Amplitude modulation of Ca²⁺ signals induced by histamine in human endothelial cells

We have addressed the problem of whether the agonist concentration sensed by endothelial cells is encoded by the sustained rise of the intracellular Ca²⁺ concentration ([Ca²⁺]_i) or by the frequency of intracellular Ca²⁺ oscillations. Single or confluent endothelial cells from umbilical veins were stimulated for 15 min with histamine (0.03 to 100 μmol/l), and the concomitant changes in [Ca²⁺]_i were measured with fura-2/AM. Application of histamine at concentrations above 0.1 μmol/l resulted always in a fast spike of [Ca²⁺]_i, followed by a slow decline to a sustained plateau level, which depends on the presence of extracellular Ca²⁺. At the same time of the development of this plateau phase, quenching of the fura-2/AM signal occurred during agonist stimulation in a Ca²⁺-free, 1.5 mmol/l Mn²⁺ containing solution, indicating influx of divalent cations during this time. From 48 cells in 1.5 mmol/l [Ca²⁺]_e we obtained a close relation between histamine concentration and time integral of [Ca²⁺]_i taken over the 15 min recording of the plateau [Ca²⁺]_i. The half-maximal increase in the integral of [Ca²⁺]_i was at 0.7 μmol/l for solitary cells, 1.2 μmol/l for clustered cells and 1.2 μmol/l for the plateau Ca²⁺ level. Repetitive Ca²⁺ spikes or Ca²⁺ oscillations appeared only in 16 out of 48 cells, but their frequency was not correlated to the agonist concentration. Ca²⁺ oscillations were only observed in a concentration window between 0.1 and 1 μmol/l histamine, both in single and in clustered endothelial cells. Our results indicate that coding of the agonist concentration in endothelial cells is not related to the frequency of Ca²⁺ oscillations, but is closely correlated with the plateau level of intracellular Ca²⁺.