The relationship between osmotic stress and calcium elevation: in vitro and in vivo rat lens models

Both in vivo and in vitro models were employed in the present study to assess the relative contribution of osmotic stress and increasing calcium levels to the development of sugar cataracts. In galactose cataract obtained from galactosemic weanling rats, the concentration of total calcium increased by nearly 10% at the first sign of visible opacification observed on the fourth day post-galactose feeding. After 7 days of galactose feeding, calcium levels continued to rise, to 0.8 mM. During the first 10 days, loss of lens transparency and calcium elevation was gradual and steady, with precipitous changes occurring on days 11 and 12. In groups of rats where galactose feeding was stopped after 7 days, cataract reversal was followed during the next 5 weeks. During the initial first week of recovery, calcium influx and elevation in the lens continued but began to decline steadily thereafter. After 3 weeks of recovery, lens transparency had returned to almost normal. Calcium levels continued to decline and reached normal levels between day 34 and 42, nearly 4 weeks after removal of the galactose diet. The relationship between osmotic stress and calcium elevation was investigated more directly by culturing normal rat lenses in hypoosmotic medium (280 mOsm) to create osmotic gradients similar to that in galactosemic lenses. The results showed that during the first day of culture (12 hr), osmotically stressed lenses gained 3 mg of water, became opaque and gained excess calcium (7 mM compared to 0.7 mM). Microscopic vacuoles appeared to accompany the process of opacification and contributed to increased light scattering and the loss of lens transparency. Additional experiments were designed to further distinguish between the effects of osmotic stress and calcium elevation on the opacification process. Thus, lenses were incubated in control and high-calcium medium (20 mM) at 300 mOsm. Within 12 hr of incubation, calcium elevation progressed to 1.37 mM, nearly doubling the normal value. Although opacification was observed in these lenses, no sign of vacuoles was evident. Collectively, the findings from this study support the premise that an early influx of calcium is brought about by osmotic stress and is responsible for the observed loss in transparency in osmotic (sugar) cataract.