Gross brain morphology of the armoured catfish Rineloricaria heteroptera,Isbrücker and Nijssen (1976), (Siluriformes: Loricariidae: Loricariinae): A descriptive and quantitative approach

The gross morphology of the brain of Rineloricaria heteroptera and its relation to the sensory/behavioural ecology of the species is described and discussed. The sexual and ontogenetic intraspecific variation in the whole brain length and mass, as well as within/between the eight different brain subdivisions volumes, is also examined and discussed. Negative allometry for the whole brain length/mass and relative growth of the telencephalon and optic tecta was observed. Positive allometry was observed for the relative growth of the olfactory bulbs and medulla oblongata . Univariate and multivariate statistical analyses did not reveal significant differences in the brain subdivision growth rates among sexes and/or developmental stages, except for the optic tectum and some portions of the medulla oblongata , with juveniles and males showing more developed optic tecta and medullary subdivisions, respectively. The growth rates for each brain subdivision were relatively constant, and the slopes of the growth equations were almost parallel, except for those of the olfactory bulbs and medulla oblongata subdivisions, suggesting some degree of tachyauxesis of subdivisions against the entire brain. The corpus cerebelli was the more voluminous brain subdivision in most specimens (principally adults), followed by the optic tectum (the more voluminous subdivision in juveniles), hypothalamus, and telencephalon, in that order. Differences in the number of lamellae and relative size of the olfactory organ were also detected among developmental stages, which were more numerous and larger in adults. Based on these results, it is possible to infer an ontogenetic shift in the habitat/resource use and behaviour of R. heteroptera . Vision, primarily routed through the optic tectum, could be fundamental in early stages, whereas in adults, olfaction and taste, primarily routed through the olfactory bulbs and medulla oblongata , play more important roles.     

Galanin and its binding sites in the brain of eels subjected to different osmolar conditions

A galanin (GAL)-like peptidergic system was investigated in the brain of Anguilla anguilla subjected to hyperosmolar and hypoosmolar conditions, by using antisera raised against porcine 1-29 GAL. A group of immunoreactive perikaria was identified in the periventricular hypothalamus, in the ventral thalamus, in the pretectal areas and in the optic tectum. Immunoreactive perikaria were present in the nucleus lateralis of the torus semicircularis in seawater (SW) adapted eels, and were absent in the freshwater (FW) adapted eels. Galaninergic fibres were observed in many areas of the brain. The immunoreactive perikaria and fibres, when localised in the same areas, were more strongly labelled and numerous in the SW adapted eels in comparison to the FW adapted animals. GAL-specific binding sites, investigated by autoradiography using iodinated porcine GAL, occurred in several regions of the brain, in particular in the dorsal telencephalon, in the dorsal thalamus and in the torus semicircularis, where a high density of binding sites was observed in the SW adapted eels. Conversely, a higher density of binding sites was observed in the caudal substantia reticularis and in the corpus cerebelli in the FW adapted eels. We conclude that different distributions and intensity in immunoreactive elements occur in response to the two environmental conditions.     

Adult neurogenesis in the brain of the Mozambique tilapia, Oreochromis mossambicus

Although the generation of new neurons in the adult nervous system ('adult neurogenesis') has been studied intensively in recent years, little is known about this phenomenon in non-mammalian vertebrates. Here, we examined the generation, migration, and differentiation of new neurons and glial cells in the Mozambique tilapia (Oreochromis mossambicus), a representative of one of the largest vertebrate taxonomic orders, the perciform fish. The vast majority of new cells in the brain are born in specific proliferation zones of the olfactory bulb; the dorsal and ventral telencephalon; the periventricular nucleus of the posterior tuberculum, optic tectum, and nucleus recessi lateralis of the diencephalon; and the valvula cerebelli, corpus cerebelli, and lobus caudalis of the cerebellum. As shown in the olfactory bulb and the lateral part of the valvula cerebelli, some of the young cells migrate from their site of origin to specific target areas. Labeling of mitotic cells with the thymidine analog 5-bromo-2'-deoxyuridine, combined with immunostaining against the neuron-specific marker protein Hu or against the astroglial marker glial fibrillary acidic protein demonstrated differentiation of the adult-born cells into both neurons and glia. Taken together, the present investigation supports the hypothesis that adult neurogenesis is an evolutionarily conserved vertebrate trait.     

Binding of the benzodiazepine ligand [H]-Ro 15–1788 to brain membrane of the saltwater fish Mullus surmuletus

The distribution and the pharmacological properties of the binding of the benzodiazepine receptor antagonist [³H]-Ro 15–1788 (8-fluoro-3-carboethoxy-5,6-dihydro-5-methyl-6-oxo-4H imidazol [1,5-a] 1,4 benzodiazepine) were compared in some brain membranes of the saltwater teleost fish, Mullus surmuletus: only a single population of [³H]-Ro 15–1788 binding sites was detected. The binding was saturable and reversible with a high affinity, revealing a significant population of binding sites (K_d value of 2.1 ± 0.2 nM and B_max value of 1400-900 fmol mg⁻¹ of protein, depending on fish length). The highest concentration of benzodiazepine recognition sites labelled with [³H]-Ro 15–1788 was present in the optic lobe and the olfactory bulb and the lowest concentration was found in the medulla oblongata, cerebellum and spinal cord. In order to explore behavioural selectivity as a consequence of multiple receptor subtypes, six benzodiazepine receptor ligands, flunitrazepam (5-(2-fluoro-phenyl)-1,3,dihydro-1-methyl-7-nitro-2H-1,4-benzodiazepine-2-one), alpidem, (N,N-dipropyl-6-chloro-2-(4-chlorophenyl) imidazo [1,2-a] pyridine-3-acetamide) zolpidem {N,N,6, trimethyl-2-(4-methyl-phenyl) imidazo [1,2-a] pyridine-3-acetamide hemitartrate}, methyl β carboline-3-carboxylate (βCCM), Ro 15–1788 and Ro 5–4864 (4′-chlorodiazepam), were tested in vitro by binding of [³H]-Ro 15–1788 to membrane preparations from various brain areas of Mullus surmuletus. Displacement studies showed a similar rank order of efficacy of various unlabelled ligands. In all regions of the brain and in the spinal cord, GABA potentiate [³H]-flunitrazepam binding in a similar order, suggesting that the BDZ recognition sites are part of the GABA_A receptor structure. These results suggest that central-type benzodiazepine receptors are present in one class of benzodiazepine binding sites in the saltwater teleost fish brain of Mullus surmuletus (type I-like). Here we report initial evidence of homogeneity of subtypes of central benzodiazepine receptors in the spinal cord of the saltwater teleost fish, Mullus surmuletus.     

3H]Ro 15-1788 binding sites to brain membrane of the saltwater Mugil cephalus

The equilibrium binding parameters of the benzodiazepine antagonist [3H]Ro 15-1788 (8-fluoro-3-carboethoxy-5,6-dihydro-5-methyl-6-oxo-4H-imidazol-[1,5-a]-1,4 benzodiazepine) were evaluated in brain membranes of the saltwater teleost fish, Mugil cephalus. To test receptor subtype specificity, displacement studies were carried out by competitive binding of [3H]Ro 15-1788 against six benzodiazepine receptor ligands, flunitrazepam [5-(2-fluoro-phenyl)-1,3-dihydro-1-methyl-7-nitro-2H-1,4-benzodiazepin-2-one], alpidem [N,N-dipropyl-6-chloro-2-(4-chlorophenyl)imidazo[1,2-a]pyridine-3-acetamide], zolpidem [N,N-6 trimethyl-2-(4-methyl-phenyl)imidazo[1,2-a]pyridine-3-acetamide hemitartrate], and beta-CCM (methyl beta-carboline-3-carboxylate). Saturation studies showed that [3H]Ro 15-1788 bound saturatably, reversibly and with a high affinity to a single class of binding sites (Kd value of 1.18-1.5 nM and Bmax values of 124-1671 fmol/mg of protein, depending on brain regions). The highest concentration of benzodiazepine recognition sites labeled with [3H]Ro 15-1788 was present in the optic lobe and the olfactory bulb and the lowest concentration was found in the medulla oblongata, cerebellum and spinal cord. The rank order of displacement efficacy of unlabelled ligands observed suggested that central-type benzodiazepine receptors are present in one class of binding sites (Type I-like) in brain membranes of Mugil cephalus. Moreover, the uptake of 36Cl- into M. cephalus brain membrane vesicles was only marginally stimulated by concentrations of GABA that significantly enhanced the 36Cl- uptake into mammalian brain membrane vesicles. The results may indicate a different functional activity of the GABA-coupled chloride ionophore in the fish brain as compared with the mammalian brain.     

Anterograde labelling from the optic nerve reveals multiple central targets in the teleost,Lethrinus chrysostomus (Perciformes)

Summary Cobaltous-lysine is transported anterogradely from the optic nerve of the teleost, Lethrinus chrysostomus (Lethrinidae, Perciformes). The marginal optic tract is labelled in longtitudinal bands of light and dark staining fibres which persists caudally within the ventral division but not in the dorsal division. This species possesses multiple central targets in the contralateral preoptic, diencephalic, pretectal, periventricular and tectal regions of the brain. In addition, a greater subdivision of the marginal optic tract is found to project to various nuclei. Ipsilateral projections are found in the suprachiasmatic nucleus and in the region of the horizontal commissure. Projections are also found in the telencephalic region of the nucleus olfactoretinalis and the thalamic region of the nucleus thalamoretinalis. The retinotopicity of some of these nuclei, found in previous studies, is discussed in relation to the possibility of specific sub-populations of retinal ganglion cells having different central targets.Abbreviations used in the Text and Figures A nucleus anteriorthalami - AO accessory optic nucleus - AOT accessory optic tract - AxOT axial optic tract - BO nucleus of the basal optic root - C cerebellum - HCv ventral division of horizontal commissure - I nucleus intermedius thalami - IL inferior lobe - MdOT medial optic tract - MO medulla oblongata - MOTd dorsal division of the marginal optic tract - MOTi intermediate division of the marginal optic tract - MOtv ventral division of the marginal optic tract - O olfactory bulb - OT optic tract - PC nucleus pretectalis centralis - PCo posterior commissure - Pd nucleus pretectalis dorsalis - PG preglomerular complex - PPd nucleus pretectalis periventricularis, pars dorsalis - PPv nucleus pretectalis periventricularis, pars ventralis - PSm nucleus pretectalis superficial pars magnocellularis - PSp nucleus pretectalis superficialis, pars parvocellularis - Sn suprachiasmatic nucleus - TEL telencephalon - TeO optic tectum - TL torus longtitudinalis - TrOlfR tractus olfactoretinalis - VCg granular layer of the valvula cerebelli - VCm molecular layer of the valvula cerebelli - VM nucleus medialis thalami - VL nucleus ventrolateralis thalami - VMdOT ventro-medial optic tract     

Melatonin Binding Sites in Senegal Sole: Day/Night Changes in Density and Location in Different Regions of the Brain

We localized melatonin binding sites in different brain regions (optic tectum, telencephalon, cerebellum, hypothalamus, olfactory bulbs, and medulla oblongata) of Senegal sole, a species of aquaculture interest, and checked day/night changes in density (B_max) at mid‐light (ZT06) and mid‐dark (ZT18). Plasma melatonin was measured using a radioimmunoassay, while binding assays were performed using 2‐[¹²⁵I]iodomelatonin as a radioligand. Plasma melatonin concentrations were significantly lower at mid‐light (189.5±46 pg/ml) than mid‐dark (455.5±163 pg/ml). Values of B_max were statistically significantly higher in the optic tectum (5.6±0.6 and 12.3±1 fmol/mg prot, at mid‐light and mid‐dark, respectively) and in the cerebellum (7.7±1.1 and 10.6±1.3 fmol/mg prot, at mid‐light and mid‐dark, respectively). Significant day/night differences were only observed in these two tissues. These results show for the first time the distribution of melatonin binding sites within the brain of a flatfish species and their lack of down‐regulation.     

Melatonin binding sites in senegal sole: day/night changes in density and location in different regions of the brain

We localized melatonin binding sites in different brain regions (optic tectum, telencephalon, cerebellum, hypothalamus, olfactory bulbs, and medulla oblongata) of Senegal sole, a species of aquaculture interest, and checked day/night changes in density (B(max)) at mid-light (ZT06) and mid-dark (ZT18). Plasma melatonin was measured using a radioimmunoassay, while binding assays were performed using 2-[(125)I]iodomelatonin as a radioligand. Plasma melatonin concentrations were significantly lower at mid-light (189.5+/-46 pg/ml) than mid-dark (455.5+/-163 pg/ml). Values of B(max) were statistically significantly higher in the optic tectum (5.6+/-0.6 and 12.3+/-1 fmol/mg prot, at mid-light and mid-dark, respectively) and in the cerebellum (7.7+/-1.1 and 10.6+/-1.3 fmol/mg prot, at mid-light and mid-dark, respectively). Significant day/night differences were only observed in these two tissues. These results show for the first time the distribution of melatonin binding sites within the brain of a flatfish species and their lack of down-regulation.     

Two distinct (-)nicotine binding sites in goldfish brain. Identification and characterization of putative neuronal nicotinic acetylcholine receptor subtypes

The binding of (-)-[3H]nicotine to membrane fragments and a detergent solubilized fraction of goldfish brain was characterized. (-)-[3H]nicotine binding was not displaced by alpha-bungarotoxin, but was displaced by (-)nicotine and carbamoylcholine with Ki of approximately 8.6 and 86 nM, respectively. Preincubation of solubilized membrane extract with alpha-bungarotoxin-coupled Sepharose resulted in the loss of approximately 50% of the (-)-[3H]nicotine binding protein from the eluent and an increase in (-)-[3H]nicotine binding to the gel compared to control, non-alpha-bungarotoxin Sepharose. 125I-alpha-bungarotoxin binding protein in the eluent from the same preincubation experiments was totally removed. In addition, incubation of the solubilized tissue extracts with alpha-bungarotoxin-coupled Sepharose resulted in an increase in the affinity for (-)-[3H]nicotine in the eluent (mean KD = 3.1) compared to control solubilized tissue extracts (KD = 6.4 nM). Specific (-)-[3H]nicotine binding sites could be eluted from the alpha-bungarotoxin-coupled Sepharose with carbamoylcholine and D-tubocurarine. Similar to previously reported 125I-alpha-bungarotoxin binding data, eye removal resulted in an approximately 40% decrease in (-)-[3H]nicotine binding in the contralateral tectum compared to that in the ipsilateral tectum. These data indicate that at least two distinct subtypes of (-)nicotine binding sites may be present in goldfish brain, one which binds alpha-bungarotoxin and (-)nicotine and another which binds only (-)nicotine.     

Melatonin binding sites in the brain of European sea bass (Dicentrarchus labrax)

Characteristics, day-night changes, guanosine 5'-O-(3-thiotriphosphate) (GTPgammaS) modulation, and localization of melatonin binding sites in the brain of a marine teleost, European sea bass Dicentrarchus labrax, were studied by radioreceptor assay using 2-[(125)I]iodomelatonin as a radioligand. The specific binding to the sea bass brain membranes was rapid, stable, saturable and reversible. The radioligand binds to a single class of receptor site with the affinity (Kd) of 9.3 +/-0.6 pM and total binding capacity (Bmax) of 39.08 +/-0.86 fmol/mg protein (mean+/-SEM, n=4) at mid-light under light-dark (LD) cycles of 12:12. Day-night changes were observed neither in the Kd nor in the Bmax under LD 12:12. Treatment with GTPgammaS significantly increased the Kd and decreased the Bmax both at mid-light and mid-dark. The binding sites were highly specific for 2-phenylmelatonin, 2-iodomelatonin, melatonin, and 6-chloromelatonin. Distribution of melatonin binding sites in the sea bass brain was uneven: The Bmax was determined to be highest in mesencephalic optic tectum-tegmentum and hypothalamus, intermediate in telencephalon, cerebellum-vestibulolateral lobe and medulla oblongata-spinal cord, and lowest in olfactory bulbs with the Kd in the low picomolar range. These results indicate that melatonin released from the pineal organ and/or retina plays neuromodulatory roles in the sea bass brain via G protein-coupled melatonin receptors.     

A reinvestigation of the Gn-RH (gonadotrophin-releasing hormone) systems in the goldfish brain using antibodies to salmon Gn-RH

Summary The organization of Gn-RH systems in the brain of teleosts has been investigated previously by immunohistochemistry using antibodies against the mammalian decapeptide which differs from the teleostean factor. Here, we report the distribution of immunoreactive Gn-RH in the brain of goldfish using antibodies against synthetic teleost peptide.Immunoreactive structures are found along a column extending from the rostral olfactory bulbs to the pituitary stalk. Cell bodies are observed within the olfactory nerves and bulbs, along the ventromedial telencephalon, the ventrolateral preoptic area and the latero-basal hypothalamus. Large perikarya are detected in the dorsal midbrain tegmentum, immediately caudal to the posterior commissure. A prominent pathway was traced from the cells located in the olfactory nerves through the medial olfactory tract and along all the perikarya described above to the pituitary stalk. In the pituitary, projections are restricted to the proximal pars distalis. A second immunoreactive pathway ascends more dorsally in the telencephalon and arches to the periventricular regions of the diencephalon. Part of this pathway forms a periventricular network in the dorsal and posterior hypothalamus, whereas other projections continue caudally to the medulla oblongata and the spinal cord. Lesions of the ventral preoptic area demonstrate that most of the fibers detected in the pituitary originate from the preoptic region.     

Distribution of salmon gonadotrophin releasing-hormone in the brain and pituitary of the sea bass (Dicentrarchus labrax)

Summary The distribution of salmon gonadotrophin-releasing hormone (sGnRH) was studied in the brain and pituitary of two-year-old immature sea bass (Dicentrarchus labrax) by means of an enzymoimmunoassay (EIA) for sGnRH and immunocytochemistry. The EIA for sGnRH is a competitive assay using a tracer made of sGnRH coupled to acetylcholinesterase from an electric eel. The separation of free and bound tracer is achieved by coating the plates with mouse anti-rabbit IgG monoclonal antibodies. Displacement curves generated by sGnRH and extracts from pituitary and different brain regions showed a good parallelism allowing the assay to be used for sGnRH measurements in this species. Although all parts of the brain contained measurable levels of sGnRH, the highest concentrations were found in the pituitary, the olfactory bulbs and the telencephalon. These data were confirmed by immunocytochemistry. Cell bodies were found in the olfactory bulbs, ventral telencephalon, preoptic region and mediobasal hypothalamus. Immunoreactive fibers could be observed in all parts of the brain including the optic tectum, the cerebellum (corpus and valvula), the vagal lobe, the medulla oblongata and the rostral spinal cord. In most cases, these fibers do not form well defined bundles; however, there was clearly a continuum of immunoreactive fibers, extending from the olfactory bulbs to the pituitary, and along which all the cell bodies described above were located. In the ventral telencephalon and the preoptic region, clear pictures of varicose positive fibers contacting immunoreactive perikarya could be observed. These data indicate that sGnRH is most likely an endogenous peptide in the brain of the sea bass, although the presence of other forms of GnRH cannot be excluded at this point. This study also demonstrates that the general organization of the GnRH systems in the sea bass is highly similar to what has been described in most freshwater teleost species, and provides basis for further studies on the neuroendocrine control of gonadotrophin release in this commercially important species.     

Comparison of melatonin-binding sites in the brain of two amphibians: an autoradiographic study

Neuroanatomic comparison of the binding capability of 2-[¹²⁵I] iodomelatonin in the crested newt Triturus carnifex Laur. and the green frog Rana esculenta, using quantitative autoradiographic techniques, revealed a heterogeneous distribution pattern. The highest and relatively high binding activities were shown to occur in the optic tracts and in the suprachiasmatic area of the hypothalamus and the optic tectum, respectively, of both species. Low or no 2-[¹²⁵I] iodomelatonin binding values were obtained in the preoptic nucleus, the tuberal hypothalamus, the medulla oblongata, the septum and the dorsal pallium. A differential binding pattern was observed in the amygdaloid nucleus pars lateralis, the striatum and the hindbrain of these amphibians. Indeed, notably high binding levels were shown to occur in the former two brain areas of the crested newt, whereas high levels were displayed in the latter brain region of the green frog. On the basis of elevated quantities of melatonin receptors in mesencephalic, hypothalamic and telencephalic sites, it seems plausible to ascribe some important sensory functions to this receptor system in both species. The remarkably different binding activities in the brain of the two amphibians could be correlated with the simpler cytoarchitectonic brain structure of urodeles and with species-specific variations.     

Distribution of neuropeptide Y-like immunoreactivity in the brain of Salmo salar and Gambusia affinis.

Through the immunohistochemical PAP technique, the distribution of immune positive neurons and fibres for an antibody anti-NPY in the encephalon of salmon fixed in Bouin have been located and studied. NPY-positive neurons are found forming three important nuclei: in the ventrolateral telencephalon; in the tegmentum mesencephali; and in the locus coeruleus. Neurons in the optic tectum, in the thalamic region and a few in the preoptic recess have also been located. The fibres were found throughout the brain, with the exception of the cerebellum, presenting a greater density in three regions: in the dorsal telencephalon; in the mesencephalon; and in the visceral lobes in the rhombencephalon. With the aim of proving if this distribution is found in other groups of teleosts, we processed, with the same technique, the advanced teleost Gambusia affinis, in order to compare it with the primitive teleost Salmo salar. The results show that in both fish this neuropeptide has the same pattern of distribution. The results also suggest that in fish this neuropeptide can be involved in several functions of the central nervous system, as has been demonstrated for mammals. The innervation of the visceral lobes and also the presence of NPY-fibres in the posterior hypothalamus are anatomical supports of the studies which suggest that NPY is related to the control of the food intake.     

Retinopetal neuronal system in the brain of an air-breathing teleost fish,Channa punctata

Summary Retinopetal neurons were visualised in the telencephalon and diencephalon of an air-breathing teleost fish, Channa punctata, following administration of cobaltous lysine to the optic nerve. The labelled perikarya (n=45–50) were always located on the side contralateral to the optic nerve that had received the neuronal tracer. The rostral-most back-filled cell bodies were located in the nucleus olfactoretinalis at the junction between the olfactory bulb and the telencephalon. In the area ventralis telencephali, two groups of telencephaloretinopetal neurons were identified near the ventral margin of the telencephalon. The rostral hypothalamus exhibited retrogradely labelled cells in three discrete areas of the lateral preoptic area, which was bordered medially by the nucleus praeopticus periventricularis and nucleus praeopticus, and laterally by the lateral forebrain bundle. In addition to a dorsal and a ventral group, a third population of neurons was located ventral to the lateral forebrain bundle adjacent to the optic tract. The dorsal group of neurons exhibited extensive collaterals; a few extended laterally towards the lateral forebrain bundle, whereas others ran into the dorsocentral area of the area dorsalis telencephali. A few processes extended via the anterior commissure into the telencephalon ipsilateral to the optic nerve that had been exposed to cobaltous lysine. However, the ventral cell group did not possess collaterals. In the diencephalon, retinopetal cells were visualised in the nucleus opticus dorsolateralis located in the pretectal area; these were the largest retinopetal perikarya of the brain. The caudal-most nucleus that possessed labelled somata was the retinothalamic nucleus; it contained the largest number of retinopetal cells. The limited number of widely distributed neurons in the forebrain, some with extensive collaterals, might participate in functional integration of different brain areas involved in feeding, which in this species is influenced largely by taste, not solely by vision.     

Coordinated evolution of brain size,structure, and eye size in Trinidadian killifish

Brain size, brain architecture, and eye size vary extensively in vertebrates. However, the extent to which the evolution of these components is intricately connected remains unclear. Trinidadian killifish, Anablepsoides hartii, are found in sites that differ in the presence and absence of large predatory fish. Decreased rates of predation are associated with evolutionary shifts in brain size; males from sites without predators have evolved a relatively larger brain and eye size than males from sites with predators. Here, we evaluated the extent to which the evolution of brain size, brain structure, and eye size covary in male killifish. We utilized wild‐caught and common garden‐reared specimens to determine whether specific components of the brain have evolved in response to differences in predation and to determine if there is covariation between the evolution of brain size, brain structure, and eye size. We observed consistent shifts in brain architecture in second generation common garden reared, but not wild caught preserved fish. Male killifish from sites that lack predators exhibited a significantly larger telencephalon, optic tectum, cerebellum, and dorsal medulla when compared with fish from sites with predators. We also found positive connections between the evolution of brain structure and eye size but not between overall brain size and eye size. These results provide evidence for evolutionary covariation between the components of the brain and eye size. Such results suggest that selection, directly or indirectly, acts upon specific regions of the brain, rather than overall brain size, to enhance visual capabilities.     

3H] muscimol receptors sites in the carp (Cyprinus carpio L.) brain: binding assay and autoradiographic distribution

Autoradiographic and binding techniques were used to study the presence of [(3)H]muscimol receptors sites in the carp brain. The radioligand was distributed with an high degree of anatomical selectivity. We found abundant labelling in the cerebellum, in the nucleus diffusus lobi inferioris, and in the torus longitudinalis. No labelling was detected within the epithalamus, thalamus and hypothalamus, while the telencephalon and the rhombencephalon displayed a low density of [(3)H]muscimol receptors sites. Binding assay showed the highest concentration of receptor sites in the nucleus diffusus lobi inferioris and the lowest in the medulla oblongata. Presence of [(3)H]muscimol binding sites within the visceral sensory area was noted. The rank order of displacement efficacy of unlabelled ligands observed, suggested that in brain membranes of carp the receptor binding of [(3)H]muscimol has the same pharmacological specificity previously reported in a large number of experiments with tissue homogenates. A general agreement in binding and autoradiography was observed. The present findings suggested that muscimol receptor could be involved in neuronal pathway controlling basic central actions like gustatory signal processing or spatial learning acquisition and retention.     

Reparative neurogenesis in the brain and changes in the optic nerve of adult trout <Emphasis Type="Italic">Oncorhynchus mykiss</Emphasis> after mechanical damage of the eye

Reparative proliferation and neurogenesis in the brain integrative centers after mechanical eye injury in an adult trout Oncorhynchus mykiss have been studied. We have found that proliferation and neurogenesis in proliferative brain regions, the cerebellum, and the optic tectum were significantly enhanced after the eye injury. The cerebellum showed a significant increase in the proliferative activity of the cells of the dorsal proliferative zone and parenchymal cells of the molecular and granular layers. One week after the injury, PCNA-positive radial glia cells have been identified in the tectum. We have found for the first time that the eye trauma resulted in the development of local clusters of undifferentiated cells forming so called neurogenic niches in the tectum and cerebellum. The differentiation of neuronal cells detected by labeling cells with antibodies against the protein HuC/D occurred in the proliferative zones of the telencephalon, the optic tectum, cerebellum, and medulla of a trout within 2 days after the injury. We have shown that the HuC/D expression is higher in the proliferative brain regions than in the definitive neurons of a trout. In addition, we have examined cell proliferation, migration, and apoptosis caused by the eye injury in the contra- and ipsilateral optic nerves and adjacent muscle fibers 2 days after the trauma. The qualitative and quantitative assessment of proliferation and apoptosis in the cells of the optic nerve of a trout has been made using antibodies against PCNA and the TUNEL method.     

Immunohistochemical localization and biochemical characterization of two novel decapeptides derived from POMC-A in the trout hypothalamus

Several vertebrate species which underwent duplication of their genome, such as trout, salmon and Xenopus, possess two proopiomelanocortin (POMC) genes. In the trout, one of the POMC molecules, called POMC-A, exhibits a unique C-terminal extension of 25 amino acids which has no equivalent in other POMCs characterized so far. This C-terminal peptide contains three pairs of basic residues, suggesting that it may be the source of novel regulatory peptides. The aim of the present study was to investigate the occurrence of these peptides in the brain of the trout Oncorhynchus mykiss by using specific antibodies raised against two epitopes derived from the C-terminal extension of POMC-A, i.e., EQWGREEGEE and YHFQ-NH2. Immunohistochemical labeling of brain sections revealed the presence of EQWGREEGEE- and YHFQ-NH2-immunoreactive cell bodies in the anterior part of the nucleus lateralis tuberis of the hypothalamus. Immunoreactive fibers were observed in the dorsal hypothalamus, the thalamus, the telencephalon, the optic tectum and the medulla oblongata. In contrast, no labeling was detected using antibodies against the non-amidated peptide YHFQG. Biochemical characterization was performed by combining high-performance liquid chromatography (HPLC) analysis with radioimmunoassay (RIA) quantification. Two peptides exhibiting the same retention time as synthetic EQWGREEGEE and ALGERKYHFQ-NH2 were resolved. However, no peptide co-eluting with YHFQ-NH2 or YHFQG could be detected. These results demonstrate that, in the trout brain, post-translational processing of POMC-A generates the two decapeptides EQWGREEGEE and ALGERKYHFQ-NH2. The wide distribution of immunoreactive fibers in the diencephalon, telencephalon, optic tectum and medulla oblongata suggests that these peptides may exert neurotransmitter and/or neuromodulator activities.