These experiments were undertaken to determine the correspondence between acidosis-induced in situ motochondrial glutamine uptake and the process by which glutamine moves across the mitochondrial membrane. Feeding rats 1.5% NH4Cl for 2 wk accelerated the in situ uptake rate from 0.12 +/- 0.08 to 1.89 +/- 0.28 mu mol/min or some 16-fold. To determine glutamine uptake independent of its metabolic conversion, D-glutamine was employed. In isolated mitochondria from non-acidotic rat kidneys, D-glutamine diffused into 71 +/- 10% of the mitochondria water volume; in acidotic mitochondria the diffusion volume increased to 127 +/- 16%. The reflection coefficients (sigma) for a series of amides, including glutamine, were determined by gravimetrically following volume decrements in increasing concentrations of solute; D-glutamine's sigma fell from 1.05 +/- 0.08 to 0.50 +/- 0.06 in acidotic rat kidney mitchondria, The permeability coefficients corresponding to the measured sigma were 10(-7) cm/s and 10(-5) cm/s in nonacidosis and acidosis, respectively. When viewed in situ proximal tubule mitochondria undergo dramatic alterations during chronic acidosis. These involved an enlargement in the mitochondria particularly in the basal region of the cell and a reduction in number. Furthermore, numerous autophagic vacuoles, containing mitochondria, appear in the basal region. The findings are consistent availability that becomes activated during acidosis as a consequence of mitochondrial glutamine permeability resulting in increased ammoniagenesis and accelerated organelle turnover.