Visualization of iontophoretic pathways with confocal microscopy and the vibrating probe electrode

The primary barrier to the permeation of water and charged compounds through mammalian skin is an extracellular lipid matrix in its outermost layer, but skin is traverse by hair follicles and sweat glands, which may act as shunts. Transdermal permeation can be electrically facilitated (iontophoresis), but the pathways of this current flow are not known. We have used a vibrating probe electrode (VPE) to identify and vectorize site-specific ionic flows at the surface of current-clamped mammalian skin. The currents located were primarily appendageal. Laser-scanning confocal microscopy (LSCM) can optically section thick tissues that are sufficiently transparent, do not strongly scatter light, and have low autofluorescence. The permeation pathways of iontophoretically driven fluorescent probes (used as model compounds) were visualized with LSCM in whole, unfixed skin. Transport of charged and polar substances appears to take place primarily via appendageal routes, whereas (non-facilitated) lipophilic fluores travel by paracellular pathways in the stratum corneum, and by both paracellular and transcellular paths in the living layers of the skin. Conventional methods (e.g., TEM) of tissue pathway visualization provide static images of pathways, and sample preparation may substantially modify tissue structure. dynamic information about current flow into and through living skin can be obtained by the VPE and by LSCM.