The transport pathways and permeation kinetics of lipophilic and hydrophilic fluorescent dyes through porcine mammary papillae were visualized and quantified. Porcine mammary papillae, removed from full-thickness abdominal tissue, were positioned in a Franz diffusion cell for passive diffusion studies. Solutions containing the fluorescent dyes were applied topically for time periods ranging from 30min to 48h. Dye concentrations in tissue and Franz diffusion compartments were analyzed using fluorescence microscopy and fluorimetry. Fluorescence micrographs elucidated two permeation pathways, transepidermal and transductal. Hydrophilic sulforhodamine B predominantly penetrated via the transepidermal route, while lipophilic nile red diffused mainly by the transductal route. An almost 4-fold higher amount of sulforhodamine B was retained within the nipple over time compared to nile red, despite both dyes permeating through the tissue at similar rates. Diffusion through the porcine nipple was 500-fold higher than through adjacent skin for both dyes, likely attributable to the two mammary ducts which provide an entry point and transport route through the tissue. These results, generated from both qualitative and quantitative evidence at a micro and macro scale, demonstrate that the mammary ducts provide a direct pathway that contributes significantly to passive transport through the nipple, particularly for lipophilic molecules.