Prostaglandin F2α induces unsynchronized intracellular calcium oscillations in monolayers of gap junctionally coupled NRK fibroblasts

Abstract

We investigated the intracellular calcium oscillations induced by prostaglandin F2alpha (PGF2alpha) in individual cells of confluent, gap junction-coupled monolayers of normal rat kidney (NRK) fibroblasts. PGF2alpha (1000 nM) induced oscillations in more than 90% of the cells in the monolayer, but the frequency of these oscillations was highly variable between individual cells (0.2-1.4 min(-1)). The initial calcium peak resulted from calcium release from IP3-sensitive stores, while subsequent calcium transients were mediated by interplay between both IP3-sensitive calcium stores and calcium influx. The oscillation frequency was increased by sensitizing the IP3 receptor with thimerosal (10 microM) and depended on the extracellular calcium concentration. Thapsigargin (5 nM), which inhibits reuptake of calcium into the stores, only seemed to reduce the amplitude of the oscillation. Patch-clamp experiments revealed that PGF2alpha did not inhibit electrical coupling of the NRK cells in the monolayer. Gap junctional permeability of NRK cells thus appears to be sufficient to allow electrical coupling, resulting in a uniform membrane potential throughout the entire monolayer, but insufficient to synchronize the intracellular calcium oscillations upon PGF2alpha stimulation.

Autocrine production of prostaglandin F2α enhances phenotypic transformation of normal rat kidney fibroblasts

“…Repetitive addition of either the CM of transformed NRK cells or TransX showed no homologous desensitization, in agreement with the finding that PGF 2␣ only partially desensitizes its natural G protein-coupled receptor (10). The binding of PGF 2␣ to this FP receptor previously was shown to result in increased intracellular IP 3 and Ca 2ϩ levels (23), and we recently demonstrated that PGF 2␣ can induce intracellular Ca 2ϩ oscillations in NRK cells cultured in quiescent monolayers (13). These oscillations resulted from an intricate interplay between Ca 2ϩ release from internal stores and Ca 2ϩ influx over the plasma membrane and were not synchronized in spite of gap junctional coupling of the NRK cells in such monolayers.…”

“…The fragmentation peaks resulted from the sequential losses of water (from m/z 377.3 to m/z 359.3 to m/z 341.3) and both water and carboxyl (from m/z 377.3 to m/z 315.3) from the parent compound. These results demonstrate that TransX contains PGF 2␣ and that the depolarizing activity of TransX can be ascribed to this prostaglandin (13).…”

“…We previously found that the minimal PGF 2␣ concentration required to constitutively depolarize monolayers of quiescent NRK cells is ϳ10 nM (13), and therefore the concentration present in the CM of transformed NRK cells was sufficient to explain the depolarizing activity of this medium. Repetitive addition of either the CM of transformed NRK cells or TransX showed no homologous desensitization, in agreement with the finding that PGF 2␣ only partially desensitizes its natural G protein-coupled receptor (10).…”

“…By performing dynamic video imaging measurements of fura-2-loaded, quiescent NRK cells, we investigated whether TransX had effects on intracellular Ca 2ϩ levels similar to those of PGF 2␣ . This prostaglandin induces unsynchronized intracellular Ca 2ϩ oscillations in NRK cells cultured in quiescent monolayers (13). Figure 4A shows the typical Ca 2ϩ response of an individual NRK cell in a monolayer upon exposure to TransX.…”

Myofibroblast communication is controlled by intercellular mechanical coupling

“…[i] oscillations have been shown to be synchronised between fibroblasts that intrinsically oscillate with similar frequencies (Harks et al, 2003).…”