The purpose of this investigation was to study the effects of altered extracellular Po(2) (Pe(O(2))) on the intracellular Po(2) (Pi(O(2))) response to contractions in single skeletal muscle cells. Single myocytes (n = 12) were dissected from lumbrical muscles of adult female Xenopus laevis and injected with 0.5 mM Pd-meso-tetra(4-carboxyphenyl)porphine for assessment of Pi(O(2)) via phosphorescence quenching. At a Pe(O(2)) of approximately 20 (low), approximately 40 (moderate), and approximately 60 (high) Torr, tetanic contractions were induced at a frequency of 0.67 Hz for approximately 2 min with a 5-min recovery between bouts (blocked order design). The Pi(O(2)) response to contractions was characterized by a time delay followed by a monoexponential decline to steady-state (SS) values. The fall in Pi(O(2)) to SS values was significantly greater at each progressively greater Pe(O(2)) (all P < 0.05). The mean response time (time delay + time constant) was significantly faster in the low (35.2 +/- 5.1 s; P < 0.05 vs. high) and moderate (43.3 +/- 6.4 s; P < 0.05 vs. high) compared with high Pe(O(2)) (61.8 +/- 9.4 s) and was correlated positively (r = 0.965) with the net fall in Pi(O(2)). However, the initial rate of change of Pi(O(2)) (calculated as net fall in Pi(O(2))/time constant) was not different (P > 0.05) among Pe(O(2)) trials. These latter data suggest that, over the range of 20-60 Torr, Pe(O(2)) does not play a deterministic role in setting the initial metabolic response to contractions in isolated frog myocytes. Additionally, these results suggest that oxidative phosphorylation in these myoglobin-free myocytes may be compromised by Pe(O(2)) at values nearing 60 Torr.