Local induction of pacemaking activity in a monolayer of electrically coupled quiescent NRK fibroblasts

SummaryCultures of normal rat kidney (NRK) fibroblasts may display spontaneous calcium action potentials which propagate throughout the cellular monolayer. Pacemaking activity of NRK cells was studied by patch clamp electrophysiology and vital calcium imaging, using a new experimental approach in which a ring was placed on the monolayer in order to physically separate pacemakers within or under the ring and follower cells outside the ring. Stimulation of cells inside the ring with IP3-generating hormones such as prostaglandin F2α (PGF2α) resulted in the induction of periodic action potentials outside the ring, which were abolished when the L-type calcium channel blocker nifedipine was added outside the ring, but not inside the ring. PGF2α-treated cells displayed asynchronous IP3-mediated calcium oscillations of variable frequency, while follower cells outside the ring showed synchronous calcium transients which coincided with the propagating action potential. Mathematical modelling indicated that addition of PGF2α inside the ring induced both a membrane potential gradient and an intracellular IP3 gradient, both of which are essential for the induction of pacemaking activity under the ring. These data show that intercellular coupling between PGF2α-treated and non-treated cells is essential for the generation of a functional pacemaker area whereby synchronization of calcium oscillations occurs by activation of L-type calcium channels.