Ca2+ versus Zn2+ transport in the gills of freshwater rainbow trout and the cost of adaptation to waterborne Zn2+

Abstract

Previous work suggested that Ca2+ and Zn2+ share a common uptake pathway in rainbow trout gills. We here report on relationships between the kinetic variables for unidirectional Ca2+ influx and unidirectional Zn2+ influx during a 1 month exposure of freshwater rainbow trout to Zn2+ (150 µg l-1=2.3 µmol l-1 as total zinc, Zn). Initial exposure to Zn2+ caused a large competitive inhibition of Ca2+ influx, as indicated by a threefold increase in apparent Km for Ca2+ (measured in the presence of Zn2+). There was also a smaller non-competitive inhibition (50 % decrease in Jmax) of the Ca2+ transport system, which was abolished after 1­2 weeks of exposure. The Km, measured in the absence of Zn2+, decreased dramatically (i.e. elevated affinity) on days 1­4 but increased thereafter; both true and apparent Km finally stabilized significantly above control levels. However, the Km values for Ca2+ (<200 µmol l-1) were low relative to the Ca2+ level in the water (1000 µmol l-1), and therefore the changes did not influence the actual Ca2+ influx of the fish, which tracked Jmax. In contrast, water [Zn2+] (2.3 µmol l-1 as total Zn) was close to the reported apparent Km (3.7 µmol l-1) for Zn2+ influx in the presence of 1000 µmol l-1 Ca2+. Unidirectional Zn2+ influx increased during the first week of exposure to waterborne Zn2+, followed by a persistent reduction to about 50 % of control levels, effects that may be largely explained by the observed changes in true Km for Ca2+. We speculate that the initial response of the fish to elevated [Zn2+] is to compensate for a reduced availability of Ca2+ by markedly increasing the affinity of a dual Ca2+/Zn2+ transporter. Once the Ca2+ influx is 'corrected' by restoration of functional transport sites (Jmax), the system is tuned to limit the influx of Zn2+ by a persistent reduction in the affinities for both ions. The changes in influx characteristics for Ca2+ and Zn2+ were correlated with internal physiological alterations indicative of adaptation to Zn2+ and increased metabolic cost. Depressed plasma [Ca] was corrected within 1 week, and there were no effects on whole-body [Ca] or [Zn]. A slight accumulation of Zn in the gills was associated with increased branchial metallothionein levels. Rates of protein synthesis and degradation in the gills were initially increased and whole-body growth was transiently impaired, effects which were reversed after 18 days of exposure. Sublethal challenge with Zn2+ (at 450 µg l-1=6.9 µmol l-1 as total Zn) always depressed plasma [Ca] in control fish, but by 1 month of exposure to Zn2+ at 150 µg l-1 (as total Zn), experimental fish were resistant to challenge. However, the fish did not acquire increased survival tolerance (LT50) to a lethal concentration of Zn2+ (4 mg l-1=61 µmol l-1 as total Zn).