Tetrodotoxin (TTX), an extremely potent low-molecular-weight neurotoxin, is widespread among marine animals including ribbon worms (Nemertea). Previously, studies on the highly toxic palaeonemertean Cephalothrix cf. simula showed that toxin-positive structures are present all over its body and are mainly associated with glandular cells and epithelial tissues. The highest TTXs concentrations were detected in a total extract from the intestine of the anterior part of the body and also in a total extract from the proboscis. However, many questions as to the TTXs distribution in the organs of the anterior part of the worm's body and the functions of the toxins in these organs are still unanswered. In the present report, we provide additional results of a detailed and comprehensive analysis of TTXs distribution in the nemertean's proboscis, buccal cavity, and cephalic gland using an integrated approach including high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS), confocal laser scanning microscopy with anti-TTX antibodies, light and electron microscopies, and observations of feeding behavior. For the proboscis, we have found a TTXs profile different from that characteristic of other organs and tissues. We have also shown for the first time that the major amount of TTXs is localized in the anterior part of the proboscis that is mainly involved in hunting. TTX-containing glandular cells, which can be involved in the prey immobilization, have been found in the buccal cavities of the nemerteans. A significant contribution of the cephalic gland to the toxicity of this animal has been shown for the first time, and the role of the gland is hypothesized to be involved not only in protection against potential enemies but also in immobilizing prey. The data obtained have made it possible to extend the understanding of the role and features of the use of TTXs in the organs of the anterior part of nemertean's body.
Keywords: HPLC–MS/MS; TTX; buccal cavity; cephalic gland; nemertea; proboscis; tetrodotoxin; tetrodotoxin analogues.