Spatial Distribution of Ca2+ Signals during Repetitive Depolarizing Stimuli in Adrenal Chromaffin Cells

AbstractExocytosis in adrenal chromaffin cells is strongly influenced by the pattern of stimulation. To understand the dynamic and spatial properties of the underlying Ca2+ signal, we used pulsed laser Ca2+ imaging to capture Ca2+ gradients during stimulation by single and repetitive depolarizing stimuli. Short single pulses (10–100ms) lead to the development of submembrane Ca2+ gradients, as previously described (F. D. Marengo and J. R. Monck, 2000, Biophysical Journal, 79:1800–1820). Repetitive stimulation with trains of multiple pulses (50ms each, 2Hz) produce a pattern of intracellular Ca2+ increase that progressively changes from the typical Ca2+ gradient seen after a single pulse to a Ca2+ increase throughout the cell that peaks at values 3–4 times higher than the maximum values obtained at the end of single pulses. After seven or more pulses, the fluorescence increase was typically larger in the interior of the cell than in the submembrane region. The pattern of Ca2+ gradient was not modified by inhibitors of Ca2+-induced Ca2+ release (ryanodine), inhibitors of IP3-induced Ca2+ release (xestospongin), or treatments designed to deplete intracellular Ca2+ stores (thapsigargin). However, we found that the large fluorescence increase in the cell interior spatially colocalized with the nucleus. These results can be simulated using mathematical models of Ca2+ redistribution in which the nucleus takes up Ca2+ by active or passive transport mechanisms. These results show that chromaffin cells can respond to depolarizing stimuli with different dynamic Ca2+ signals in the submembrane space, the cytosol, and the nucleus