K+, and osmotic concentrations
The crabs were anesthetized and dissected, and ~300 µl
(for H. tridens and C. dehaani ) or ~150 µl
(for M. japonicus ) of hemolymph was collected from each crab.
They were centrifuged at 2390 × g for 5 min at 4°C and the supernatant
was collected to be used as the sample solution. Na+and K+ concentrations in 1:10 diluted and undiluted
sample solutions, respectively, were measured using sodium (model C-122)
and potassium ion meters (model C-131) (HORIBA, Kyoto, Japan),
respectively. The osmotic concentrations were measured with a vapor
pressure osmometer (Model 5600, Wescor, Logan, UT, USA).