Energetics of acclimation to NaCl by submerged, anoxic rice seedlings

Abstract

Background and aims: Our aim was to elucidate how plant tissues under a severe energy crisis cope with imposition of high NaCl, which greatly increases ion fluxes and hence energy demands. The energy requirements for ion regulation during combined salinity and anoxia were assessed to gain insights into ion transport processes in the anoxia-tolerant coleoptile of rice.

Methods: We studied the combined effects of anoxia plus 50 or 100 mm NaCl on tissue ions and growth of submerged rice (Oryza sativa) seedlings. Excised coleoptiles allowed measurements in aerated or anoxic conditions of ion net fluxes and O₂ consumption or ethanol formation and by inference energy production.

Key results: Over 80 h of anoxia, coleoptiles of submerged intact seedlings grew at 100 mm NaCl, but excised coleoptiles, with 50 mm exogenous glucose, survived only at 50 mm NaCl, possibly due to lower energy production with glucose than for intact coleoptiles with sucrose as substrate. Rates of net uptake of Na⁺ and Cl^- by coleoptiles in anoxia were about half those in aerated solution. Ethanol formation in anoxia and O₂ uptake in aerobic solution were each increased by 13-15 % at 50 mm NaCl, i.e. ATP formation was stimulated. For acclimation to 50 mm NaCl, the anoxic tissues used only 25 % of the energy that was expended by aerobic tissues. Following return of coleoptiles to aerated non-saline solution, rates of net K⁺ uptake recovered to those in continuously aerated solution, demonstrating there was little injury during anoxia with 50 mm NaCl.

Conclusion: Rice seedlings survive anoxia, without the coleoptile incurring significant injury, even with the additional energy demands imposed by NaCl (100 mm when intact, 50 mm when excised). Energy savings were achieved in saline anoxia by less coleoptile growth, reduced ion fluxes as compared to aerobic coleoptiles and apparent energy-economic ion transport systems.

Keywords: Anoxia tolerance; NaCl ×; Oryza sativa; anaerobic catabolism; anoxia interaction; anoxia plus NaCl; coleoptile; complete submergence; energetics; energy crisis; energy efficient transport; ethanolic fermentation; germination; salinity tolerance.

Fig. 1.

Fresh weight increment (mg g⁻¹ f. wt d⁻¹) of coleoptiles of rice seedlings and excised coleoptile tips between 0 and 72 h of exposure to 50 and 100 mm NaCl in anoxia. Caryopses were germinated in aerated solution and following a hypoxic pretreatment seedlings were transferred to anoxia at 66 h after imbibition. NaCl was increased 18 h after the start of anoxia by steps of 50 mm NaCl. Coleoptile fresh weight at start of anoxia was 2·9 mg. Data are means ± s.e., n = 3. Different letters indicate significant differences at P < 0·05. Data from Experiments 1 and 2.

Fig. 2.

K⁺ net uptake or loss in excised coleoptile tips at 0·3 and 50 mm NaCl, when NaCl was given 18 h after starting anoxia, shown as time zero on the horizontal axis (Experiment 3). K⁺ was at 0·3 mm in the medium during salinity and exogenous glucose was at 50 mm in anoxia and 5 mm in aerated solution. The reaeration after exposure to 50 mm NaCl and anoxia was carried out using non-saline solutions either without or with iso-osmotic mannitol (key for symbols in panel). Data points are plotted at the end of each sampling period for which K⁺ in the medium was measured. Data are means ± s.e., n = 3.

Fig. 3.

Na⁺ (A), Cl⁻ (B) and K⁺ (C) concentrations of excised rice coleoptile tips after reaeration in non-saline solution following prior exposure to NaCl treatments (0·3 or 50 mm) in anoxia for 90 h (Experiment 3). The reaeration started at time 0 on the horizontal axis and for coleoptile tips previously at 50 mm NaCl the recovery phase was carried out either without or with iso-osmotic mannitol (see symbols in the panel). Asterisks indicate significant differences (P < 0·05) between treatments at a sampling time. Data are means ± s.e., n = 3.

Fig. 4.

Time-courses of ethanol formation in anoxia as affected by 50 mm NaCl by excised coleoptile tips (A) and time-course of O₂ consumption in air-equilibrium solution by excised rice coleoptile tips (B) (Experiment 5). Tissues in A had been in anoxia for 18 h before NaCl was increased to 50 mm (time 0 on the horizontal axis is when NaCl was first added). Coleoptile tips in B were excised from seedlings that had initially grown in aerated solution and at 36 h after imbibition were given a 30-h hypoxic pre-treatment, after which the coleoptile tips were excised, healed for 5 h in hypoxia and transferred to stirred cuvettes with air-equilibrium solution and periodically closed and O₂ consumption monitored with a Clark-type electrode. Asterisks indicate significant differences (P < 0·05) between treatments at a sampling time. Data are means ± s.e., n = 3.

Fig. 5.

Na⁺, Cl⁻ and K⁺ concentrations (mm, tissue water basis) in the coleoptile (left: A, C, E) and endosperm (right: B, D, F) of seedlings exposed to 50 and 100 mm NaCl in anoxia (Experiment 4). At 36 h after imbibition and aerated conditions, the seedlings were in a hypoxic pre-treatment for 30 h. The seedlings were then transferred to anoxia and after 18 h of anoxia there was a first step up to 50 mm NaCl (time 0 on the horizontal axis) and for the 100 mm NaCl treatment a second step up after another 24 h (time indicated by dashed arrow). Asterisks indicate significant differences (P < 0·05) between treatments at a sampling time. Data are means ± s.e., n = 4.

Fig. 6.

Na⁺ (A), Cl⁻ (B) and K⁺ (C) concentrations in excised rice coleoptile tips at 0·3 and 50 mm NaCl, during both anoxia and aeration (Experiment 5). At 36 h after imbibition and aerated conditions, the seedlings were in a hypoxic pretreatment for 30 h, and then excised coleoptile tips were healed for 5 h in hypoxic conditions and then transferred to anoxia. After 18 h of anoxia, NaCl treatments were imposed. Exogenous K⁺ was 0·3 mm and exogenous glucose at 50 mm in anoxia and 5 mm in aerated solution. Asterisks indicate significant differences (P < 0·05) between treatments at a sampling time. Data are means ± s.e., n = 3.

Fig. 7.

Net uptake rates of Na⁺ (A) and Cl⁻ (B) by excised rice coleoptile tips during 50 mm NaCl for 90 h in aerated or in anoxic solution, calculated based on increases in tissue Na⁺ and Cl⁻ concentrations between each sequence of sampling times (data from mean values in Fig. 6; Experiment 5). Data points are plotted in the middle of each period between sampling times of tissues.