G. Gargiulo

The effects of cadmium on the gills of the goldfish Carassius auratus L.: Metal uptake and histochemical changes

Abstract 1. Uptake of cadmium and histochemical changes of mucopolysaccharides and of cytochromeoxidase in the gill epithelium of Carassius auratus were examined at different times after 10ppm Cd 2+ exposure in hard water (520 mg/l CaCO 3 ). 2. All animals survived treatment and no significant behavioural changes were observed. 3. Cd 2+ precipitates completely as CdCO 3 in the first 72 hr. Nevertheless, metal uptake in the organ was observed beyond this period. 4. After 96 hr an increase in the amount of sulphate mucins and a reduction in intensity of the cytochrome-oxidase reaction were observed. 5. Cadmium toxicity in very hard water is interpreted as being due to CdCO 3 ingestion which might cause histochemical alterations in the gills.

Immunohistochemical and immunochemical characterization of the distribution of leptin-like proteins in the gastroenteric tract of two teleosts (Dicentrarchus labrax and Carassius auratus L.) with different feeding habits.

Leptin is a modulator of food intake and energy homeostasis both in mammals and in some species of nonmammals vertebrates. In this study, we reported for the first time, using an immunohistochemical and immunochemical approach, the presence and distribution of immunoreactivity to leptin‐like protein in the gastroenteric tract of Dicentrarchus labrax (bass) and Carassius auratus (goldfish), two teleostean species with different feeding and different adaptative morphological organization of the gastroenteric tract. Bass stomach showed intense immunoreactivity to leptin‐like protein in all regions, with immunoreactive cells located at the base of the mucosal plicae and at the apical margin of the gastric crypts. Immunoreactive fibers and neuronal cells were observed close to vascular structures in the pyloric region. In bass and goldfish intestine, rare immunoreactive cells were observed along the mucosal epithelium mostly at the base or the apex of intestinal folds in the proximal and medium intestine; numerous immunoreactive nerve fibers in the circular and longitudinal layers of the tunica muscolaris as well as in the myenteric plexus were observed. Western blot analysis recognized a ∼15 kDa signal with a similar expression pattern for goldfish and sea bass. Our results could contribute to confirm the evolutive conservation of leptin‐like proteins and their probably precocious functional diversification in fish. Microsc. Res. Tech., 2011.

PACAP (pituitary adenylate cyclase-activating peptide)-like immunoreactivity in the gill arch of the goldfish, Carassius auratus: distribution and comparison with VIP.

Abstract Pituitary adenylate cyclase-activating peptide (PACAP) is a novel vasoactive intestinal peptide (VIP)-like peptide isolated from ovine hypothalamus. It is present in neuronal elements of a number of peripheral organs. We have examined whether PACAP occurs in the gill arch of Carassius auratus L. in which our recent studies have shown the presence of VIP-like peptide. Immunohistochemistry has revealed PACAP-like immunoreactivity in the anterior branches of the post-trematic glossopharyngeal and vagus nerves. PACAP-immunoreactive nerve cell bodies and fibers are present in connective tissue on the oral side of the gill arch. Colocalization studies carried out by the application of double immunofluorescence show that a PACAP-like peptide coexists with VIP in the same nerve cell bodies and fibers. The localization pattern of PACAP in the gill arch of goldfish suggests its possible involvement in the regulation of secretory activities.

Distribution of ghrelin peptide in the gastrointestinal tract of stomachless and stomach-containing teleosts.

The occurrence and localization of ghrelin peptide in the gastrointestinal tract of Carassius auratus and Dicentrarchus labrax, two fish species that exhibit different feeding behavior, different habitats, and different anatomical organizations of the gastroenteric tract, were examined by immunohistochemical methods and western blotting analysis. All of the gastrointestinal segments studied displayed immunohistochemical localizations of ghrelin peptide. Numerous single or clustered immunoreactive cells were found along the gastric folds, particularly in the pyloric region of Dicentrarcus labrax, whereas scattered ghrelin immunoreactive cells were observed in the intestinal epithelium of both fish species. Double immunolabeling PGP 9.5/ghrelin demonstrated the localization of ghrelin peptide also in nerve fibers and neuronal cells of the submucosal and myenteric plexuses, often in association with vascular structures. Western blotting analysis confirmed the presence of ghrelin peptide in the gatrointestinal tract of both species studied, whose molecular weight was similar to that of the corresponding mammalian prepro‐ghrelin. The findings could support the hypothesis that this peptide is an important appetite regulator in fish and could confirm the presence of the ghrelin peptide, starting from its precursor proteins, in the gastrointestinal tract of the goldfish and the sea bass. Microsc. Res. Tech. 2009.

Anatomical distribution of NPY-like immunoreactivity in the domestic chick brain (Gallus domesticus)

Neuropeptide Y‐immunoreactive (NPY‐ir) fibers and neurons in the brain of the domestic chick (Gallus domesticus) were described using an immunohistochemical technique. NPY‐ir neurons were seen in the lobus parolfactorius; hyperstriatum, neostriatum, paleostriatum, and archistriatum; hippocampal and parahippocampal areas; dorsolateral corticoid area; piriform cortex; two thalamic areas contiguous to the n. rotundus; n. dorsolateralis anterior thalami, pars lateralis, and pars magnocellularis; n. periventricularis hypothalami; n. paraventricularis magnocellularis; regio lateralis hypothalami; n. infundibuli; inner zone of the median eminence; dorsal and lateral portions of the n. opticus basalis; n. raphes; and n. reticularis paramedianus. NPY‐ir fibers were seen throughout the entire chick brain, but were more abundant in the hypothalamus where they formed networks and pathways. They were also observed in some circumventricular organs. The anatomical data of the present study regarding the distribution of NPY ir in the chick brain, together with the physiological findings of other studies, suggest that NPY plays a key role in the regulation of the neuroendocrine, vegetative, and sensory systems of birds by acting as a neuromodulator and/or neurotransmitter. Anat Rec 263:186–201, 2001.

Action of cadmium on the gills of Carassius auratus L. in the presence of catabolic NH3

Histochemical changes of oxidative enzymes and cadmium uptake in the Carassius auratus gills were examined up to 7 days after a 87.6 μM cadmium exposure in very alkaline water in the presence and absence of catabolic ammonia. In the presence of catabolic NH3, the toxic action of cadmium is much more intense and determines a 20% survival of goldfish with a heavy increase in the excretion of mucus. Animals treated with cadmium only and controls survived in the order of 100%. More drastic histochemical alterations in the activity of oxidative enzymes (SDH, G-6-PDH, LDH) were documented in animals treated both with cadmium and ammonia, if compared with those treated with cadmium only. A reduction in the activity of enzymes after 24 h, a heavy increase of the LDH reaction after 72 h, and a strong increase after 7 days in the number of positive SDH cells (which correspond to chloride cells) were documented. After a hyperaccumulation at 24 h (returning afterwards to much lower values), at 7 days the cadmium uptake was higher in the animals treated both with cadmium and ammonia.The authors assume that these effects should be ascribed to the complexation of cadmium with catabolic ammonia.

Neurotensin‐like immunoreactivity in the brain of the chicken, Gallus domesticus

The distribution of neurons containing neurotensin in the central nervous system of the chicken was studied immunohistochemically. The majority of the neurotensin‐immunoreactive (‐ir) cell bodies were located in the hypothalamus. Extensive groups of labelled perikarya were found in the hypothalamic periventricular nucleus and in the magnocellular periventricular nucleus. In addition, ir‐perikarya were scattered throughout the lateral hypothalamic area and in the ventromedial hypothalamic nucleus. The only extrahypothalamic site of ir‐perikarya was in the region immediately under the lateral forebrain bundle. Immunoreactive fibres were detected in the hippocampus, the parahippocampal area, the hypothalamus, the region of the tractus corticohabenular and corticoseptal tracts, the median eminence, the region above the posterior commissure and in the intercollicular nucleus. The distribution pattern of the neurotensin‐ir neurons suggests that neurotensin‐like peptides are involved in the hypophysiotropic functions.

Ghrelin in the Gastroenteric Tract of Birds: Immunoreactivity Expression

S. Neglia*, N. Arcamone, V. Esposito and G. Gargiulo Department of Biological Structures, Functions and T echnology – Faculty of Veterinary Medicine, University of Naples ‘‘Federico II’’ – Italy *Correspondence: Dipartimento di Strutture, Funzioni e T ecnologie Biologiche – Facolta di Medicina Veterinaria, Universita di Napoli ‘‘Federico II’’, V ia Veterinaria, 1, I-80137 Napoli, Italy E-mail: sineglia@unina.it

The teleost kidney expresses Trk Neurotrophin Receptor-Like Proteins

Neurotrophins are involved in the development and maintenance of some neuronal populations, acting through tyrosine protein kinase (Trk) receptors, TrkA, TrkB and TrkC. In addition, recent studies have demonstrated the occurrence of Trk receptors in a wide variety of adult and embryonic non-neuronal tissues in vertebrates, including kidney. Nevertheless no data are available on Trk proteins distribution in teleost kidney. The present study, by using an immunoperoxidase technique, analyses Trk receptor protein distribution in the kidney of three species of freshwater and saltwater teleost. TrkA-like immunoreactivity was the only one detected in all examined species, while TrkB-like immunoreactivity was completely absent and TrkC-like immunoreactivity was detected only in goldfish. The TrkA immunoreactive cells were mainly localised in the collecting duct system, as this system was the only one to also display TrkC. Such data could provide new clues to Trk in fish and aid assessment of the role of Trk protein receptors during vertebrate evolution.

Immunoreactivity to vasoactive intestinal polypeptide (VIP) in the hypothalamus of the domestic fowl, Gallus domesticus

The distribution of VIP-immunoreactive neurons and fibers was detected in the hypothalamus of the chick by immunohistochemistry and light microscopy. A large amount of VIP cellular bodies was localized in the anterior and medial area of the hypothalamus with the highest density of cells in supraoptic, magnocellular preoptic, suprachiasmatic and paraventricular nuclei. Only few VIP-immunoreactive neurons were observed in the caudal section of infundibuli nucleus. A considerable concentration of VIP-positive fibers was also detected in the external layer of the anterior and posterior median eminence. Their presence might have origin both from the neurons of the infundibuli nucleus and from the cells of the paraventricular nucleus. Few VIP-immunoreactive fibers were revealed in the organum vasculosum of the lamina terminalis. These results tend to suggest that VIP may play more than one role in the hypothalamic regions, particularly in the preoptico-hypothalamic area. The presence of this peptide in the median eminence supports even more the hypothesis that it may be released into the portal circulation and transported to the pars distalis of the pituitary gland.