In the first of two sets of experiments, the lumen-to-cell and cell-to-bath transport rates for glycine were measured in the isolated-perfused medullary pars recta (S₃ cells) of the rabbit proximal tubule at multiple luminal glycine concentrations (0-2.0 mM). The lumen-to-cell transport of glycine saturated which permitted the calculation of the J_max (pmol min^-1 (mm tubular length)^-1), Km (mM) and k (paracellualar leak, pmol min^-1 (mm tubular length)^-1 mM^-1) values for this transport mechanism, these values were 4.3, 0.3, and 0.03, respectively. The cell-to-bath transport did not saturate but showed a linear relationship to cellular glycine concentration, 0.58 (pmol min^-1 (mm tubular length)^-1 mM^-1). The second set of experiments characterized the transport rate, cellular accumulation and metabolic rate of lumen-to-cell transported ³H-glycine in all segments (cell types) of the proximal tubule, pars convoluta (S₁ cells), cortical pars recta (S₂ cells) and medullary pars recta (S₃ cells). These proximal tubular segments were isolated and perfused at a single glycine concentration, 11.4µM. From the results of this study and previous work (3), we conclude that the axial heterogeneity for glycine lumen-to-cell and cell-to-bath transport capacity extends to the medullary pars recta (S₃ cells) (S₁ >> S₂ < S₃ for lumen-to-cell transport and S₁>S₂>>S₃ for cell-to-bath transport). Also, we conclude that lumen-to-cell transported glycine can be metabolized and its metabolic rate displays axial heterogeneity (S₁>S₂>S₃). The physiological significances of these transport and metabolic characteristics of the S₃ cell type permits the medullary pars recta to effectively recover glycine from very low luminal glycine concentrations and makes glycine available for protective and maintenance metabolism of the medullary pars recta.