Pancreatic beta cells, tightly organized in the islet of Langerhans, secrete insulin in response to glucose in a calcium-dependent manner. The calcium input required for this secretory activity is thought to be provided by an oscillatory electrical activity occurring in the form of "bursts" of calcium action potentials. The previous observation that islet intracellular free Ca2+ levels undergo spontaneous oscillations in the presence of glucose, together with the fact that islet cells are coupled through gap junctions, hinted at a highly effective co-ordination between individual islet cells. Through the use of simultaneous recordings of intracellular calcium and membrane potential it is now reported that the islet calcium waves are synchronized with the beta cell bursting electrical activity. This observation suggests that each calcium wave is due to Ca2+ entering the cells during a depolarized phase of electrical activity. Moreover, fura-2 fluorescence image analysis indicates that calcium oscillations occur synchronously across the whole islet tissue. The maximal phase shift between oscillations occurring in different islet cells is estimated as 2 s. This highly co-ordinated oscillatory calcium signalling system may underlie pulsatile insulin secretion and the islet behaviour as a secretory "syncytium". Since increasing glucose concentration lengthens calcium wave and burst duration without significantly affecting wave amplitude, we further propose that it is the fractional time at an enhanced Ca2+ level, rather than its amplitude, that encodes for the primary response of insulin-secreting cells to fuel secretagogues.

Intracellular Ca2+ levels were monitored in single, acutely isolated mouse islets of Langerhans by dual emission Indo-l fluorometry. High-frequency (3.1 min-i) [Ca*+], oscillations with a brief rising time (1-2 s) and 10 s half-width ('fast' oscillations) were detected in 11 mM glucose. Raising the glucose concentration to 16.7 mM increased the duration of these oscillations, which were otherwise absent in 5.5 mM glucose. [Caz'], waves of lower frequency (0.5 mini) and longer rising time ('slow' oscillations) were also recorded. The data indicate that "fast" oscillations are directly related to p-cell bursting electrical activity, and suggest the existence of extensive networks of electrically coupled cells in the islet.

Glucose‐induced insulin release from single islets of Langerhans is pulsatile. We have investigated the correlation between changes in cytosolic free calcium concentration ([Ca2+]i) and oscillatory insulin secretion from single mouse islets, in particular examining the basis for differences in secretory responses to intermediate and high glucose concentrations. Insulin release was monitored in real time through the amperometric detection of the surrogate insulin marker 5‐hydroxytryptamine (5‐HT) via carbon fibre microelectrodes. The [Ca2+]i was simultaneously recorded by whole‐islet fura‐2 microfluorometry. In 82 % of the experiments, exposure to 11 mM glucose evoked regular high‐frequency (average, 3.4 min−1) synchronous oscillations in amperometric current and [Ca2+]i. In the remaining experiments (18 %), 11 mM glucose induced an oscillatory pattern consisting of high‐frequency [Ca2+]i oscillations that were superimposed on low‐frequency (average, 0.32 min−1) [Ca2+]i waves. Intermittent high‐frequency [Ca2+]i oscillations gave rise to a similar pattern of pulsatile 5‐HT release. Raising the glucose concentration from 11 to 20 mM increased the duration of the steady‐state [Ca2+]i oscillations without increasing their amplitude. In contrast, both the duration and amplitude of the associated 5‐HT transients were increased by glucose stimulation. The amount of 5‐HT released per secretion cycle was linearly related to the duration of the underlying [Ca2+]i oscillations in both 11 and 20 mM glucose. The slopes of the straight lines were identical, indicating that there is no significant difference between the ability of calcium oscillations to elicit 5‐HT/insulin release in 11 and 20 mM glucose. In situ 5‐HT microamperometry has the potential to resolve the high‐frequency oscillatory component of the second phase of glucose‐induced insulin secretion. This component appears to reflect primarily the duration of the underlying [Ca2+]i oscillations, suggesting that glucose metabolism and/or access to glucose metabolites is not rate limiting to fast pulsatile insulin release.

Although isolated rat islets are widely used to study in vitro insulin secretion and the underlying metabolic and ionic processes, knowledge on the properties of glucose-induced electrical activity (GIEA), a key step in glucose-response coupling, has been gathered almost exclusively from microdissected mouse islets. Using a modified intracellular recording technique, we have now compared the patterns of GIEA in collagenase-isolated rat and mouse islets. Resting membrane potentials of rat and mouse ␤-cells were approximately -50 and -60 mV, respectively. Both rat and mouse ␤-cells displayed prompt membrane depolarizations in response to glucose. However, whereas the latter exhibited a bursting pattern consisting of alternating hyperpolarized and depolarized active phases, rat ␤-cells fired action potentials from a nonoscillating membrane potential at all glucose concentrations (8.4-22.0 mmol/l). This was mirrored by changes in the intracellular Ca 2+ concentration ([Ca 2+ ] i ), which was oscillatory in mouse and nonoscillatory in rat islets. Stimulated rat ␤-cells were strongly hyperpolarized by diazoxide, an activator of ATP-dependent K + channels. Glucose evoked dosedependent depolarizations and [Ca 2+ ] i increases in both rat (EC 50 5.9-6.9 mmol/l) and mouse islets (EC 50 8.3-9.5 mmol/l), although it did not affect the burst plateau potential in the latter case. We conclude that there are important differences between ␤-cells from both species with respect to early steps in the stimulussecretion coupling cascade based on the following findings: 1) mouse ␤-cells have a larger resting K + conductance in 2 mmol/l glucose, 2) rat ␤-cells lack the compensatory mechanism responsible for generating membrane potential oscillations and holding the depolarized plateau potential in mouse ␤-cells, and 3) the electrical and [Ca 2+ ] i dose-response curves in rat ␤-cells are shifted toward lower glucose concentrations. Exploring the molecular basis of these differences may clarify several a priori assumptions on the electrophysiological properties of rat ␤-cells, which could foster the development of new working models of pancreatic ␤-cell function.

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