Calcium transport in gill cells of Ucides cordatus, a mangrove crab living in variable salinity environments

Abstract

Crustaceans show discontinuous growth and have been used as a model system for studying cellular mechanisms of calcium transport, which is the main mineral found in their exoskeleton. Ucides cordatus, a mangrove crab, is naturally exposed to fluctuations in calcium and salinity. To study calcium transport in this species during isosmotic conditions, dissociated gill cells were marked with fluo-3 and intracellular Ca2+ change was followed by adding extracellular Ca2+ as CaCl2 (0, 0.1, 0.25, 0.50, 1.0 and 5mM), together with different inhibitors. For control gill cells, Ca2+ transport followed Michaelis–Menten kinetics with Vmax=0.137±0.001 ∆Ca2+i (μM×22.104cells−1×180s−1; N=4; r2=0.99); Km=0.989±0.027mM. The use of different inhibitors for gill cells showed that amiloride (Na+/Ca2+ exchange inhibitor) inhibited 80% of Ca2+ transport in gill cells (Vmax). KB-R, an inhibitor of Ca influx in vertebrates, similarly caused a decrease in Ca2+ transport and verapamil (Ca2+ channel inhibitor) had no effect on Ca2+ transport, while nifedipine (another Ca2+ channel inhibitor) caused a 20% decrease in Ca2+ affinity compared to control values. Ouabain, on the other hand, caused no change in Ca2+ transport, while vanadate increased the concentration of intracellular calcium through inhibition of Ca2+ efflux probably through the plasma membrane Ca2+-ATPase. Results show that transport kinetics for Ca2+ in these crabs under isosmotic conditions is lower compared to a hyper-regulator freshwater crab Dilocarcinus pagei studied earlier using fluorescent Ca2+ probes. These kinds of studies will help understanding the comparative mechanisms underlying the evolution of Ca transport in crabs living in different environments.