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Statoliths of Aurelia coerulea were successfully labeled with 34S and Sr and were visualized for the first time by nanoscale secondary ion mass spectrometry (NanoSIMS) and electron probe microanalysis (EPMA). Medusae were incubated in artificial seawater with an enrichment of 34S and Sr for 25 hours. Statocysts for analyses were fixed at two time-points: immediately after the exposure (zero-day) and after 3 weeks of incubation in non-enriched seawater (post-incubated). In the analyses by NanoSIMS, deposition of 34S was observed as an inner ring enclosed within the crystal in post-incubated specimens. In zero-day specimens, 34S/32S ratio in the peripheral region of statolith was much higher than those in the core region. In the analyses by EPMA, small statoliths with dense deposition of Sr were located in the basal part of statocysts in zero-day specimens. The larger statoliths tended to be located more distant from the basal part. These findings were consistent with the hypothesis that the basal part of statocysts is the area of origin of statoliths. Statoliths without deposition of Sr tend to be larger in size than those with deposition, suggesting the saturation of growth after crystals attain a certain size. [ABSTRACT FROM AUTHOR]

Taxonomic work on soft-bodied and gelatinous cnidarians is a challenging task because preservation often causes difficulties for storage and identification of the specimens. Moreover, examination of scyphozoan and cubozoan statoliths, which can probably provide information on systematics and the age of medusae, are difficult since the crystals dissolve when exposed to air or aqueous solutions. In order to overcome such limitations, we evaluated the benefits of a non-invasive approach using X-ray micro-computed tomography (micro-CT). Various cnidarian taxa (Scyphozoa, Cubozoa, Hydrozoa, Staurozoa) were investigated with particular focus on scyphozoan and cubozoan statoliths. We further evaluated the effect of different fixatives (e.g. ethanol, glutaraldehyde, Trumps) and contrast-enhancing agents (iodine or osmium tetroxide). The reconstruction of tomographic data revealed that osmium tetroxide staining significantly enhanced the contrast of diagnostic morphological features of soft tissues whereas iodine staining was less effective. The reconstruction of scyphozoan and cubozoan statocysts allowed statolith analyses in their natural arrangement as well as automated morphometric measurements in three dimensions. In conclusion, micro-CT allows a detailed reconstruction of taxonomically important characters of soft-tissues and provides a sophisticated tool for reconstructing the spatial arrangement of complex internal structures expanding the opportunities for future research on marine organisms. [ABSTRACT FROM AUTHOR]