Histochemical identification of the black-brown pigment granules found in the alimentary canal of Siberian sturgeon (Acipenser baeri) during the lecitotrophic stage

Abstract

The histochemical characteristics of the black-brown pigment granules accumulated during the lecitotrophic stage in the digestive tract of Siberian sturgeon, Acipenser baeri larvae, were studied in order to identify the nature of such pigments. Pigment granules appeared just after hatching in contact with the endodermal epithelium and in the periphery of the yolk-sac matrix, and increased in number with digestive system differentiation. At 3 days post hatch, the accumulation of pigment granules in the spiral valve formed a pigmented plug, which was ejected with first faeces after first feeding. Pigment granules were insoluble in all solubilising solutions tested (distilled water, H2O2, NH4OH, KMnO4, absolute methanol and different acid, basic and oxidising solutions). Pigment granules were completely bleached with the oxidising processes of Gomori and Casella (15 min) and with H2O2 (20%) during 48 h. Histochemical characteristics of bleached pigmented granules revealed that they were basophilic (Eosine negative), presented a moderate methachromasia (Toluidine Blue positive) and argentaffinity (Masson Fontana method positive), and contained carboxylated and sulphated (strongly and weakly ionised) glycoconjugated residues (Alcian Blue pH 0.5, 1.0, 2.5 positive). Bleached pigment granules also contained reductor groups (Ferric ferricyanide-Fe III positive), while they were negative for Bromophenol Blue, Perl's Prussian Blue, Red Oil O and Sudan Black B, indicating the absence of proteins, iron and lipidic substances respectively. According to such results, the black-brown pigment granules observed in the alimentary canal of Siberian sturgeon larvae during the lecitotrophic stage correspond to melanins.