Stimulation of insulin release by glucose is widely thought to be coupled to a decrease in the activity of ATP-sensitive K + channels (K ATP channels) that is caused by a decreased concentration of free ADP. To date, most other investigators have reported only on total cellular ADP concentrations, even though only a small fraction of all ADP is free and only the free ADP affects K ATP channels. We tested the hypothesis that amino acids elicit insulin release via a decrease in the activity of K ATP channels owing to a decrease in the level of free ADP. We estimated the concentration of free ADP in ␤HC9 hyperplastic insulin-secreting cells based on the cell diameter and on luminometric measurements of ATP, phosphocreatine, and total creatine. The concentration of free ADP fell exponentially as the concentration of glucose increased. A physiological mixture of amino acids greatly stimulated insulin release at 0-30 mmol/l glucose but affected the concentration of free ADP only to a minor degree and significantly so only at ≤2 mmol/l glucose. In the presence of 2-deoxyglucose and NaN 3 , amino acids were unable to stimulate insulin release. When K ATP channels were held open with diazoxide (and the plasma membrane partially depolarized with high extracellular KCl), amino acids still stimulated insulin release. We conclude that amino acid-induced insulin release depends on two components: a yet-unknown amino acid sensor and K ATP channels, which serve to attenuate hormone release when cellular energy stores are low. We propose that glucoseinduced insulin release may be regulated similarly by two components: glucokinase and K ATP channels. Diabetes 50:291-300, 2001A ccording to the most widely accepted hypothesis, glucose induces insulin release as follows. Glucose rapidly equilibrates across the plasma membrane and is phosphorylated with the help of glucokinase, which determines flux through glycolysis (1). Pyruvate from glycolysis enters the citric acid cycle and determines ATP production from oxidative phosphorylation. As a result, the concentrations of ATP and free ADP reflect the concentration of blood glucose. The plasma membrane contains ATP-sensitive K + channels (K ATP channels). ATP inhibits the channels, and ADP relieves this inhibition (2). In the absence of glucose, K ATP channels are quite active and dictate the membrane potential. In the presence of glucose, initially, the balance between K + flux through K ATP channels and ionic fluxes through yet unknown "leak conductances" determines the membrane potential (2). Once the membrane potential is more positive than about -40 mV, Ca 2+ channels open intermittently, allowing the influx of extracellular Ca 2+ (2). As intracellular Ca 2+ rises, the exocytotic machinery is activated, which moves secretory granules containing insulin to the plasma membrane surface. In partial support of the above hypothesis, patients with a mutant glucokinase of abnormal affinity to glucose have an abnormal set point for glucose homeostasis (3-5), and other patients with K ATP c...

No abstract