Species: Diplostomum pseudospathaceum Niewiadomska, 1984    Trematoda

Preparation Type: whole mount

Prepared: A.A. Petrov

Description: RF-amidergic nervous system and the musculature of a 22-day-old metacercaria of Diplostomum pseudospathaceum (Trematoda, Platyhelminthes). Overall view of the metacercaria showing the distribution of RF-amidergic elements in the brain, anterior nerves, longitudinal nerve cords and the nerve innervating the internal organs. The preparation also shows the musculature of the body wall and internal organs (oral and ventral suckers, pharynx, intestine and holdfast).

Method: F-actin in muscle fibers (Phalloidin), FMRFamide

Method Description: The metacercariae were fixed for 15-17 hours with 4% formaldehyde in 0.01M PBS and were kept at 6°С in 0.1% sodium azide in 0.01М PBS. All subsequent procedures were performed at room temperature. Before staining, the worms were processed for 2 hours in 0.5% Triton X-100 in 0.01M PBS, incubated for 1-2 hours in a blocking solution (Image-iT® FX Signal Enhancer), and rinsed 3 times for 15 minutes in 0.5% Triton X-100 in 0.01M PBS. After rinsing, the worms were incubated for 12 hours in antibodies to FMRFamide (diluted 1:600, ref. number ab10352; Abcam), rinsed 4 times for 15 minutes in 0.5% Triton X-100 in 0.01M PBS and then incubated for 5 hours in secondary antibodies (diluted 1: 300, Chromeo 488, ref. number ab60314; Abcam). After incubation in antibodies, the worms were stained for 3 hours in phalloidin-TRITC (diluted 1:150, Sigma-Aldrich, ref. number P1951; Sigma-Aldrich), rinsed for 15 minutes in 0.01M PBS and mounted in Vectashield (Vector Laboratories Inc.).

Institution: Zoological Institute RAS

Description: General view of a 22-day-old metacercaria of Diplostomum pseudospathaceum (Platyhelminthes) showing the serotonergic nervous system and the musculature. The series shows the brain (cerebral ganglia), the anterior nerves and longitudinal nerve cords that extend from the brain, the commissures connecting the nerve cords and the nerves innervating the internal organs (sucker, lappets and the holdfast).

Organs: сentral nervous system, nerve cord, cerebral brain or its part, ganglion / ganglia, commissure, nerve, larval nervous system, musculature innervation / musculature, peripheral nervous system

Methods: Immunohistochemistry - FMRFamide,
Histochemistry - F-actin in muscle fibers (Phalloidin)