Rapid effects of nitrogen form on leaf morphogenesis in tobacco

Ammonium (NH4+) instead of nitrate (NO3-) as the nitrogen (N) source for to bacco (Nicotiana tabacum L.) cultivated in a pH-buffered nutrient solution resulted in decreased shoot and root biomass, Reduction of shoot fresh weig ht was mainly related to inhibition of leaf growth, which was already detec table after short-term NH4+ treatments of 24 h, and even at a moderate conc entration level of 2 mM, Microscopic analysis of the epidermis of fully exp anded leaves revealed a decrease in cell number (50%) and in cell size (30% ) indicating that both cell division and cell elongation were affected by N H4+ application. Changes in various physiological parameters known to be as sociated with NH4+-induced growth depression were examined both in long-ter m and short-term experiments: the concentrations of total N, soluble sugars and starch as well as the osmotic potential, the apparent hydraulic conduc tivity and the rate of water uptake were not reduced by NH4+ treatments (du ration 1-12 d), suggesting that leaf growth was neither limited by the avai lability of N and carbohydrates, nor by a lack of osmotica or water supply. Although the concentration of K+ in leaf press sap declined in expanding l eaves by approximately 15% in response to NH4+ nutrition, limitation of min eral nutrients seems to be unlikely in view of the fast response of leaf gr owth at 24 h after the start of the NH4+ treatment. No inhibitory effects w ere observed when NH4+ and NO3- were applied simultaneously (each 1 mM) res ulting in a NO3-/NH4+ net uptake ratio of 6 : 4. These findings suggest tha t the rapid inhibition of leaf growth was not primarily related to NH4+ tox icity, but to the lack of NO3--supply, Growth inhibition of plants fed sole ly with NH4+ was associated with a 60% reduction of the zeatine + zeatine r iboside (Z + ZR) cytokinin fraction in the xylem sap after 24 h, Furthermor e Z + ZR levels declined to almost zero within the next 4 d after start of the NH4+ treatment. In contrast, the concentrations of the putative Z + ZR precursors isopentenyl-adenine and isopentenyl-adenosine (i-Ade + i-Ado) we re not affected by NH4+ application. Since cytokinins are involved in the r egulation of both cell division and cell elongation, it seems likely that t he presence of NO3- is required to maintain biosynthesis and/or root to sho ot transfer of cytokinins at a level that is sufficient to mediate normal l eaf morphogenesis.