The time course of changes in metabolite concentrations and ultrastructure during ischemia was studied in rat kidney cortex by incubating tissues at 37°C for 15, 30, 60, and 120 min or at 0°C for 0.5, 6, 12, 24, 48, 72, and 96 hr. Ischemia at 37°C for 15 min caused a rapid fall of ATP (13% of control) and ADP (42%) with a concomitant increase in AMP (2.9-fold) and P 1 (2.8-fold). At 60 and 120 min, AMP also decreased resulting in the decrease of total adenine nucleotide pool. At 0°C, ATP was 50% of control at 0.5 hr and was maintained at this level up to 72 hr. The changes in other adenine nucleotides were also moderate at 0°C. The accumulation of lactate was the highest at all time intervals studied (at 37°C, 11-fold at 15 min and 26-fold at 120 min; at 0°C, 5-fold at 0.5 hr and 14-fold at 96 hr). The two rate limiting enzymes of glycolysis, i.e., phosphofructokinase and pyruvate kinase were activated during ischemia to different extents depending on the temperature and length of time of ischemia based on the metabolite patterns. At 37°C, swelling of mitochondria with loss of matrix granules and the formation of flocculent densities in proximal tubule cells were the prominent ultrastructural features which progressed with time. At 0°C, swelling, cytoplasmic clumping and dilatation of endoplasmic reticulum were observed. Mitochondrial flocculent densities appeared at 96 hr. Based on metabolic and ultrastructural observations, 1 hr of ischemia at 37°C and 72 hr at 0°C were found to be the “point-of-no-return” from ischemic injury in renal cortex.