3.4| Effects of ambient minor cations on survival and hemolymph composition of euryhaline crabs under hypo-osmotic conditions
To investigate the effects of ambient minor cations on the survival of euryhaline crabs at low ambient salinity, H. tridens , M. japonicus , and C. dehaani were reared in 8.6 mmol/L NaCl solution supplemented with 25.2 mmol/L MgCl2, 9.9 mmol/L CaCl2, and 10.7 mmol/L KCl (referred to as 8.6 mmol/L NaCl+MCK solution), 8.6 mmol/L NaCl solution, and 17.1 mmol/L NaCl solution without additional salts. Additional salts did not have any effect on the survival of H. tridens and C. dehaani , showing 100% survival in both species (data not shown). In contrast, the survival rate considerably decreased in M. japonicus in 8.6 mmol/L NaCl+MCK solution (data not shown). Because some M. japonicus individuals still appeared intact in 8.6 mmol/L NaCl+MCK solution, further experiments were conducted using 4.3 mmol/L NaCl, 4.3 mmol/L NaCl supplemented with 30.5 mmol/L MgCl2, 11.7 mmol/L CaCl2, and 13.4 mmol/L KCl (referred to as 4.3 mmol/L NaCl+1.2 MCK solution), and 17.1 mmol/L NaCl+1.2 MCK solution. The survival rate of H. tridens and C. dehaani was 100%, but that of M. japonicus significantly decreased to 17% in 4.3 mmol/L NaCl+1.2 MCK solution (Fig. 7). A few M. japonicusindividuals survived in this solution but were moribund. In contrast, most M. japonicus individuals survived in 4.3 mmol/L NaCl and 17.1 mmol/L NaCl+1.2 MCK solution (Fig. 7B). These results indicated that higher concentrations of additional salts in combination with a lower NaCl concentration caused high mortality in M. japonicus in 4.3 mmol/L NaCl+1.2 MCK solution.
To examine which minor cations have lethal effect on M. japonicus , 30.5 mmol/L MgCl2 and 11.7 mmol/L CaCl2 (1.2MC) and 13.4 mmol/L KCl (1.2K) were added to 4.3 mmol/L NaCl solution separately. In M. japonicus reared in 4.3 mmol/L NaCl solution supplemented with 13.4 mmol/L KCl (referred to as 4.3 mmol/L NaCl+1.2 K solution), the survival rate significantly decreased to a comparable level as that in 4.3 mmol/L NaCl+1.2 MCK solution (Fig. 7D). Even after diluting KCl to 2.7 mmol/L (almost one-fourth of the concentration in seawater), the survival rate did not hardly recover (Fig. 7E), indicating that K+ accounts for the lethality in 4.3 mmol/L NaCl+ 1.2 MCK solution. In contrast, supplementation of 4.3 mmol/L NaCl with 30.5 mmol/L MgCl2 and 11.7 mmol/L CaCl2 did not have a lethal effect on M. japonicus ; instead, these divalent cations attenuated the lethal effect of K+ 15 h after initiation of rearing, although it did not block the lethal effect at all 48 h after initiation of rearing (Fig. 7D, F).
Possible effects of ambient minor cations under hypo-osmotic conditions on hemolymph ionic composition and osmotic concentration were examined in H. tridens , M. japonicus , and C. dehaani. They were incubated in the 4.3 mmol/L NaCl and 4.3 mmol/L NaCl+1.2 MCK solutions, and hemolymph Na+, K+, and osmotic concentrations were determined 6 h after incubation. There were no significant differences in the hemolymph Na+concentration between crabs incubated in 4.3 mmol/L NaCl and 4.3 mmol/L NaCl+1.2 MCK solutions in all three species (Fig. 8A–C). In contrast, the hemolymph K+ concentration in crabs incubated in 4.3 mmol/L NaCl+1.2 MCK solution was significantly higher compared to 4.3 mmol/L NaCl solution in H. tridens and M. japonicusbut not in C. dehaani (Fig. 8E–G). Moreover, addition of minor cations increased osmotic concentration significantly in M. japonicus , but not in H. tridens and M. japonicus (Table 3). Additional experiments were conducted to know specific role of K+ on the hemolymph ionic composition in M. japonicus. Administration of KCl increased the hemolymph K+ concentration as combined administration of MgCl2, CaCl2, and KCl (Fig. 8H).