Substitution; D002460:Cell Line; D003922:Diabetes Mellitus, Type 1 / Q000235:genetics* / Q000276:immunology; D005786:Gene Expression Regulation; D006801:Humans; D007231:Infant, Newborn; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000378:metabolism*; D007700:Kinetics; D009154:Mutation; D024661:Potassium Channels, Inwardly Rectifying / Q000187:drug effects / Q000235:genetics*; D014162:Transfection; D000099161:Kcnj11 Channel
D000255:Adenosine Triphosphate; D007328:Insulin; D024661:Potassium Channels, Inwardly Rectifying; D000099161:Kcnj11 Channel