Examination of barriers and barrier alteration in transscleral iontophoresis

Abstract

The flux enhancing mechanisms of transscleral iontophoresis are not well understood. The objective of the present study was to investigate the ocular barrier and barrier alterations in transscleral iontophoretic delivery with magnetic resonance imaging (MRI). Experiments involving constant current transscleral iontophoresis of 2 mA (current density 10 mA/cm(2)) and subconjunctival injection were conducted with rabbits in vivo and postmortem and with excised sclera in side-by-side diffusion cells in vitro. The postmortem and in vitro experiments were expected to be helpful in clarifying the importance of vascular clearance and other transport barriers in transscleral iontophoresis. Manganese ion (Mn(2+)) and manganese ethylenediaminetetraacetic acid complex (MnEDTA(2-)) were the model permeants. The results show that pretreatment of the eye with an electric field by iontophoresis enhanced subconjunctival delivery of the permeants to the anterior segment of the eye in vivo. This suggests that electric field-induced barrier alterations can be an important absorption enhancing mechanism of ocular iontophoresis. Penetration enhancement was magnified in the postmortem experiments with larger amounts of the permeants delivered into the eye and to the back of the eye. The different results observed in the in vivo and postmortem studies can be attributed to ocular clearance in ocular delivery.

Figure 1

Rabbit eye diagram.

Figure 2

Representative MR images of in vivo studies in Experiments A (blank iontophoresis of 2 mA followed by 0.1 mL subconjunctival injection) and Experiment III (0.1 mL subconjunctival injection without iontophoresis pretreatment). a: Experiment A, injection of 40 mM Mn²⁺ showing significant penetration enhancement, (b) Experiment III, injection of 40 mM Mn²⁺, (c) Experiment A, injection of 0.1 M MnEDTA²⁻ showing some penetration enhancement, and (d) Experiment III, injection of 0.1 M MnEDTA²⁻. From left to right, images obtained at (a) 40 and 70 min, (b) 30 and 138 min, (c) 21 and 124 min, and (d) 40 and 120 min after injection, respectively. Images in (b) and (d) are obtained from a previous study. The numbers indicate the MR signals from the region of interest (average of 9 voxels or 4 mm³). The orientations of the eyes in the images (facing left or right) denote the left or right eye.

Figure 3

Representative MR images of postmortem studies in (a) Experiment B, blank iontophoresis of 2 mA followed by 0.1 mL subconjunctival injection of 40 mM Mn²⁺ and (b) Experiment IV, 0.1 mL subconjunctival injection of 40 mM Mn²⁺. From left to right, for (a) and (b), images obtained at 30 and 60 min after injection. Images in (b) are obtained from a previous study. The numbers indicate the MR signals from the region of interest (average of 9 voxels or 4 mm³).

Figure 4

Representative MR images of postmortem transscleral iontophoresis experiment of 2 mA of (a) 0.1 M MnEDTA²⁻, (b) 4 mM Mn²⁺, and (c) 40 mM Mn²⁺ in Experiment C. From left to right, images obtained at (a) 9 and 19 min during iontophoresis, and 10, 55 and 140 min after iontophoresis, (b) 10 and 19 min during iontophoresis, and 40, 60, and 190 min after iontophoresis, and (c) 9 and 18 min during iontophoresis, and 10, 60, and 140 min after iontophoresis. Note that high Mn²⁺ and MnEDTA²⁻ concentrations such as 40 mM Mn²⁺ and 0.1 M MnEDTA²⁻ in the electrode device are represented by dark voxels in the images due to the parabolic signal versus concentration relationship. The numbers indicate the MR signals from the region of interest (average of 9 voxels or 4 mm³).

Figure 5

Representative MR images of postmortem transscleral iontophoresis of 4 mM Mn²⁺ and 2 mA postmortem in Experiments C and D. From left to right, images obtained at (a) 10 and 19 min during iontophoresis, and 40 and 60 min after iontophoresis, with electrode at the pars plana and (b) 11 and 20 min during iontophoresis, and 40 and 86 min after iontophoresis, with electrode at the back of the eye near the fornix. The numbers indicate the MR signals from the region of interest (average of 9 voxels or 4 mm³).