SUMMARY1. 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 114 m-mole/kg at 5 mm.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 same rate as the slow fraction.8. Increasing the external phosphate from 0 04 to 5 mm increased the amount of labelled ATP.

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B. ANNER AND OTHERS 10. The influx of phosphate was calculated for different external phosphate concentrations using the rate of uptake of radiophosphate measured 45 min after the application of isotope and the corresponding efflux rate coefficients and the intracellular specific activities of the labelled compounds.11. A correction was also introduced for diffusion, using the 'limited biophase model'.12. The Na-sensitive influx, i.e. the difference between influx in Locke and influx in choline-Locke, showed saturation kinetics.13. A Lineweaver-Burk plot for Na-sensitive uncorrected influxes gave a vmax of 16-7 #tmole/kg wet wt. min and an apparent Km of 0-42 mM;for the corrected fluxes these values were 18-2 and 0-36. 14. It is concluded that a large part of phosphate influx and some of the phosphate efflux is mediated by a specific Na-dependent phosphate transport system.15. This system seems to be present also in other types of nervous tissues and probably in many types of animal cells.

SU-MMARY1. Uptake and release of radiophosphate were measured in desheathed rabbit vagus nerve.2. During incubation in Locke containing 0-2 mM-[32P]phosphate a slow labelling of water soluble compounds of the nerve was found; the labelling of the non water soluble compounds was much smaller. During washing with inactive Locke, the label was almost entirely released from the water soluble compounds; the radioactivity of these compounds was therefore used as the basis for the calculation of the efflux rate constants.3. The efflux of radiophosphate increased with increasing phosphate concentration of the washing fluid.4. A similar effect of external phosphate on the efflux of radiophosphate was seen when the phosphate concentration was suddenly changed. The rate constants were in 0 02 mM-phosphate PM29 x 10-m d-1, in 0-2 mM 1-95 x 10-3 min-I, and in 2 mm 3-21 x 10-3 min-' at 37 'C. After changing the external solution the efflux reached a new level with a time constant of about 9 mi.5. Addition of arsenate also increased the efflux of radiophosphate; on a molar basis the effect of arsenate was slightly smaller than the effect of phosphate. the ratio between the Na dependent effluxe8 in 2 and 0-2 mM-phosphate was approximately equal to the ratio between the Na dependent influxes in 2 and 0*2 mMphosphate.10. The efflux of 22Na had a rate constant of 0 050 min-' in Locke and 0x046 minin Tris-Locke.11. It is concluded that part of the efflux of phosphate is mediated by a 508 J. FERRERO AND OTHERS Na-dependent transport system; the system appears to be able to exchange phosphate between the inside and the outside and to mediate net movements of phosphate in both directions.

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