CA2-MOBILIZING HORMONES INDUCE SEQUENTIALLY ORDERED CA2+ SIGNALS IN MULTICELLULAR SYSTEMS OF RAT HEPATOCYTES()

The development of hormone-mediated Ca2+ signals was analysed in polar ized doublets, triplets and quadruplets of rat hepatocytes by video im aging of fura2 fluorescence. These multicellular models showed dilated bile canaliculi, and gap junctions were observed by using an anti-con nexin-32 antibody. They also showed highly organized Ca2+ signals in r esponse to vasopressin or noradrenaline. Surprisingly, the primary ris es in intracellular Ca2+ concentration ([Ca2+](i)) did not start rando mly from any cell of the multiplet. It originated invariably in the sa me hepatocyte (first-responding cell), and then was propagated in a se quential manner to the nearest connected cells (cell 2, then 3, in tri plets; cell 2, 3, then 4 in quadruplets). The sequential activation of the cells appeared to be an intrinsic property of multiplets of rat h epatocytes. (1) In the continued presence of hormones, the same sequen tial order was observed up to six times, i.e. at each train of oscilla tions occurring between the cells. (2) The order of [Ca2+](i) response s was modified neither by the repeated addition of hormones nor by the hormonal dose. (3) The mechanical disruption of an intermediate cell slowed down the speed of the propagation, suggesting a role of gap jun ctions in the rapidity of the sequential activation of cells. (4) The same multiplet could have a different first-responding cell for vasopr essin or noradrenaline, suggesting a role of the hormonal receptors in the sequentiality of cell responses. It is postulated that a function al heterogeneity of hormonal receptors, and the presence of functional gap junctions, are involved in the existence of sequentially ordered hormone-mediated [Ca2+](i) rises in the multiplets of rat hepatocytes.