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ATP- and gap junction-dependent intercellular calcium signaling in osteoblastic cells.

Jorgensen NR, Geist ST, Civitelli R, Steinberg TH - J. Cell Biol. (1997)

Bottom Line: ROS 17/2.8 cells, which express the gap junction protein connexin43 (Cx43), are well dye coupled, and lack P2U receptors, transmitted slow gap junction-dependent calcium waves that did not require release of intracellular calcium stores.These studies demonstrate that activation of P2U purinergic receptors can propagate intercellular calcium, and describe a novel Cx43-dependent mechanism for calcium wave propagation that does not require release of intracellular calcium stores by IP3.These studies suggest that gap junction communication mediated by either Cx43 or Cx45 does not allow passage of IP3 well enough to elicit release of intracellular calcium stores in neighboring cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine, Washington University School of Medicine, St. Louis, Missouri 63110, USA.

ABSTRACT
Many cells coordinate their activities by transmitting rises in intracellular calcium from cell to cell. In nonexcitable cells, there are currently two models for intercellular calcium wave propagation, both of which involve release of inositol trisphosphate (IP3)- sensitive intracellular calcium stores. In one model, IP3 traverses gap junctions and initiates the release of intracellular calcium stores in neighboring cells. Alternatively, calcium waves may be mediated not by gap junctional communication, but rather by autocrine activity of secreted ATP on P2 purinergic receptors. We studied mechanically induced calcium waves in two rat osteosarcoma cell lines that differ in the gap junction proteins they express, in their ability to pass microinjected dye from cell to cell, and in their expression of P2Y2 (P2U) purinergic receptors. ROS 17/2.8 cells, which express the gap junction protein connexin43 (Cx43), are well dye coupled, and lack P2U receptors, transmitted slow gap junction-dependent calcium waves that did not require release of intracellular calcium stores. UMR 106-01 cells predominantly express the gap junction protein connexin 45 (Cx45), are poorly dye coupled, and express P2U receptors; they propagated fast calcium waves that required release of intracellular calcium stores and activation of P2U purinergic receptors, but not gap junctional communication. ROS/P2U transfectants and UMR/Cx43 transfectants expressed both types of calcium waves. Gap junction-independent, ATP-dependent intercellular calcium waves were also seen in hamster tracheal epithelia cells. These studies demonstrate that activation of P2U purinergic receptors can propagate intercellular calcium, and describe a novel Cx43-dependent mechanism for calcium wave propagation that does not require release of intracellular calcium stores by IP3. These studies suggest that gap junction communication mediated by either Cx43 or Cx45 does not allow passage of IP3 well enough to elicit release of intracellular calcium stores in neighboring cells.

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ROS cells propagate slow intercellular calcium waves. Monolayers of  ROS cells were loaded with  fura-2, and a single cell (arrowhead) was mechanically  stimulated during fluorescence ratio imaging. Time after stimulation in seconds is  indicated on each panel. The  pseudocolor map represents  the estimated [Ca2+]i.
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Figure 2: ROS cells propagate slow intercellular calcium waves. Monolayers of ROS cells were loaded with fura-2, and a single cell (arrowhead) was mechanically stimulated during fluorescence ratio imaging. Time after stimulation in seconds is indicated on each panel. The pseudocolor map represents the estimated [Ca2+]i.

Mentions: In both ROS and UMR cell lines, mechanical stimulation increased [Ca2+]i in the stimulated cell and subsequently induced a wave of calcium transients spreading to adjacent cells in the monolayer. However, intercellular calcium waves in ROS monolayers were much slower, and propagated to fewer cells, than calcium waves in UMR monolayers. In monolayers of ROS cells, calcium waves spread 5–15 neighboring cells over several minutes (Fig. 2). Although the rate of propagation of these waves showed variability depending on cell density and geometry, these “slow” waves in ROS cells generally propagated at a rate of ∼0.5 μm/s. Frequently there was a time lag of 15–30 s between the calcium rise in one cell and its neighbor. In contrast, the poorly dye-coupled UMR cells transmitted waves to most of the cells in the field of view (30–50 cells) within 15–20 s (Fig. 3), and with little time lag between neighboring cells. The velocity of propagation was ∼10 μm/s in UMR. In some experiments, the stimulated cell was in fact ruptured, as demonstrated by loss of dye from that cell. We detected no differences in the propagated calcium wave between experiments where cells were ruptured and experiments where cells were not ruptured. In either instance, ROS cells demonstrated slower calcium waves and UMR cells demonstrated faster calcium waves.


ATP- and gap junction-dependent intercellular calcium signaling in osteoblastic cells.

Jorgensen NR, Geist ST, Civitelli R, Steinberg TH - J. Cell Biol. (1997)

ROS cells propagate slow intercellular calcium waves. Monolayers of  ROS cells were loaded with  fura-2, and a single cell (arrowhead) was mechanically  stimulated during fluorescence ratio imaging. Time after stimulation in seconds is  indicated on each panel. The  pseudocolor map represents  the estimated [Ca2+]i.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2139805&req=5

Figure 2: ROS cells propagate slow intercellular calcium waves. Monolayers of ROS cells were loaded with fura-2, and a single cell (arrowhead) was mechanically stimulated during fluorescence ratio imaging. Time after stimulation in seconds is indicated on each panel. The pseudocolor map represents the estimated [Ca2+]i.
Mentions: In both ROS and UMR cell lines, mechanical stimulation increased [Ca2+]i in the stimulated cell and subsequently induced a wave of calcium transients spreading to adjacent cells in the monolayer. However, intercellular calcium waves in ROS monolayers were much slower, and propagated to fewer cells, than calcium waves in UMR monolayers. In monolayers of ROS cells, calcium waves spread 5–15 neighboring cells over several minutes (Fig. 2). Although the rate of propagation of these waves showed variability depending on cell density and geometry, these “slow” waves in ROS cells generally propagated at a rate of ∼0.5 μm/s. Frequently there was a time lag of 15–30 s between the calcium rise in one cell and its neighbor. In contrast, the poorly dye-coupled UMR cells transmitted waves to most of the cells in the field of view (30–50 cells) within 15–20 s (Fig. 3), and with little time lag between neighboring cells. The velocity of propagation was ∼10 μm/s in UMR. In some experiments, the stimulated cell was in fact ruptured, as demonstrated by loss of dye from that cell. We detected no differences in the propagated calcium wave between experiments where cells were ruptured and experiments where cells were not ruptured. In either instance, ROS cells demonstrated slower calcium waves and UMR cells demonstrated faster calcium waves.

Bottom Line: ROS 17/2.8 cells, which express the gap junction protein connexin43 (Cx43), are well dye coupled, and lack P2U receptors, transmitted slow gap junction-dependent calcium waves that did not require release of intracellular calcium stores.These studies demonstrate that activation of P2U purinergic receptors can propagate intercellular calcium, and describe a novel Cx43-dependent mechanism for calcium wave propagation that does not require release of intracellular calcium stores by IP3.These studies suggest that gap junction communication mediated by either Cx43 or Cx45 does not allow passage of IP3 well enough to elicit release of intracellular calcium stores in neighboring cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine, Washington University School of Medicine, St. Louis, Missouri 63110, USA.

ABSTRACT
Many cells coordinate their activities by transmitting rises in intracellular calcium from cell to cell. In nonexcitable cells, there are currently two models for intercellular calcium wave propagation, both of which involve release of inositol trisphosphate (IP3)- sensitive intracellular calcium stores. In one model, IP3 traverses gap junctions and initiates the release of intracellular calcium stores in neighboring cells. Alternatively, calcium waves may be mediated not by gap junctional communication, but rather by autocrine activity of secreted ATP on P2 purinergic receptors. We studied mechanically induced calcium waves in two rat osteosarcoma cell lines that differ in the gap junction proteins they express, in their ability to pass microinjected dye from cell to cell, and in their expression of P2Y2 (P2U) purinergic receptors. ROS 17/2.8 cells, which express the gap junction protein connexin43 (Cx43), are well dye coupled, and lack P2U receptors, transmitted slow gap junction-dependent calcium waves that did not require release of intracellular calcium stores. UMR 106-01 cells predominantly express the gap junction protein connexin 45 (Cx45), are poorly dye coupled, and express P2U receptors; they propagated fast calcium waves that required release of intracellular calcium stores and activation of P2U purinergic receptors, but not gap junctional communication. ROS/P2U transfectants and UMR/Cx43 transfectants expressed both types of calcium waves. Gap junction-independent, ATP-dependent intercellular calcium waves were also seen in hamster tracheal epithelia cells. These studies demonstrate that activation of P2U purinergic receptors can propagate intercellular calcium, and describe a novel Cx43-dependent mechanism for calcium wave propagation that does not require release of intracellular calcium stores by IP3. These studies suggest that gap junction communication mediated by either Cx43 or Cx45 does not allow passage of IP3 well enough to elicit release of intracellular calcium stores in neighboring cells.

Show MeSH
Related in: MedlinePlus