Ocean acidification: effects of pH on Ca uptake by lobster branchiostegites.

Gill chambers of the Atlantic lobster, Homarus americanus, possess three structures that are involved with respiration and ion regulation: gill filaments, epipodites, and branchiostegites. This paper describes ion transport mechanisms present in the plasma membranes of branchiostegite epithelial cells and the effects of pH on the uptake of Ca by these processes. Partially purified membrane vesicles (PPMV) of branchiostegite cells were produced by a homogenization/centrifugation method that has previously been used to define ion transport processes in both crab and lobster gill tissues. In the present study, lobster branchiostegite PPMV Ca uptake was highest at pH 8.5 and lowest at pH values between 6.0 and 7.0 (p < 0.02). At pH 8.0, Ca uptake was a biphasic process consisting of a saturable process at low [Ca] and a linear process at higher [Ca]. At pH 6.0, Ca uptake was only a linear process and paralleled linear uptake at pH 8.0. A valinomycin/K-induced membrane potential (PD, inside negative) doubled Ca uptake at pH 7.0 above that in the absence of a PD (p < 0.05). An induced PD at pH 8.0 did not significantly (p > 0.05) affect Ca uptake observed in the absence of a PD, but was threefold greater than uptake at pH 7.0 in the absence of a PD (p < 0.05). Amiloride (2 mM) did not affect Ca uptake at pH 8.0, but 2 mM amiloride + 100 µM verapamil reduced uptake by approximately 50%. In the presence of both 2 mM amiloride + 100 µM verapamil, 15 s Ca influx at pH 8.5 was a hyperbolic function of [Ca] (0.1-5 mM) (K = 4.2 ± 0.3 mM; J = 9792 ± 439 pmol/mg protein × 15 s). Ca influxes at pH 7.5 under the same conditions were also hyperbolic with K = 8.3 ± 1.4 mM; J = 10732 ± 1250 pmol/mg protein × 15 s. K values were significantly different (p < 0.05), but J values were not (p > 0.05). These results suggest that Ca uptake by lobster branchiostegites may have occurred by the combination of diffusion through a verapamil-inhibited calcium channel and carrier-mediated transport by amiloride-insensitive, electroneutral, 1Ca/2H antiporters. Decreased pH, as might occur during ocean acidification, did not appear to modify calcium diffusion through the channels, but protons acted as competitive inhibitors of calcium transport by carrier-mediated antiport. Decreased calcium uptake with continued ocean acidification may significantly affect calcification processes during periodic molting, potentially influencing mortality.