3D model radula of Gastropteron rubrum

The molluscan feeding structure is the radula, a chitinous membrane with teeth, which are highly adapted to the food and the substrate to which the food is attached. In Polyplacophora and Patellogastropoda, the handling of hard ingesta can be facilitated by high content of chemical compounds containing Fe or Si in the tooth cusps. Other taxa, however, possess teeth that are less mineralized, even though animals have to avoid structural failure or high wear during feeding as well. Here, we investigated the gastropod Gastropteron rubrum, feeding on hard Foraminifera, diatoms and Porifera. Tooth morphologies and wear were documented by scanning electron microscopy and their mechanical properties were tested by nanoindentation. We determined, that gradients of hard- and stiffness run along each tooth, decreasing from cusp to basis. We also found, that inner lateral teeth are harder and stiffer than the outer ones. These findings allowed us to propose hypotheses about the radula-ingesta interaction. In search for the origins of the gradients, teeth were visualized using confocal laser scanning microscopy, to determine the degree of tanning, and analyzed with energy-dispersive X-ray spectroscopy, to test the elemental composition. We found that the mechanical gradients do not have their origins in the elemental content, as the teeth did not contain high proportions of metals or other minerals, which indicates that their origin might in the degree of tanning. However, in the tooth surfaces, which interact with the ingesta, high Si and Ca content was determined, which is likely an adaptation to reduce wear.

Methods
For the 3D visualization, mature radular teeth of the working zone of two radulae were extracted manually with forceps. Each tooth was mounted on SEM specimen holders by double-sided adhesive carbon tape, sputter-coated with platinum (5 nm layer), and visualized under the SEM from all sides. Using the 3D software Blender v2.83 (Blender Foundation), the teeth were then modelled by hand constantly comparing the 3D visualization with the SEM images taken from different sides (see also protocol in Krings et al., 2020, 2021e). In the same manner, the position and embedment of the teeth within the membrane were reconstructed.