Subcellular NH4 + flux analysis in leaf segments of wheat ( Triticum aestivum ).

Summary • We report the first use of tracer ¹³ NH₄ ⁺ ( 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 µm to 10 mm, 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 mm, where they declined. Concentrations of NH ₄ ⁺ in the cytosol ([NH ₄ ⁺ ] _c ) increased from 2.6 to 400 mm. • 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. [ABSTRACT FROM AUTHOR]