Silver ions disrupt K+ homeostasis and cellular integrity in intact barley (Hordeum vulgare L.) roots

Abstract

The heavy metals silver, gold, and mercury can strongly inhibit aquaporin-mediated water flow across plant cell membranes, but critical examinations of their side effects are rare. Here, the short-lived radiotracer 42K is used to demonstrate that these metals, especially silver, profoundly change potassium homeostasis in roots of intact barley (Hordeum vulgare L.) plants, by altering unidirectional K+ fluxes. Doses as low as 5 μM AgNO3 rapidly reduced K+ influx to 5% that of controls, and brought about pronounced and immediate increases in K+ efflux, while higher doses of Au3+ and Hg2+ were required to produce similar responses. Reduced influx and enhanced efflux of K+ resulted in a net loss of >40% of root tissue K+ during a 15 min application of 500 μM AgNO3, comprising the entire cytosolic potassium pool and about a third of the vacuolar pool. Silver also brought about major losses of UV-absorbing compounds, total electrolytes, and NH4+. Co-application, with silver, of the channel blockers Cs+, TEA+, or Ca2+, did not affect the enhanced efflux, ruling out the involvement of outwardly rectifying ion channels. Taken together with an examination of propidium iodide staining under confocal microscopy, the results indicate that silver ions affect K+ homeostasis by directly inhibiting K+ influx at lower concentrations, and indirectly inhibiting K+ influx and enhancing K+ efflux, via membrane destruction, at higher concentrations. Ni2+, Cd2+, and Pb2+, three heavy metals not generally known to affect aquaporins, did not enhance K+ efflux or cause propidium iodide incorporation. The study reveals strong and previously unknown effects of major aquaporin inhibitors and recommends caution in their application.

Rapid Ammonia Gas Transport Accounts for Futile Transmembrane Cycling under NH3/NH4 + Toxicity in Plant Roots

“…3B). We have previously shown that sudden exposure to Ag + causes extensive damage to both major membrane systems (plasmalemma and tonoplast) in barley roots (Coskun et al, 2012). By contrast, we observed no effect of mercury (Hg 2+ ) application on tracer release (Fig.…”

“…Importantly, this qualifies the use of Hg 2+ as a potential inhibitor of AQPs (see below). With respect to the Ag + -induced stimulation in tracer efflux, this effect allowed for quantification of released substrate (in terms of mmol g -1 root fresh weight) via integration of the 13 N loss and estimated intracellular specific activity, as shown previously (Coskun et al, 2012 ; Fig. 3D).…”

“…A subset of experiments involved either chemical (pH 7.25, 8.25, and 9.25 or complete NH 3 /NH 4 + withdrawal) or cold (4°C) treatment for the final 5 min of elution (see "Results"). Another subset involved a longer elution protocol (30 min; Coskun et al, 2012) and the sudden (at 15 min) application of 500 mM AgNO 3 to disrupt membranes and release tracer accumulated in the cell (Coskun et al, 2012). Following elution, roots were handled as above, and radioactivity in roots, shoots, and efflux eluates were counted.…”

“…To quantify the chemical amount of released NH 3 /NH 4 + during Ag + application (above), an integration technique was employed as described in detail elsewhere (Coskun et al, 2012). In brief, the summation of radioactivity released (in counts per min) during Ag + treatment was divided by the internal specific activity at the time of Ag + application (taking into account the exponential rise in specific activity during loading and its decline during elution up to the time of Ag + application) and corrected for root fresh weight.…”

Boron deprivation immediately causes cell death in growing roots ofArabidopsis thaliana(L.) Heynh.

“…4b). Thus, Ag + perturbed the immediate B-deprivation responses of roots observed in the elongation zone, although this treatment might have had some adverse side effects on root cell viability, as suggested in another study on barley (Coskun et al 2012). These results suggested that ethylene mediated the early responses of A. thaliana roots to B deprivation, probably upstream of ROS production.…”