Polyamines improve K+/Na+ homeostasis in barley seedlings by regulating root ion channel activities

Abstract

Polyamines are known to increase in plant cells in response to a variety of stress conditions. However, the physiological roles of elevated polyamines are not understood well. Here we investigated the effects of polyamines on ion channel activities by applying patch-clamp techniques to protoplasts derived from barley (Hordeum vulgare) seedling root cells. Extracellular application of polyamines significantly blocked the inward Na(+) and K(+) currents (especially Na(+) currents) in root epidermal and cortical cells. These blocking effects of polyamines were increased with increasing polycation charge. In root xylem parenchyma, the inward K(+) currents were blocked by extracellular spermidine, while the outward K(+) currents were enhanced. At the whole-plant level, the root K(+) content, as well as the root and shoot Na(+) levels, was decreased significantly by exogenous spermidine. Together, by restricting Na(+) influx into roots and by preventing K(+) loss from shoots, polyamines were shown to improve K(+)/Na(+) homeostasis in barley seedlings. It is reasonable to propose that, therefore, elevated polyamines under salt stress should be a self-protecting response for plants to combat detrimental consequences resulted from imbalance of Na(+) and K(+).

Figure 1.

Effect of exogenous spermidine on Na⁺ and K⁺ distribution in barley seedlings. A, Na⁺ and K⁺ contents in the roots and shoots of barley seedlings. B, [K⁺]/[Na⁺] in the roots and shoots of barley seedlings. C, Shoot [K⁺]/[Na⁺] was enhanced by exogenous spermidine in a dose-dependent manner. The applied spermidine concentration was 0.5 mm. Spd in A is an abbreviated form of spermidine. Asterisks indicate the difference at P < 0.05 (*) or P < 0.01 (**) by Student's t test. Data represent mean ± sd of three independent experiments.

Figure 2.

Effects of extracellular and intracellular spermidine on the inward K⁺ currents in root epidermal cells. A, During the recordings, the holding potential was −52 mV, the currents were recorded at the membrane potentials from −190 to 30 mV with increment of 20 mV. B, Whole-cell inward K⁺ currents (capacitance = 6.5 pF). C, The same cell as in B was treated with 1 mm spermidine in the bath solution for 15 min. D, The current amplitudes (mean ± sd) from control cells (▪) and cells treated with 1 mm spermidine in the bath solution (▵) are presented as I-IV curves (n = 15). E, Whole-cell inward K⁺ currents (capacitance = 5.4 pF). F, The same cell as in E was treated with 1 mm spermidine in the pipette solution (capacitance = 6.3 pF). G, The current amplitudes (mean ± sd) from 21 control cells (▪) and 25 cells treated with 1 mm spermidine in the pipette solution (▵) are presented as I-V curves.

Figure 3.

Characteristics of the Na⁺-conducted currents in root epidermal cells. A, The dependence of instantaneous Na⁺ currents in root epidermal cells on external NaCl concentration. Concentrations are given in mm. Na⁺ was present as the chloride salt. Arrows indicate E_rev. The data represent mean ± sd. B, I-V relations of inward instantaneous currents in root epidermal cells treated with extracellular application of Ca²⁺, TEA⁺, and quinine. The current amplitudes from control cells (♦, n = 15), Ca²⁺-treated cells (○, n = 15), TEA⁺-treated cells (▵, n = 18), and quinine-treated cells (▴, n = 21) are presented as I-V curves. Concentrations are indicated in mm. Na⁺ was present as the Na gluconate. Data were obtained after 10-min exposure to Ca²⁺, TEACl, or quinine. The data represent mean ± sd.

Figure 4.

Effects of extracellular and intracellular spermidine on the inward Na⁺ currents in root epidermal cells. A, Plasma membrane currents were recorded with voltage pulses ranging from −170 to 70 mV at 20 mV increments. Holding potential was kept at −52 mV. B, Whole-cell inward Na⁺ currents (capacitance = 4.5 pF). C, The same cell as in B was treated with 1 mm spermidine in the bath solution for 15 min. D, The current amplitudes (mean ± sd) from control cells (▪) and cells treated with 1 mm spermidine in the bath solution (▵) are presented as I-V curves (n = 16). E, Whole-cell inward Na⁺ currents (capacitance = 6.5 pF). F, The same cell as in E was treated with 1 mm spermidine in the pipette solution (capacitance = 5.7 pF). G, The current amplitudes (mean ± sd) from 14 control cells (▪) and 23 cells treated with 1 mm spermidine in the pipette solution (▵) are presented as I-V curves.

Figure 5.

Effects of natural polyamines on the inward Na⁺ and K⁺ currents in root epidermal cells. A, The responses of inward Na⁺ current to different polyamines in the bath solution are shown as I-V curves (control, n = 13; putrescine, n = 15; spermidine, n = 17; spermine, n = 19). Each polyamine concentration was 1 mm. The data were presented as mean ± sd. B, The responses of inward K⁺ current to different polyamines in the bath solution are shown as I-V curves (control, n = 16; putrescine, n = 14; spermidine, n = 18; spermine, n = 17).

Figure 6.

Extracellular spermidine blocked the inward K⁺ currents in root cortical cells. A, The holding potential was −52 mV and the currents were recorded at the membrane potentials from −190 to 50 mV with increments of 20 mV. B, Whole-cell inward K⁺ currents (capacitance = 15.5 pF). C, The same cell as in B was treated with 1 mm spermidine in the bath solution for 15 min. D, The current amplitudes (mean ± sd) from control cells (▪) and cells treated with 1 mm spermidine in the bath solution (▵) are presented as I-V curves (n = 17).

Figure 7.

Extracellular spermidine blocked the inward Na⁺ currents in root cortical cells. A, The currents were recorded at the membrane potentials from −170 to 70 mV (in 20 mV steps) with the holding potential of −52 mV. B, Whole-cell inward Na⁺ currents (capacitance = 18 pF). C, The same cell as in B was treated with 1 mm spermidine in the bath solution for 15 min. D, The current amplitudes (mean ± sd) from control cells (▪) and cells treated with 1 mm spermidine in the bath solution (▵) are presented as I-V curves (n = 18).

Figure 8.

Effect of extracellular spermidine on the outward K⁺ currents in root xylem parenchyma. A, The membrane potential was clamped at −52 mV and stepped to values from −90 to 130 mV with 20 mV increments. B, Whole-cell inward K⁺ currents (capacitance = 7.2 pF). C, The same cell as in B was treated with 1 mm spermidine in the bath solution for 15 min. D, The current amplitudes (mean ± sd) from control cells (▪) and cells treated with 1 mm spermidine in the bath solution (▵) are presented as I-V curves (n = 14).

Figure 9.

Effect of extracellular spermidine on the inward K⁺ currents in root xylem parenchyma. A, During the recordings, the holding potential was −52 mV, the currents were recorded at the membrane potentials from −190 to 90 mV with increments of 20 mV. B, Whole-cell inward K⁺ currents (capacitance = 9 pF). C, The same cell as in B was treated with 1 mm spermidine in the bath solution for 15 min. D, The current amplitudes (mean ± sd) from control cells (▪) and cells treated with 1 mm spermidine in the bath solution (▵) are presented as I-V curves (n = 18).

Figure 10.

Effect of extracellular spermidine on the outward Na⁺ currents in root xylem parenchyma cells. A, During the recordings, the holding potential was −52 mV, the currents were recorded at the membrane potentials from −130 to 110 mV with increments of 20 mV. B, Whole-cell outward Na⁺ currents (capacitance = 8.6 pF). C, The same cell as in B was treated with 1 mm spermidine in the bath solution for 15 min. D, The current amplitudes (mean ± sd) from control cells (▪) and cells treated with 1 mm spermidine in the bath solution (▵) are presented as I-V curves (n = 15).