Visualization of putative ion-transporting epithelia in Amphibalanus amphitrite using correlative microscopy: Potential function in osmoregulation and biomineralization

Thoracican barnacles are a unique suborder of crustaceans typified by their calcified exterior, which provides protection to the sessile juvenile and adult. Biomineralization is mediated by a mantle epithelium that appears to be involved in calcium uptake and the secretion of calcium laden matrix. Larval and adult intertidal Balanomorph barnacles tolerate a wide range of salinities and it is hypothesized that active ion transport is the primary mechanism for osmoregulation. We observed adult Amphibalanus amphitrite producing an electrolyte-rich secretion emanating from the junction of the basis and parietal plates. Further study of this region using silver staining microscopic techniques, verified by scanning electron microscopy-energy dispersive spectroscopy, revealed a chloride ion rich mantle epithelium. A distinctive pattern of silver chloride stained epithelia was revealed in all A. amphitrite life stages. These epithelia were observed to contain mitochondria rich cells in nauplius and cyprid larvae (as shown by DASPMI staining visualized with confocal laser scanning microscopy) and therefore exhibit potential for active ion transport. Rhod-5 N (a low affinity cellular Ca2+ indicator) labeling was also observed in all barnacle life stages, in tissues shown to be chloride positive. We suspect that the observed chloride ion rich epithelia facilitate ionic regulation via active transport, and biomineralization via cellular Ca2+ uptake, storage and mobilization.