Evidence for Cotransport of Nitrate and Protons in Maize Roots : II. Measurement of NO(3) and H Fluxes with Ion-Selective Microelectrodes

We report here on an investigation of net nitrate and proton fluxes in root cells of maize (Zea mays L.) seedlings grown without (noninduced) and with (induced) 0.1 millimolar nitrate. A microelectrode system described previously (IA Newman, LV Kochian, MA Grusak, WJ Lucas 1987] Plant Physiol 84: 1177-1184) was utilized to quantify net ionic fluxes from the measurement of electrochemical potential gradients for NO₃⁻ and H⁺ within the unstirred layer at the root surface. The nitrate-inducibility, pH dependence, and concentration dependence of net NO₃⁻ uptake correlated quite closely with the electrical response of maize roots to nitrate under the same experimental conditions (as described in PR McClure, LV Kochian, RM Spanswick, JE Shaff 1990] Plant Physiol 93: 281-289). Additionally, it was found that potential inhibitors of the plasmalemma H⁺-ATPase (vandate, diethylstilbestrol), which were shown to abolish the electrical response to NO₃⁻ (in PR McClure, LV Kochian, RM Spanswick, JE Shaff 1990] Plant Physiol 93: 281-289), dramatically inhibited NO₃⁻ absorption. These results strongly indicate that the NO₃⁻ electrical response is due to the operation of a NO₃⁻ transport system in the plasmalemma of maize root cells. Furthermore, the results from the H⁺-ATPase inhibitor studies indicate that the NO₃⁻ transport system is linked to the H⁺-ATPase, presumably as a NO₃⁻/H⁺ symport. This is further supported by the pH response of the NO₃⁻ transport system (inhibition at alkaline pH values) and the change in net H⁺ flux from a moderate efflux in the absence of NO₃⁻, to zero net H⁺ flux after exposing the maize root to exogenous nitrate. Although these results can be explained by other interpretations, the simplest model that fits both the electrical responses and the NO₃⁻/H⁺ flux data is a NO₃⁻/H⁺ symport with a NO₃⁻:H⁺ flux stoichiometry >1, whose operation results in the stimulation of the H⁺-ATPase due to the influx of protons through the cotransport system.