Fluid Transport across the Isolated Porcine Ciliary Epithelium

Abstract

To quantify spontaneous fluid transport across the isolated porcine ciliary epithelium and determine its sensitivity to the electrolyte transport inhibitors ouabain and bumetanide, as well as bath Cl(-) and HCO(3)(-) levels. A complete annulus of ciliary body was mounted in a custom-designed chamber appropriate for quantifying net fluid movement, as well as the transepithelial potential difference (PD) across the in vitro ciliary epithelium. A spontaneous and stable fluid flow (FF) in the blood-to-aqueous direction was measured over a 4-hour period. This flux solely reflected the secretory activity of the isolated ciliary epithelium (CE), given the absence of externally applied osmotic or pressure gradients. In contrast to FF, the PD declined during the 4 hours in vitro, suggesting that the integrity of the tight junctions may have been compromised during this time so that an increased movement of counter ions via the paracellular pathway could have shunted the PD, while at the same time transcellular fluid transport remained unaffected. The FF in the blood-to-aqueous direction (2.3 +/- 0.2 muL/hr; n = 7) was eliminated by a unilateral reduction in the bath Cl(-) levels on the blood side of the preparation and restored on reintroducing the anion to the bathing medium. This linkage between FF and blood side [Cl(-)] is consistent with the existence of a net Cl(-) flux across the porcine CE in the same direction as the fluid transport. Addition of bumetanide to the blood-side bath inhibited FF by approximately 40%, whereas the removal of CO(2)/HCO(3)(-) from the blood-side bathing solution elicited a approximately 50% reduction in FF. Ouabain inhibited the FF from either side of the preparation, although the effects were more rapid when the glycoside was applied to the blood side of the tissue. Overall, these findings indicate the dependence of FF on active ionic transport by the isolated CE. Isolated porcine ciliary epithelial preparations transport fluid in the blood-to-aqueous direction, indicating that measurements of volumetric fluid flow across this preparation may serve as a suitable model for future studies directed toward the pharmacological control of secretion.