Larval salt gland ofArtemia salina nauplii

Summary

The naupliar stage ofArtemia salina has been found to contain large amounts of an ATPase which requires the presence of Mg⁺⁺, is stimulated by Na⁺ and K⁺, and is sensitive to ouabain. Upon cell fractionation, sixty percent of the Na+K-activated ATPase activity was present in a heavy membrane fraction which accompanied mitochondria, and twenty percent of the activity was found in the microsomal fraction. Separation of the heavy membranes containing the Na+K-activated ATPase from mitochondria was accomplished by sucrose density gradient centrifugation.

Kinetic characterization of the Na+K-activated ATPase was examined with both mitochondrial and microsomal preparations. No significant differences were observed between the two preparations. Maximal enzyme activity occurred at pH 7.2 and a temperature of 45 ° C. Half maximal inhibition of enzymatic activity occurred with ouabain concentration of 8 × 10⁻⁶ M. Half maximal activation of the enzyme by ATP and MgCl₂ occurred at 8 × 10⁻⁴ and 1 × 10⁻³ M, respectively, with an optimal Mg⁺⁺/ATP ratio of 2.0. NaCl and KC1 (or NH₄C1) were required for activity, with half maximal activation at 1 × 10⁻² and 9 × 10⁻³ M (or 2.1 × 10⁻² M), respectively. The optimal Na⁺/K⁺ ratio was 4.0, although enzymatic activity occurred through a wide range of ratios. In contrast, survival of nauplii was reduced in media in which the Na⁺/K⁺ ratio was less than 10.

Anatomical localization of the enzyme in nauplii indicated that the abdominal region contained 44% of the total Na+K-activated ATPase activity, while the cephalothoracic region contained 54% of the total activity. Isolated larval salt glands contained 10% or less of the total activity. In contrast, the adult leg segments containing the salt-transporting organs showed 54% of the total Na+K-activated ATPase activity.