Samples were subsequently washed in cacodylate buffer, dehydrated in a graded series of ethanol, and isoamyl acetate better (10 minutes each at room temperature), critical point dried in CO2, coated with gold, and examined with a Philips XL30 SEM.2.4. Transmission Electron Microscopy (TEM)Small pieces of the integument were fixed in 2% paraformaldehyde (Scharlau) and 2% glutaraldehyde in cacodylate buffer for approximately 3h at 4��C. Then, tissues were washed three times (1h each) in the same buffer and postfixed in 1% osmium tetroxide in cacodylate buffer for 2h at 4��C. After rinsing the tissues in buffer, they were then dehydrated in a graded series of acetone and subsequently, embedded in Spurr’s resin. Toluidine blue-(Sigma) stained semithin sections were used to determine the area of study.

Ultrathin sections were stained with uranyl acetate and lead citrate and analyzed with either a Philips CM20 or a Jeol JEM1010 TEM.3. Results3.1. General Features and Scanning Electron MicroscopyThe foot epithelium ultrastructure of Haliotis tuberculata is schematized in the drawings shown in Figures 1(a) and 1(b). The side foot is lined by two types of columnar epithelial cells showing a prominent brush border interspersed with ciliated cells and four different types of epithelial secretory cells (Figure 1(a)). By contrast, the sole foot is characterized by taller columnar ciliated cells with three kind of epithelial secretory cells among them. Moreover clusters of secretory cells are embedded in the subepithelial space of the sole foot, and, due to their arrangement in glandular complexes, we refer to them as subepithelial glands (Figure 1(b)).

Figure 1Schematic drawing of the side (a) and sole (b) foot epithelia. Secretory cells (A, B, C, D, E, and F), basal membrane (bm), cilia (ci), Golgi complex (g), cell junctions (cj), microfilaments (mf), mitochondria (mt), microvillus border (mv), nuclei (n), …The external surface of the side foot is relatively rough containing many vertical folds that form crests and grooves (Figure 2(a)). At SEM, microvillus epithelial cells are observed alternating with the openings of secretory cells (Figure 2(b)). Moreover, cells with small ciliary tufts occur in a very low density separated from each other more than 100��m. The diameter of ciliary tufts is around 4��m, and they are comprised of approximately 30 cilia surrounded by microvilli (Figures 2(b) and 2(c)).

However, the external surface of the sole foot bears a dense field Carfilzomib of long cilia, which are generally covered by a thick layer of mucus (Figure 2(d)). This layer of mucus is important to protect the foot during locomotion as well as to help with the adhesion to the rock’s surface.Figure 2Scanning electron micrograph. (a) General view of the side foot showing the folds (f) in the external surface. Bar: 200��m.