Mechanism of calcium-induced disintegrative globulization of rat lens fiber cells

Abstract

Purpose: To study the role of calcium and calcium-dependent processes in the disintegrative globulization of isolated single rat lens cortical fibers.

Methods: The authors isolated viable and morphologically intact single fiber cells from rat lens cortex and studied the effect of 1 mM [Ca2+]o on the globulization of fiber cells from the outer and inner cortex. They investigated the effects of the calcium-channel blocker, verapamil; an inhibitor of calcium transport, lanthanum; various protease inhibitors; Na+ -free and K+ -free media; calcium ionophore, A23187; and calcium chelator, BAPTA, on the globulization of fiber cells exposed to 1 mM [Ca2+]o.

Results: Perfusion with Ringer's solution containing 1 mM [Ca2+]o, caused disintegration and globulization of the isolated fibers in 32.3 +/- 1 minute, and the addition of 10 microM A23187 to the superfusing solution reduced the time to complete globulization (tg) to 19.4 +/- 0.3 minutes. However, the addition of protease inhibitors, leupeptin, calpain inhibitor I, E-64, or pepstatin (0.5 mM each) to the superfusing solution, increased tg to 105 +/- 3.5, 84.2 +/- 7.8, 60.7 +/- 3.5, and 48.3 +/- 3.1 minutes, respectively. The tg also increased (96.4 +/- 3.5 minutes) when the fibers were preincubated with BAPTA-AM or when they were exposed to 1 mM [Ca2+]o in Na+ - or K+ -free Ringer's solution (tg = 66.7 +/- 5.3 and 58.9 +/- 3.9 minutes, respectively) or in Ringer's solution containing 1 mM [Ca2+]o + 50 microM verapamil (tg = 49.3 +/- 3.8 minutes). Single fibers isolated from the outer cortex of the lens were less susceptible to extracellular calcium than those isolated from the inner cortex.

Conclusions: Increased calcium influx and the attendant elevation of [Ca2+]i are necessary for disintegrative globulizaiton of lens fiber cells. Calcium influx appears to be mediated partially by the L-type calcium channels and the background calcium leak. Protection by protease inhibitors suggests that membrane fragmentation, caused by elevated [Ca2+]i, results from proteolytic damage to the fiber cytoskeleton. Besides underscoring the central role of calcium homeostasis in preserving the morphologic integrity of the cortical fibers, this study suggests a possible cellular mechanism for the formation of supranuclear cataract.