Fig. 3. Localization of 5-HT_1Hel and 5-HT_7Hel transcripts in whole-mounts of H. trivolvis embryos. In situ hybridization of digoxigenin (DIG)-labeled anti-sense RNA probes was immunohistochemically localized with alkaline phosphatase-conjugated anti-DIG antibodies and reactants that form either a purple (A–C; light microscope) or fluorescent red (D and E; confocal microscope) precipitate. Using anti-sense probes in stage E45 embryos (A, B and D), 5-HT_1Hel RNA expression was localized mainly to the ciliated epithelium of the foot (open arrows), anterior head (filled arrows), tentacles (^*), ciliated mantle (filled arrowheads) and gut (open arrowheads). Using sense probes to 5-HT_1Hel, no signal was observed (C). In stage E30 embryos (E), 5-HT_7Hel RNA expression was localized to the ciliated foot epithelium (open arrows), developing gut (open arrowheads), and ciliated mantle (filled arrowhead). Using sense probes to 5-HT_7Hel, no signal was observed (not shown). Scale bar, 100 µm.