Agonist-induced cytosolic calcium oscillations originate from a specific locus in single hepatocytes

Digital imaging fluorescence microscopy of fura-2-loaded hepatocytes in primary culture has been used to examine the changes of cytosolic free Ca²⁺ ([Ca²⁺](i)) in response to receptor activation by α₁-adrenergic agonists and vasopressin at the subcellular level. Agonist-induced Ca²⁺ oscillations did not occur synchronously within the cell but originated from a specific region adjacent to the cell membrane and then propagated throughout the rest of the cell, with each oscillation within a series originating from the same locus. Furthermore, hormones acting through different receptors produced Ca²⁺ waves with similar rates of progress (20-25 μm·s^-1) which originated from the same subcellular locus. For a given cell, the rate of progress and amplitude of the Ca²⁺ waves were independent of applied agonist concentration and were unaffected by depletion of extracellular Ca²⁺. The kinetics of Ca²⁺ increase at different points within the cell indicated that the Ca²⁺ waves were not driven by diffusion but were characteristic of a self-propagating mechanism. Significantly, when cells were treated with AIF₄^- to directly activate the G-protein which couples receptor occupancy to [Ca²⁺](i) mobilization, the origin and kinetics of the Ca²⁺ waves were identical to those observed with hormonal stimulation. It is proposed that the spatial organization of the intracellular Ca²⁺ release mechanisms may have significance in the regulation of the asymmetric metabolic functions of hepatocytes and other functionally polarized cells.