The parallel gap junction electrical conductance between a β-cell and its nearest neighbors was measured by using an intracellular microelectrode to clamp the voltage of a β-cell within a bursting islet of Langerhans. The holding current records consisted of bursts of inward current due to the synchronized oscillations in membrane potential of the surrounding cells. The membrane potential record of the impaled cell, obtained in current clamp mode, was used to estimate the behavior of the surrounding cells during voltage clamp, and the coupling conductance was calculated by dividing the magnitude of the current bursts by that of the voltage bursts. The histogram of coupling conductance magnitude from 26 cells was bimodal with peaks at 2.5 and 3.5 nS, indicating heterogeneity in extent of electrical communication within the islet of Langerhans. Gap junction conductance reversibly decreased when the temperature was lowered from 37 to 30°C and when the extracellular calcium concentration was raised from 2.56 to 7.56 mm. The coupling conductance decreased slightly during the active phase of the burst. Activation of adenylate cyclase with forskolin (10 μm) resulted in an increase in cell-to-cell electrical coupling. We conclude that β-cell gap junction conductance can be measured in situ under near physiological conditions. Furthermore, the magnitude and physiological regulation of β-cell gap junction conductance suggest that intercellular electrical communication plays an important role in the function of the endocrine pancreas.