Simultaneous measurement of ammonium,nitrate and proton fluxes along the length of eucalypt roots

Knowledge of the preferred source of N for Eucalyptus nitens will lead to improved fertiliser management practices in plantations. Ion selective microelectrodes were used non-invasively to measure simultaneously net fluxes of NH₄ ⁺, NO₃ ^– and H⁺ along the tap root of solution-cultured E. nitens. Measurements were conducted in solutions containing 100 m NH₄NO₃. The pattern of fluxes was such that there was a large influx of NH₄ ⁺, a smaller influx of NO₃ ^– and large H⁺ efflux. The ratio of these fluxes was constant, according to the ratio 3:1:–6 (NH₄ ⁺:NO₃ ^–:H⁺). Within the region 20–60 mm from the root apex of E. nitens seedlings there was spatial and temporal variation in fluxes but flux patterns remained constant. Root hair density did not affect fluxes nor did proximity to lateral roots. Variation was less than that found in previous studies of localised root fluxes using similar high-resolution measurement techniques. It was concluded that small-scale spatial variation in fluxes may have confounded previous studies. There were associations between fluxes of all three ions, the strongest associations being between NH₄ ⁺ and H⁺, and NH₄ ⁺ and NO₃ ^–. Overall, these results are consistent with NH₄ ⁺ being the preferred source N for E. nitens.     

Local measurements of nitrate and potassium fluxes along roots of maritime pine. Effects of ectomycorrhizal symbiosis

In order to assess the actual role of ectomycorrhizae in ion uptake by the ectomycorrhizal root system, we used a microelectrode ion flux estimation methodology that provided access to local values of net fluxes. This made it possible to investigate the heterogeneity of ion fluxes along the different types of roots of Pinus pinaster associated or not with ectomycorrhizal species. We compared two fungi able to grow with nitrate in pure culture, Rhizopogon roseolus and Hebeloma cylindrosporum, the former having a positive effect on host tree shoot growth (c. +30%) and the latter a negative effect (c.? 30%). In non‐mycorrhizal plants (control), NO₃^– was taken up at higher rates by the short roots than by the long ones, whereas K⁺ uptake occurred mainly in growing apices of long roots. In mycorrhizal plants, H. cylindrosporum did not modify K⁺ uptake and even decreased NO₃^– uptake at the level of ectomycorrhizal short roots, whereas R. roseolus strongly increased K⁺ and NO₃^– fluxes at the level of ectomycorrhizal short roots without any modification of the fluxes measured along the fungus‐free long roots. The measurement of ion influxes at the surface of the ectomycorrhizal roots can provide a way to reveal actual effects of mycorrhizal association on ion transport in relation to mycorrhizal efficiency in natural conditions.     

Ammonium, nitrate, and proton fluxes along the maize root

Ion-selective microelectrodes were used to measure NH₄⁺, NO₃^– and H⁺ fluxes along the primary root of maize seedlings. Plants were exposed to nutrient solutions containing NH₄⁺, NO₃^– or both ions. Nitrogen fluxes along the root varied substantially among the different treatments. Net NH₄⁺ and NO₃^– uptake and H⁺ extrusion were low at the very apex of the root and generally increased in the more basal regions. In the absence of nitrogen or in the presence of NO₃^– alone, net H⁺ uptake (and root surface alkalinization) occurred at the root tip (0–1 mm), whereas net H⁺ extrusion occurred in all other regions. In the presence of NH₄⁺ alone, a dramatic increase in net H⁺ extrusion was detected in all regions except for the region 6–11 mm from the apex. In contrast, when NO₃^– alone was supplied, net H⁺ extrusion was depressed at all locations except for the tip (0–1 mm). When both NH₄⁺ and NO₃^– were supplied, NO₃^– uptake was suppressed at all locations while net H⁺ extrusion was increased relative to NO₃^– alone. The capacities to absorb NH₄⁺ and NO₃^– at the tip were similar, as indicated by flux rates when NH₄⁺ or NO₃^– were supplied as sole sources, but when supplied together, net NO₃^– uptake was half that of net NH₄⁺ uptake, indicating that NH₄⁺ may satisfy the nitrogen requirements of the poorly vascularized apical tissue in the most energy-efficient way. The high spatial resolution of the measurements enabled us to establish that acidification in the root expansion zone is maintained regardless of nitrogen source.     

Assimilation of nitrate and ammonium by the Scots pine (Pinus sylvestris) seedling under conditions of high nitrogen supply

Seedlings of Scots pine ( Pinus sylvestris L.) were grown on perlite for 21 days under controlled conditions. Apart from the water control, KNO₃ (15 m M ), (NH₄)₂SO₄ (7.5 m M ), and NH₄NO₃ (15 m M ) were offered to study the effects of a high nitrogen supply on nitrogen assimilation. In some experiments 1.3 m M potassium was added to the basic ammonium solutions. In labelling studies nitrate and ammonium were 2.3 atom%¹⁵N-enriched. It was found that over the 21-day period approximately three times more ammonium-N was taken up than nitrate-N. However, nitrate and ammonium, applied simultaneously, were taken up to the same extent as if they were applied separately (additivity). The presence of K⁺ in the medium did not affect N-uptake. Among the soluble N-containing compounds nitrate, ammonium and 8 amino acids were quantified. It was found that assimilation of nitrate can cope with the uptake of NO⁻₃ under all circumstances. Neither free nitrate nor ammonium or amino acids accumulated to an extent exceeding the values of water-grown seedlings. On the other hand, in case of high ammonium supply considerably more nitrogen was taken up than could be incorporated into nonsoluble N-containing substance ('protein'). The remaining nitrogen was found to accumulate in intermediary storage pools (free NH₄⁺, glutamine, asparagine, arginine). Part of this accumulated N could be incorporated into protein when potassium was offered in the nutrient solution. It is concluded that potassium is a requirement for a high rate of protein synthesis not only in crop plants but also in conifers.     

A comparison of NH4+ and NO3– net fluxes along roots of rice and maize

Net fluxes of NH₄⁺ and NO₃^– along adventitious roots of rice ( Oryza sativa L.) and the primary seminal root of maize ( Zea mays L.) were investigated under nonperturbing conditions using ion-selective microelectrodes. The roots of rice contained a layer of sclerenchymatous fibres on the external side of the cortex, whereas this structure was absent in maize. Net uptake of NH₄⁺ was faster than that of NO₃^– at 1 mm behind the apex of both rice and maize roots when these ions were supplied together, each at 0·1 mol m^–3. In rice, NH₄⁺ net uptake declined in the more basal regions, whereas NO₃^– net uptake increased to a maximum at 21 mm behind the apex and then it also declined. Similar patterns of net uptake were observed when NH₄⁺ or NO₃^– was the sole nitrogen source, although the rates of NO₃^– net uptake were faster in the absence of NH₄⁺. In contrast to rice, rates of NH₄⁺ and NO₃^– net uptake in the more basal regions of maize roots were similar to those near the root apex. Hence, the layer of sclerenchymatous fibres may have limited ion absorption in the older regions of rice roots.     

Appearance of nitrate reductase, nitrite reductase and glutamine synthetase in different organs of the Scots pine (Pinus sylvestris) seedling as affected by light, nitrate and ammonium

Appearance of nitrate reductase (NR, EC 1.6.6.1–3), nitrite reductase (NiR, EC 1.7.7.1) and glutamine synthetase (GS, EC 6.3.1.2) under the control of nitrate, ammonium and light was studied in roots, hypocotyls and needles (cotyledonary whorl) of the Scots pine ( Pinus sylvestris L.) seedling. It was found that appearance of NiR was mainly controlled by nitrate whereas appearance of GS was strongly controlled by light. In principle, the NR activity level showed the same dependency on nitrate and light as that of NiR. In the root, both nitrate and ammonium had a stimulatory effect on GS activity whereas in the whorl the induction was minor. The level of NiR (NR) activity is high in the root and hypocotyl and low in the cotyledonary whorl, whereas the GS activity level per organ increases strongly from the root to the whorl. Thus, in any particular organ the operation of the glutamine synthetase/glutamate synthase (GS/GOGAT) cycle is not closely connected to the operation of the nitrate reduction pathway. The strong control of GS/GOGAT by light and the minor sensitivity to induction by nitrate or ammonium indicate a major role of the GS/GOGAT cycle in reassimilation of endogeniously generated ammonium.     

Localization and quantification of net fluxes of H+ along maize roots by combined use of pH-indicator dye videodensitometry and H+-selective microelectrodes

Two methods for measuring proton fluxes along intact maize roots grown with NH ₄ ⁺ or NO ₃ ⁻ at pH 6.5 were compared. Videodensitometric measurement of changes in a pH-indicator dye by video camera was used to map pH around roots and determine the amounts of protons released by various root regions. This method was compared with potentiometric determination of the concentration of H⁺ in the unstirred layer at the root surface using ion-selective microelectrodes. With NH ₄ ⁺ the roots released large amounts of H⁺ in preferential regions where the rate of flux can reach 1.4 or even 2.5 nmol m⁻¹ s⁻¹. Videodensitometry indicated a first region of root acidification in the subapical zone, but this was more difficult to localize with microelectrodes. With NO₃ ⁻ both methods showed that the roots released small amounts of H⁺ and that the apical region took up H⁺ in the first 10 mm then sometimes released H⁺ over the following 10 mm of root. The H⁺ flux profiles obtained by both methods were in good agreement in terms of both order of magnitude of the fluxes and spatial differences along the root. These results suggest that videodensitometry, which is easier to use than potentiometry, can be used to screen different plant species or cultivars under various experimental conditions. The microelectrode technique is indispensable, however, for studying the underlying mechanisms of net H⁺ fluxes. This revised version was published online in June 2006 with corrections to the Cover Date.