Widespread distribution of histamine in the nervous system of a trematode flatworm.

In general, most flatworms contain very little histamine (HA) and their nervous systems often lack, or contain very few, histaminergic elements. However, preliminary studies in our laboratory have revealed that the frog lung parasite, Haplometra cylindracea (Trematoda: Digenea), contains HA in a very high concentration. For this reason, the present study was undertaken to study the localization and synthesis of HA in this worm by using immunocytochemistry and high-pressure liquid chromatography (HPLC). Essentially all parts of the nervous system of H. cylindracea showed HA-like immunoreactivity. The paired cerebral ganglia and nerves emanating from these, including the longitudinal nerve cords, were intensely immunoreactive. The musculature of the pharynx, oral and ventral suckers, and those of the reproductive organs were all innervated by HA-immunoreactive fibers. Fiber plexuses beneath the tegument and throughout the parenchyma also showed HA-like immunoreactivity. HPLC studies revealed one of the highest HA concentrations in the animal kingdom, 6.49 +/- 1.36 nmole/mg protein, in the worm. The frog lung and blood contained very low concentrations of HA and could be excluded as sources for HA, while an enzyme assay revealed that the worm produces HA by decarboxylation of histidine. Thus, it is likely that H. cylindracea uses HA as a neurotransmitter or modulator.

GABA in the nervous system of parasitic flatworms.

In an immunocytochemical study, using an antiserum and a monoclonal antibody specific for the amino acid, gamma-aminobutyric acid (GABA), GABA-like immunoreactivity (GLIR) has been demonstrated for the first time in parasitic flatworms. In Moniezia expansa (Cestoda), GLIR was seen in nerve nets which were closely associated with the body wall musculature and in the longitudinal nerve cords. In the liver fluke Fasciola hepatica (Trematoda), the GLIR occurred in the longitudinal nerve cords and lateral nerves in the posterior half of the worm. GLIR was also detected in subtegumental fibres in F. hepatica. The presence of GABA was verified, using high-pressure liquid chromatography coupled with fluorescence detection. The concentration of GABA (mean +/- S.D.) in M. expansa anterior region was 124.8 +/- 15.3 picomole/mg wet weight, while in F. hepatica it was 16.8 +/- 4.9 picomole/mg. Since several insecticides and antinematodal drugs are thought to interfere with GABA-receptors, the findings indicate that GABAergic neurotransmission may be a potential target for chemotherapy in flatworms too.

Binding of GRP(14-27) but not bombesin or GRP(1-27) to hypothalamic magnocellular elements: an immunohistochemical study.

Bombesin, gastrin-releasing peptide (1-27), and gastrin-releasing peptide (14-27) abolished the specific immunocytochemical staining revealed by antiserum directed to the C-terminus of gastrin releasing peptide (GRP) and bombesin (BN) in rat hypothalamus. When the antiserum was preabsorbed with GRP(14-27), a strong reaction appeared in hypothalamic magnocellular neurons. This staining of magnocellular elements was produced by lower concentrations of GRP(14-27) than were needed to block immunocytochemical staining revealed by the antiserum in other hypothalamic locations. The distribution of GRP(14-27)-induced immunostaining was similar to that of neurophysin. Since only GRP(14-27) but not GRP(1-27) or bombesin was found to bind to magnocellular cells, it was concluded that binding was due to the N-terminus of GRP(14-27), which resembles the structure of oxytocin and vasopressin. In agreement with this, oxytocin and vasopressin were found to prevent the binding of GRP(14-27) to magnocellular cells. The similarity in localization and the effect of oxytocin and vasopressin suggest that GRP(14-27) may bind to neurophysin at low concentrations. The results suggest that enhancement of staining after preabsorption of antisera with antigens must be interpreted with care. Enhancement can occur at antigen concentrations lower than those required to block the immunostaining. These results fail to support the premise that antigen-induced enhancement of staining is due to antigen binding to specific receptors and subsequent detection of the receptor-bound antigen with the antiserum.

Immunohistochemical demonstration of substance P in the lower respiratory tract of the rabbit and not of man.

Substance P (SP)-immunoreactive nerve fibres were searched for at all levels of both fetal and adult human lower respiratory tract. Because the demonstrability of substance P immunoreactivity varies between different animal species, rabbit pulmonary tissue was also subjected to SP immunohistochemistry. Human irises and corneas served as positive human controls. The specimens were taken from 10 human lungs during pulmonary operations. Tracheal tissue was obtained from three patients during bronchoscopy. Five fetal human lungs were examined. Human specimens examined included the trachea, main bronchi, segmental bronchi, and peripheral pulmonary tissue. In addition, the tracheobronchial tissues of four rabbits were studied. SP immunoreaction was demonstrated in formaldehyde-fixed cryostat sections by either the indirect immunofluorescence technique or the peroxidase-antiperoxidase procedure. Both monoclonal and conventional antibodies to SP were tested. In the rabbit SP-immunoreactive nerves were found in both the submucosa and the smooth muscle layer of the main bronchi and trachea. Specimens from human trachea, bronchi, and bronchioli were all negative. Since the SP immunoreaction was easily demonstrated in both human cornea and human iris, it was concluded that there are no SP-immunoreactive nerves in the human pulmonary tissues or that their SP content is very low and below the sensitivity of all the techniques used.