Ca(2+) regulation in differentiating lens cells in culture.
In the ocular lens, cataract formation is associated with an elevated intracellular Ca(2+) concentration (Ca(2+)(i)) resulting from the loss of lens cell Ca(2+) regulation. The mechanisms regulating Ca(2+)(i) have been characterized previously in lens epithelial cells, but have not been well characterized in the more differentiated lens fiber cells. The mechanisms regulating Ca(2+)(i) in clusters of fiber-like cells (lentoids) in a sheep lens primary cell culture system in which the epithelial cells differentiate into enlarged fiber-like cells were investigated. Only approximately 50% of the lentoids responded to thapsigargin and/or agonists (ATP and epinephrine), compared to>95% of the epithelial cells. Remarkably, most (90%) lentoids exhibited a resting cytosolic Ca(2+)(i) that was approximately three-fold greater than that in epithelial cells (approximately 100n M). This elevated resting cytosolic Ca(2+)(i) was not affected by thapsigargin treatment, but decreased upon removal of extracellular Ca(2+) or addition of the Ca(2+) channel blocker Gd(3+) (5mM ). These results suggest that a plasma membrane Ca(2+) channel is more active in lentoids than in epithelial cells. Indeed, when plasma membrane cation channel activity was monitored by Mn(2+) influx and quenching of fura-2 fluorescence, quenching was faster in lentoids than epithelial cells. Following thapsigargin treatment, capacitative Ca(2+) entry was activated in epithelial cells but not lentoids. In conclusion, during differentiation in primary cell culture, lens cells lose their ability to respond to agonists and exhibit an elevated resting Ca(2+)(i) that was dependent on the activation of a Ca(2+) influx pathway. The results of this study support the possibility that a sustained elevation in resting Ca(2+)(i) is one of the factors controlling lens cell differentiation, possibly by triggering events such as calpain activation.
Department of Biology, Georgia State University, Atlanta, GA, 30303, U.S.A.
Grant C
GC
Churchill
Charles F
CF
Louis
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000502:physiology*; D002454:Cell Differentiation / Q000502:physiology*; D002478:Cells, Cultured; D003600:Cytosol / Q000502:physiology; D004837:Epinephrine / Q000494:pharmacology; D004848:Epithelium / Q000502:physiology; D007908:Lens, Crystalline / Q000187:drug effects / Q000502:physiology*; D012756:Sheep; D019284:Thapsigargin / Q000494:pharmacology
D019284:Thapsigargin; D000255:Adenosine Triphosphate; D002118:Calcium; D004837:Epinephrine