Salinity-induced noise in membrane potential of Characeae Chara australis: effect of exogenous melatonin.

Salt sensitive Characeae Chara australis responds to 50 mM NaCl by a prompt appearance of noise in the trans-membrane potential difference (PD). The noise diminishes with time in saline and PD depolarization, leading to altered current-voltage characteristics that could be modeled with H(+)/OH(-) channels. Beilby and Al Khazaaly (JMB 230:21-34, 2009) suggested that the noise might arise from cooperative transient opening of H(+)/OH(-) channels. Presoaking cells in 10 µM melatonin over 24 h abolished the noise in some cells, postponed its appearance in others or changed its characteristics. As melatonin is a very effective antioxidant, we postulated opening of H(+)/OH(-) channels by reactive oxygen species (ROS). Measurement of ROS using dihydrodichlorofluorescein diacetate confirmed substantial reduction in ROS production in melatonin-treated cells in saline and sorbitol media. However, ROS concentration decreased as a function of time in saline medium. Possible schemes for activation of H(+)/OH(-) channels under salinity stress are considered.

Zinc ions block H⁺/OH⁻ channels in Chara australis.

Chara australis cells exposed to media of pH 10 and above exhibit high conductance, arising from the opening of H⁺/OH⁻ channels in the plasma membrane. This high conductance can be totally inhibited by 1.0 mm ZnCl2 and restored by 0.5 mm 2-mercaptoethanol (ME). Important for carbon fixation, H⁺/OH⁻ channels play a key role in cell pH banding. Banding was also shown to be abolished by 1.0 mm ZnCl2 and restored in some cells by ME. The proton pump is also involved in banding, but was little affected by ZnCl2 over the periods needed for the inhibition of H⁺/OH⁻ channels. Previously, we postulated that H⁺/OH⁻ channels open transiently at the onset of saline stress in salt-sensitive C. australis, causing membrane potential difference (PD) noise; and remain open in latter stages of saline stress, contributing to cell deterioration. ZnCl2 totally inhibited the saline noise and the upwardly concave I/V characteristics associated with the putative H⁺/OH⁻ currents. Again, ME reversed both these effects. We discuss the mode of action of zinc ions and ME with reference to animal voltage-gated H⁺ channels and water channels.

The role of H(+)/OH(-) channels in the salt stress response of Chara australis.

We investigate the electrophysiological salt stress response of the salt-sensitive charophyte Chara australis as a function of time in saline artificial pond water (saline APW) containing 50 mM NaCl and 0.1 mM CaCl(2). The effects are due to an increase in Na(+) concentration rather than an increase in Cl(-) concentration or medium osmolarity. A previous paper (Shepherd et al. Plant Cell Environ 31:1575-1591, 2008) described the rise in the background conductance and inhibition of proton pumping in saline APW in the first 60 min. Here we investigate the shift of membrane potential difference (PD) to levels above -100 mV and the change of shape of the current-voltage (I/V) profiles to upwardly concave. Arguing from thermodynamics, the I/V characteristics can be modeled by channels that conduct H(+) or OH(-). OH(-) was chosen, as H(+) required an unrealistic increase in the number/permeability of the channels at higher pH levels. Prolonged exposure to saline APW stimulated opening of more OH(-) channels. Recovery was still possible even at a PD near -50 mV, with partial return of proton pumping and a decrease in OH(-) current following APW wash. Upon change of pH from 7 to 9, the response was consistent with previously observed I/V characteristics of OH(-) channels. For a pH change to 6, the response was transient before channel closure but could still be modeled. The consequences of opening of H(+) or OH(-) channels while the cell is under salt stress are discussed.

Membrane potential fluctuations in Chara australis: a characteristic signature of high external sodium.

We have studied fluctuations in membrane PD in Chara australis at frequencies between 1 and 500 mHz, by classical noise analysis and by inspection of the PD time-course. The former shows (1) a quasi-Lorentzian (1/f (2)) rise of noise power as frequency falls, and (2) a marked increase in noise power when the cell is exposed to high salinity (Chara australis is a salt-sensitive species). The latter shows that, as well as initiating depolarization, exposure to 50 mM Na as either chloride or sulfate usually initiates a continuous but random series of small depolarizations which gives rise to the increase in noise and whose mechanism is discussed.

Mechano-perception in Chara cells: the influence of salinity and calcium on touch-activated receptor potentials, action potentials and ion transport.

This paper investigates the impact of increased salinity on touch-induced receptor and action potentials of Chara internodal cells. We resolved underlying changes in ion transport by current/voltage analysis. In a saline medium with a low Ca(2+) ion concentration [(Ca(2+))(ext)], the cell background conductance significantly increased and proton pump currents declined to negligible levels, depolarizing the membrane potential difference (PD) to the excitation threshold [action potential (AP)(threshold)]. The onset of spontaneous repetitive action potentials further depolarized the PD, activating K(+) outward rectifying (KOR) channels. K(+) efflux was then sustained and irrevocable, and cells were desensitized to touch. However, when [Ca(2+)](ext) was high, the background conductance increased to a lesser extent and proton pump currents were stimulated, establishing a PD narrowly negative to AP(threshold). Cells did not spontaneously fire, but became hypersensitive to touch. Even slight touch stimulus induced an action potential and further repetitive firing. The duration of each excitation was extended when [Ca(2+)](ext) was low. Cell viability was prolonged in the absence of touch stimulus. Chara cells eventually depolarize and die in the saline media, but touch-stimulated and spontaneous excitation accelerates the process in a Ca(2+)-dependent manner. Our results have broad implications for understanding the interactions between mechano-perception and salinity stress in plants.

Exogenous melatonin affects photosynthesis in characeae Chara australis.

Melatonin was found in the fresh water characeae Chara australis. The concentrations (~4 µg/g of tissue) were similar in photosynthesizing cells, independent of their position on the plant and rhizoids (roots) without chloroplasts. Exogenous melatonin, added at 10 µM to the artificial pond water, increased quantum yield of photochemistry of photosystem II by 34%. The increased efficiency appears to be due to the amount of open reaction centers of photosystem II, rather than increased efficiency of each reaction center. More open reaction centers reflect better functionality of all photosynthetic transport chain constituents. We suggest that melatonin protection against reactive oxygen species covers not only chlorophyll, but also photosynthetic proteins in general.