Role of P-glycoprotein in restricting propranolol transport in cultured rabbit conjunctival epithelial cell layers

Purpose. To determine the role of P-glycoprotein (P-gp) in propranolol transport in cultured rabbit conjunctival epithelial cell layers (RCEC). Methods. The localization of P-gy in the cultured RCEC as well as in the excised conjunctiva was determined by immunofluorescence technique. The role of P-gp in transepithelial transport and uptake of propranolol in conjunctival epithelial cells cultured on Transwell filters was evaluated in the presence and absence of P-gp competing substrates, an anti-P-gp monoclonal antibody (4E3 mAb), or a metabolic inhibitor, 2, 4-dinitrophenol (2,4-DNP). Results. Immunofluorescence studies revealed positive staining in the apical membrane of cultured RCEC and in the apical surface of the superficial cell layers in the excised conjunctiva, but not the basolateral membrane of cultured RCEC. Transport of propranolol showed preference in the basolateral-to-apical direction. The net secretory flux was saturable with a K-m of 71.5 +/- 24.0 nM and a J(max) of 1.45 +/- 0.17 pmol/cm(2)/hr. Cyclosporin A, progesterone, rhodamine 123, verapamil, 4E3 mAb and 2,4-DNP all increased apical 50 nM propranolol uptake by 43% to 66%. On the other hand, neither beta-blockers (atenolol, metoprolol, and alprenolol) nor organic cation transporter substrates (tetraethylammonium (TEA) and guanidine), affected apical 50 nM propranolol uptake. Conclusions. The energy-dependent efflux pump P-gp appears to be predominantly located on the apical plasma membrane of the conjunctival epithelium. It may play an important role in restricting the conjunctival absorption of some lipophilic drugs.