Subcellular NH 4 + flux analysis in leaf segments of wheat ( Triticum aestivum )

• We report the first use of tracer 13 NH 4 + ( 13 N-ammonium) efflux and retention data to analyse subcellular fluxes and compartmentation of NH 4 + in the leaves of a higher plant (wheat, Triticum aestivum ). • Leaf segments, 1–2 mm, were obtained from 8-d-old seedlings. The viability of the segments, and stability of NH 4 + acquisition over time, were confirmed using oxygen-exchange and NH 4 + -depletion measurements. Fluxes of NH 4 + and compartment sizes were estimated using tracer efflux kinetics and retention data. • Influx and efflux across the plasma membrane, half-lives of exchange and cytosolic pool sizes were broadly similar to those in root systems. As the external concentration of NH 4 + ([NH 4 + ] o ) increased from 10 μ M to 10 m M , both influx and efflux greatly increased, with a sixfold increase in the ratio of efflux to influx. Half-lives were similar among treatments, except at [NH 4 + ] o = 10 m M , where they declined. Concentrations of NH 4 + in the cytosol ([NH 4 + ] c ) increased from 2.6 to 400 m M . • Although [NH 4 + ] c became large as [NH 4 + ] o increased, the ratio of [NH 4 + ] c to [NH 4 + ] o decreased more than sixfold. The apparently futile cycling of NH 4 + at high [NH 4 + ] o suggested by the large fluxes of NH 4 + in both directions across the membrane indicate that leaf cells respond to potentially toxic NH 4 + concentrations in a manner similar to root cells.