Involvement of Carrier-Mediated Transport in the Retinal Uptake of Clonidine at the Inner Blood–Retinal Barrier

Abstract

In the present study, the blood-to-retina transport across the inner BRB was investigated for clonidine, a compound which is expected to exhibit a neuroprotective effect for the treatment of severe retinal diseases. In the in vivo study, the integration plot analysis for [(3)H]clonidine exhibited an apparent influx permeability clearance of 457 μL/(min·g retina) in the retina. The in vivo inhibition study suggests that the blood-to-retina transport of clonidine at the BRB is organic cation-sensitive since clonidine, pyrilamine, and propranolol, at a concentration of 40 mM, significantly reduced the retinal uptake index (RUI) of [(3)H]clonidine, and an inhibitory effect on the RUI was also exhibited by verapamil, at a concentration of 3 mM. The in vitro study with TR-iBRB2 cells, an in vitro model cell line of the inner BRB, suggests that carrier-mediated transport is involved in the blood-to-retina transport of clonidine at the inner BRB since the results obtained demonstrated time-, temperature-, pH-, and concentration-dependent [(3)H]clonidine uptake, with a Km of 286 μM. In the in vitro inhibition study, the [(3)H]clonidine uptake was significantly reduced by several organic cations, such as clonidine, verapamil, pyrilamine, and propranolol, and was competitively inhibited by 200 μM verapamil, in spite of slight or no significant alteration being produced with organic anions. Furthermore, the typical substrates and inhibitors of well-known organic cation transporters had no significant effect on the uptake of [(3)H]clonidine to suggest the involvement of novel transporter molecules in the transport of clonidine across the inner BRB. These results suggest that the blood-to-retina transport of clonidine across the inner BRB involves a carrier-mediated transport manner, suggesting the contribution of a novel organic cation transporter expressed by the retinal capillary endothelial cells.

Influx Transport of Cationic Drug at the Blood–Retinal Barrier: Impact on the Retinal Delivery of Neuroprotectants

“…2C), and were inhibited in the presence of cationic drugs, such as verapamil and pyrilamine. 24,25) These results suggest the involvement of facilitative transport system in the influx transport of propranolol and clonidine at the BRB, supporting the utility of these transport systems in the efficient and safe delivery of cationic neuroprotectants to the retina.…”

“…Similarly, in the in vitro uptake study of [ Figure 4B shows the results obtained in the concentration-dependent uptake of [ 3 H] clonidine by TR-iBRB2 cells. 24,25) In addition, the uptake study of pyrilamine by TR-iBRB2 cells also suggests the involvement of carrier-mediated processes, including high-and low-affinity transport, in the influx transport of pyrilamine across the inner BRB. 21) Table 2 presents the kinetic parameters suggested for cationic drug transport at the inner BRB.…”

“…5). The in vitro inhibition study revealed that substrate specificity of the cationic drug transport at the BRB is different from those of well-characterized organic cation transporters, such as plasma membrane monoamine transporter (PMAT/ SLC29A4/ENT4), multidrug and toxin extrusion (SLC47/A1), organic cation/carnitine transporters (OCTNs/SLC22A4-5) and organic cation transporters (SLC22A1-3), 21,24,25) suggesting the involvement of novel organic cation transporters in the influx transport of cationic drugs across the BRB.…”

“…The figure was prepared by reference to previous reports. 7,16,17,21,24,25) PAH, p-aminohippuric acid. 16,17) of verapamil at the BRB.…”

“…4C), suggesting the involvement of organic cation transporter in verapamil transport at the inner BRB. 21 21,24,25) strongly suggesting that the delivery of neuroprotectants through carrier-mediated cationic drug transport is a promising way to improve the treatment of retinal diseases.…”