Regeneration of resistance and ion transport in rabbit corneal epithelium after induced surface cell exfoliation

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Abstract
Exposure of the rabbit corneal surface to a 20-μm digitonin-0.9% NaCl solution leads to permeabilization of the most superficial cells of the stratified epithelium. The devitalized cells exfoliate spontaneously from the corneal surface. Detergent exposure limited to 4–8 min leads to permeabilization and rapid exfoliation of a monolayer of surface cells. Consistent with the presence of the epithelial paracellular permeability barrier in this cell layer, their permeabilization results in complete loss of transepithelial resistance (R t ). Within minutes after detergent removal an initial recovery ofR t can be noticed indicating generation of a new paracellular permeability barrier by the viable subsurface cells. This recovery proceeds rapidly andR t reaches within 70 min a maximum equal to > 90% of the preexfoliation values (=2.43 kΩ·cm2,n=22). TheR t recovery is fully blocked in a reversible manner by 10 μm dihydrocytochalasin B. The recovery is not affected by inhibition of protein synthesis with 5 μm cycloheximide. When the ocular surface is treated again with digitonin the permeabilization and exfoliation of a monolayer of cells and loss ofR t are repeated. After the second detergent exposure an initial recovery ofR t occurs as before within minutes. However, the pace ofR t recovery is much slower: 4–5 hr are required to reach a stable maximalR t values amounting to about 73% of initial control. This recovery can be fully blocked by 5 μm cycloheximide indicating that protein synthesis is required for generation of tight junctions by the second subcellular layer. With only a fraction ofR t recovered, short-circuit currents amounting to, at least, 50% of control values and attributable in part to cell-to-tear movement of Cl− through the apical surface can be measured. This suggests that apical-type Cl− channels are either present in the apically facing membrane of subsurface cells or that they are rapidly inserted in it from preexisting intracellular pools immediately following the devitalization of the surface cells by digitonin.