Abstract
Purpose::
The regulation of fluid transport across the corneal endothelium by extra- and intracellular messengers is poorly understood. We have examined the effects of modulation of PKC with activators and inhibitors on corneal fluid transport.
Methods::
Corneas were obtained from New Zealand albino rabbits (~2 kg) using procedures in accordance with NIH guidelines. Transendothelial fluid movement was determined by measuring the changes in corneal thickness with the Dikstein-Maurice method. Corneal thickness was determined with a computer-controlled specular microscope, which automatically sensed and recorded it at 5-minute intervals. Fluid transport was calculated from the rates of corneal thickness change as reported previously.
Results::
Inhibition of PKC activity with calphostin C, an inhibitor of conventional and novel PKC isoforms, resulted in a dose-dependent increase in fluid movement and a reduction of corneal thickness. The effect of PKC activators was found to be more complex. Phorbol-12, 13- dibutyrate (PDBu) and Phorbol-12-myristate-13-acetate (PMA) produce an initial increase followed by a reduction in fluid transport. We have further attempted to identify the membrane component affected by PKC modulation. Corneal swelling in bicarbonate-free solutions was temporarily arrested by exposure to 1 µM calphostin C. However, if corneal swelling was induced by inhibition of anion channels using a cocktail of 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB) and niflumic acid (NA) both 100 µM, then exposure to Calphostin C did not affect the rate of swelling.
Conclusions::
Activation of PKC may result in an initial activation of fluid transport followed by inhibition of it. Such activation and/or inhibition may be due to effects on anion channels.
Keywords: cornea: endothelium • ion transporters • ion channels