plasma membrane, active transport, protein pumps, protein channels, ion channels, osmolality, intracellular messengers, erythrocytes
The mechanisms by which cells compensate for volume fluctuations are not clearly understood and vary among species. Research efforts in our lab have focused on elucidating the pathways involved in regulatory volume decrease (RVD), the process activated in response to cell swelling that allows for volume recovery. Previously, fluorescence microscopy studies performed by Light et al. (2005) revealed that in salmon red blood cells, cell swelling elicits a rise in intracellular Ca2+ concentration (visualized using fluorescence microscopy and the Ca2+ indicator fluo-4-AM). This was most likely due Ca2+ influx from the extracellular environment, because it was not observed in cells bathed in a hypotonic, low Ca2+ medium. The goal of this study, therefore, was to confirm a role for extracellular Ca2+ in the RVD response, using both Salmo salar (Atlantic salmon) and Alligator mississippiensis (American alligator) red blood cells. This was done by exposing cells to different extracellular environments and pharmacological agents that block Ca2+ influx pathways or Ca2+-mediated intracellular signaling cascades. To asses the effects of these manipulations on RVD, median cell volume changes over a 90 minute time course were determined by electronic sizing using a Coulter counter. Salmon cells exposed to a low Ca2+ environment failed to recover from cell swelling, indicating that extracellular Ca2+ was needed for a successful RVD response. Similarly, volume regulation of alligator red blood cells occurred by a Ca2+-dependent mechanism. Additionally, RVD in alligator cells appeared to occur through an intracellular signaling cascade involving Ca2+ activation of phospholipase A2 and the subsequent formation of arachidonic acid. Arachidonic acid itself, as opposed to one of its potential breakdown products, aided in volume recovery by stimulating K+ efflux. In conclusion, the results from this study indicate that Ca2+ plays a pivotal role in the RVD response of both salmon and alligator red blood cells.
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