Measurements of unidirectional calcium fluxes in stripped intestinal epithelium of the tilapia, Oreochromis mossambicus, in the presence of ouabain or in the absence of sodium indicated that calcium absorption via the fish intestine is sodium dependent. Active Ca2+ transport mechanisms in the enterocyte plasma membrane were analyzed. The maximum capacity of the ATP-dependent Ca2+ pump (Vm: 0.63 nmol.min-1.mg-1, Km:27 nM Ca2+) is calculated to be 2.17 nmol.min-1.mg-1, correcting for 29% inside-out oriented vesicles in the membrane preparation. The maximum capacity of the Na+/Ca2+ exchanger with high affinity for Ca2+ (Vm:7.2 nmol.min-1.mg-1, Km:181 nM Ca2+) is calculated to be 13.6 nmol.min-1.mg-1, correcting for 53% resealed vesicles and assuming symmetrical behavior of the Na+/Ca2+ exchanger. The high affinity for Ca2+ and the sixfold higher capacity of the exchanger compared to the ATPase suggest strongly that the Na+/Ca2+ exchanger will contribute substantially to Ca2+ extrusion in the fish enterocyte. Further evidence for an important contribution of Na+/Ca2+ exchange to Ca2+ extrusion was obtained from studies in which the simultaneous operation of ATP- and Na(+)-gradient-driven Ca2+ pumps in inside-out vesicles was evaluated. The fish enterocyte appears to present a model for a Ca2+ transporting cell, in which Na+/Ca2+ exchange activity with high affinity for Ca2+ extrudes Ca2+ from the cell.

The tilapia, Oreochromis mossambicus, is a truly eurybaline species in that it lives, grows, a n d reproduces in freshwater as well as in full-strengtb seawater. The gills, intestine, a n d kidneys show ionoregulatory adaptations fu n d a m en ta l fo r the calcium balance o f this fish in these vastly different ionic media. This re view focuses on calcium flows in these ionoregulatory organs a n d the changes that occur in theThe kidney offreshw ater fishes produces a typical dilute a n d hypocalcic urine; in seawater, urine production decreases a n d the urine calcium concen tration is always higher than that o f the plasma. Exchange activity ofN a* a n d Ca2* is undetectably low or absent in renal tissue plasm a membranes. However, a high-ajfinity, high-capacity Ca2*-ATPase activity appears to correlate with Ca2*

The inhibition of Ca(2+)-ATPase, (Na+ + K+)-ATPase and Na+/Ca2+ exchange by Cd2+ was studied in fish intestinal basolateral plasma membrane preparations. ATP driven 45Ca2+ uptake into inside-out membrane vesicles displayed a Km for Ca2+ of 88 +/- 17 nM, and was extremely sensitive to Cd2+ with an IC50 of 8.2 +/- 3.0 pM Cd2+, indicating an inhibition via the Ca2+ site. (Na+ + K+)-ATPase activity was half-maximally inhibited by micromolar amounts of Cd2+, displaying an IC50 of 2.6 +/- 0.6 microM Cd2+. Cd2+ ions apparently compete for the Mg2+ site of the (Na+ + K+)-ATPase. The Na+/Ca2+ exchanger was inhibited by Cd2+ with an IC50 of 73 +/- 11 nM. Cd2+ is a competitive inhibitor of the exchanger via an interaction with the Ca2+ site (Ki = 11 nM). Bepridil, a Na+ site specific inhibitor of Na+/Ca2+ exchange, induced an additional inhibition, but did not change the Ki of Cd2+. Also, Cd2+ is exchanged against Ca2+, albeit to a lesser extent than Ca2+. The exchanger is only partly blocked by the binding of Cd2+. In vivo cadmium that has entered the enterocyte may be shuttled across the basolateral plasma membrane by the Na+/Ca2+ exchanger. We conclude that intracellular Cd2+ ions will inhibit plasma membrane proteins predominantly via a specific interaction with divalent metal ion sites.

The euryhaline tilapia Oreochromis mossambicus kept in fresh water takes up cal cium mainly from the water via the gills, like other freshwater fish. In the gills specialized mitochondria-rich cells, the chloride cells, are thought to mediate a transcellular Ca2+ transport. Second messenger operated calcium channels (SMOCs) in the apical membrane, regulated by the hormone stanniocalcin, allow minute-to-minute control over the entry of Ca2 +. In the basolateral plasma membranes of these cells, an ATP-consuming Ca2+ transporting enzyme provides the major driving force for extrusion of Ca2+ into the blood; in addition, an Na + /Ca2+ exchanger is present in these membranes. The kinetics of the exchanger in vitro indicate that this extrusion mechanism dominates when intracellular calcium levels reach micromolar levels. In the gills, the transport of calcium appears primarily ATPase mediated and therefore largely independent of the Na + status of the transporting cell. In enterocytes, similar mechanisms for transcellular transport of Ca2 + exist. However, in the intestinal epithelium the extrusion of Ca2 + is primarily via the Na + /Ca2 + exchanger and to a very limited extent mediated via the Ca2 +-ATPase. Indeed, calcium transport over the intestinal epithelium is dependent on the Na +-status and the N a+/K +-ATPase activity of the epi thelium. Prolactin and cortisol are endocrine factors determining the relative densities of calcium pumps in basolateral plasma membranes.At least 80% of the magnesium required for growth and homeostasis is absorbed from the food via the intestine. Magnesium is transported transcellularly and actively via enterocytes. The movement of Mg2 + over the apical membrane is passive, down an electrochemical gradient. The cytosolic Mg2 + concentration is kept well below its equilibrium concentration. The extrusion over the basolateral plasma membrane is mediated by an ATP-consuming enzyme. The gills contribute less than 20% to magnesium uptake, but up to 50% in tilapia fed a magnesium deficient diet. Evidence is accruing that prolactin is involved in the adaptation to low magnesium diets.