Effects of leucine on insulin secretion and beta cell membrane potential in mouse islets of Langerhans.

Leucine is known to enhance insulin secretion from islets of Langerhans, and insulin promotes leucine uptake in peripheral tissues. The present studies were designed to elucidate the effects of leucine on glucose responsiveness and stimulus secretion coupling in mouse islets of Langerhans. The effects of 20 mM leucine on insulin secretion and membrane potential were studied over a range of glucose concentrations (0-27.7 mM). Microdissected, perifused pancreatic islets from normal adult mice were used for both studies of insulin secretion and electrophysiology in order to make a close comparison between these measurements. Leucine enhanced the insulin secretion in the presence of 5.6, 11.1, and 22.2 mM glucose. In the presence of leucine, 27 mM glucose inhibited insulin secretion. In the absence of glucose-leucine did not induce electrical activity of the beta cell membrane, whereas in the presence of 5.6, 11.1, and 22.2 mM glucose leucine increased spike frequency. Thus, leucine shifts both the glucose-dependent insulin secretion and electrical activity toward lower glucose concentrations. It is concluded that leucine and glucose share a common metabolic pathway (citric acid cycle) for stimulatory effects. Leucine is deaminated to form 2-ketoisocaproic acid (KIC) and produce NH4+. We propose that in the absence of glucose this increases cytosolic pH, which in turn increases K+ permeability, and inhibits electrical activity and insulin secretion.

Differences in K+ permeability between cultured adult and neonatal rat islets of Langerhans in response to glucose, tolbutamide, diazoxide, and theophylline.

The effects of glucose, tolbutamide, and diazoxide on K+ permeability in neonatal and adult rat pancreatic islets, maintained in culture 1 week, were investigated by measuring the 86Rb outflow rate from prelabeled islets. In the absence of glucose, the 86Rb efflux was significantly lower in neonatal than adult islets. Raising the glucose concentration to 2.8, 5.6, 8.3, and 11.1 mM produced a marked reduction in the 86Rb efflux in adult islets but only a minor reduction in neonatal islets. The effect of tolbutamide to reduce, and diazoxide to increase, the 86Rb efflux was also less in neonatal islets. These results are discussed with respect to previously reported differences in insulin secretion from neonatal and adult islets in culture.

Valinomycin inhibition of the electrical activity of mouse pancreatic beta-cells.

The effect of the K-ionophore valinomycin (VAL) on the electrical activity of single mouse pancreatic beta-cells was measured using the glass micro-electrode technique. In the presence of 11.1 mM glucose, after 4 min of exposure to 10 nM VAL the spike activity was abolished, bursts became irregular and after 10 min the membrane hyperpolarized 4 mV. The electrical activity was completely blocked by 100 nM VAL within 3 min and the membrane hyperpolarization averaged 10 mV. The effect of VAL was irreversible. VAL (100 nM) also inhibited the electrical activity induced by 11.1 mM glucose plus 10 mM tetraethylammonium chloride (TEA), a specific blocker of the voltage sensitive PK. Total inhibition was observed 6-7 min after VAL addition and the cell hyperpolarized 14 mV. With the simultaneous application of 10 mM TEA and 100 microM quinine, a specific blocker of the [Ca2+]i-dependent PK, in the presence of 16.7 mM glucose, the membrane showed continuous activity and progressive depolarization from -45 to -5 mV within 23 min. The subsequent addition of 100 nM VAL also hyperpolarized the cell, with the membrane potential reaching -17 mV after 7 min. On the basis of these data, we suggest that VAL affects the glucose-dependent depolarization of the beta-cell membrane by increasing K+ permeability.

Effect of glycemic index on whole-body substrate oxidation in obese women.

BACKGROUND: Glycemic index is hypothesized to determine fuel partitioning through serum plasma insulin modifications induced by dietary carbohydrates, thereby modulating fat accretion or oxidation. OBJECTIVE: To assess the glycemic effects on postprandial fuel oxidation and blood response. DESIGN: In all, 12 obese women were fed on a randomized crossover design with two test meals (breakfast+lunch). High- or low-glycemic meals were provided on separate days. Energy intake on high-glycemic meal was 7758+/-148 kJ and for low-glycemic meal was 7806+/-179 kJ. Carbohydrates supplied were 273+/-5 and 275+/-6 g, respectively. Macronutrient distribution was 55% carbohydrates, 30% fat and 15% protein. Fuel oxidation was measured continuously in a respiratory chamber for 10 h. Serum glucose, free fatty acids (FFA), insulin and glucagon samples were taken for 5 h after breakfast. RESULTS: Glucose AUC changed significantly in response to different glycemic breakfast. Low- vs high-glycemic breakfast was 211+/-84 and 379+/-164 mmol/l (P<0.05). Similarly, insulin changed from 94+/-37 and 170+/-87 nmol/l (P<0.05), respectively. The rate of increment for serum glucose and insulin reached by the high- vs low-glycemic meal was 1.8 times more with the high-glycemic breakfast. Serum FFA were similarly suppressed by both meal types by 3 h after meal intake, but then raised significantly more with the low-glycemic meal by the fourth and fifth hour (P<0.05). Plasma glucagon did not show a significant variation with glycemic index. Carbohydrate and fat oxidation was not modified by glycemic meal characteristics, being virtually the same for low- vs high-glycemic comparisons in the 5 h following breakfast and lunch (P=NS). CONCLUSION: This study demonstrates that dietary glycemic characteristics were unable to modify fuel partitioning in sedentary obese women.

The kinetics of electrical activity of beta cells in response to a "square wave" stimulation with glucose or glibenclamide.

The effect of a "square wave" stimulation with glucose or glibenclamide on the electrical activity of beta-cells has been studied with microelectrode techniques in isolated perifused mouse islets. While glucose evoked a burst activity with periodic oscillations of the membrane potential between two levels, glibenclamide produced a constant depolarization with a continuous spike activity. In both cases the time course of activity was biphasic and resembled closely the corresponding patterns of insulin release. The effect of glibenclamide on the membrane was irreversible. The results provide further evidence for a direct correlation between insulin release and electrical activity. Furthermore, it is suggested that the membrane potential is, at least in part, involved in the mechanism of biphasic insulin release.