Uptake of orthophosphate by rabbit vagus nerve fibres

1. The uptake of orthophosphate and its incorporation into ATP, ADP, and creatine phosphate (CrP) were studied in desheathed rabbit vagus nerve. 2. Using -32P labelled orthophosphate, the total amount of labelled phosphate taken up by the preparation was continuously recorded in a perfusion apparatus. For measuring the incorporation into phosphorylated compounds, phosphate esters and inorganic phosphate were extracted, separated and their total amount and radioactivity determined. 3. The total uptake of phosphate was found to be a biexponential function of time. 4. The time constant of the first process was 10-20 min and independent of the extracellular phosphate concentration, the final amount labelled by this process was relatively small and proportional to external phosphate, increasing from 0.026 m-mole/kg wet nerve at 0-04 mM phosphate to 1-14 m-mole/kg at 5nM. 5. The time constant of the second process depended on the extracellular phosphate concentration varying from 4624 min at 0-04 mM to 210 min at 5 mM. The final amount labelled by this process was 5-6 m-mole/kg wet wt. and independent of the extracellular phosphate. 6. The kinetics of the slow uptake were consistent with the presence of a saturable process and a non-saturable one. 7. Extraction of ATP, ADP, and the sum of CrP and Pi, showed that the total amount of these compounds remained constant for 2 hr while their radioactivity increased slowly, approximately at the same rate as the slow fraction. 8. Increasing the external phosphate from 0-04 to 5 nM increased the amount of labelled ATP. 9. A comparison with the metabolic turnover of phosphate, estimated from the oxygen consumption, shows that uptake is much slower than metabolism, so that the slow appearance of labelled nucleotides is very probably due to a limitation of the influx. 10. From the experimental data the influx can then be calculated for various phosphate concentrations. It is close to that found in squid axons.