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A centrifugal visual system showing FMRF-amide-like immunoreactivity has been demonstrated in Lampetra fluviatilis by using immunocytochemical and hodological techniques. From 50 to 60 immunoreactive neurons, labelled after contralateral intraocular injection of rhodamine beta-isothiocyanate, form a small, clearly defined, nucleus in the lateral neural plate of the magnocellular preoptic nucleus. These cells give rise to immunoreactive axons which have been observed at the base of the nucleus, in the optic chiasma and in the optic nerve, to project into the intermediate plexiform layer of the retina, which separates the layer of internal horizontal cells from the layer of external horizontal cells. This FMRF-amide-like immunoreactive centrifugal visual system is compared to that described in Gnathostomes.

Thin varicose centrifugal visual fibers, between 30-45 in number and displaying cGnRH-I immunoreactivity, were identified in Crocodylus niloticus. Approximately 80% of these fibers were also FMRF-amide-like immunoreactive. The cGnRH-I fibers extended from the preoptic region to the retina where they appeared to terminate in the external portion of the inner plexiform layer. The location of their neurons of origin could not be determined precisely following the intraocular injection of the retrograde axonal tracer RITC. Nevertheless, the presence of cGnRH-I-immunoreactive neurons exclusively within the complex comprising the terminal nerve and the septo-preoptic region, and of several retinopetal fibers labelled retrogradely with the axonal tracer at the septo-preoptic junction, indicates that the cGnRH-immunoreactive centrifugal visual system originates from within this complex.

Serotonin (5-HT) immunoreactive (-ir) profiles within the isthmo-optic nucleus (ION) of the centrifugal visual system (CVS) were studied in the pigeon using light microscopic immunohistofluorescent and electron microscopic immunocytochemical pre-embedding techniques. The brainstem origin of the 5-HT input upon the ION was determined by combining 5-HT immunohistofluorescence (FITC) and retrograde transneuronal tracing after intraocular injection of Rhodamine beta-isothiocyanate. The light microscopic results showed that 5-HT endings were mainly localised within the neuropillar zones of the ventral ION. The 5-HT-ir cell bodies, belonging to a lateral extension of the dorsal raphe system, were observed within the same region as the centrifugal ectopic neurons (EN) underlying the ION and some displayed dendritic processes which penetrated the nucleus. Double-labeled neurons, representing 5-HT-ir afferents to the ION, were identified only within the n. linearis caudalis region of the ventral raphe. The electron microscopic results confirmed the presence of 5-HT-ir dendritic processes within the ventral part of the nucleus and showed that they were contacted by axon terminals belonging to intrinsic interneurons. The functional organisation of the ION and the possible contribution of serotonergic raphe afferents and efferents are discussed in relation to present hypotheses linking the avian CVS to mechanisms of visual attention.

The present study demonstrated a direct serotonergic retinopetal projection in the mouse stemming from the lateral portion of the dorsal raphe nucleus bilaterally. A double-labeling technique was employed combining: (1) radioautography and retrograde axonal tracing following intraocular injection of [(3)H] 5-HT and (2) immunocytochemical identification of endogenous 5-HT. Radiolabeled neurons were only observed within the dorsal raphe nucleus and were always double-labeled with the 5-HT antibody. The radiolabeling appeared to be specific resulting from the retrograde transport of a radioactive 5-HT derivative product following uptake of the neurotransmitter by intraretinal terminals.

The distribution of retinal ganglion cells (RGCs) providing input to the thalamofugal visual system in the pigeon was studied with an anatomical transneuronal transport technique using the fluorescent dye rhodamine beta-isothiocyanate (RITC). Unilateral injections of RITC made into the telencephalic visual Wulst resulted in the retrograde (1) first-order labeling (FOL) of dorsal thalamic (n. dorsolateralis anterior and n. superficialis parvocellularis: SPC) and brainstem somata as well as (2) second-order labeling of other cell populations within the brain and of retinal ganglion cells in both eyes obtained after transneuronal transfer of the tracer from neurons labeled directly via FOL. The mapping and counting of labeled RGCs in retinal flat-mounts showed that they were mainly distributed within the nasal portion of the retinal yellow field (YF) and that their total numbers were consistently higher (averaging 57%) in the eye contralateral to the tracer injection. Labeled RGCs in the retinal red field (RF) represented 13.4% and 12.0% of total labeled cells in the ipsilateral and contralateral eye, respectively. Moreover, the average densities of labeled cells/mm(2) in the RF and YF were respectively 8.4 and 42.8 (ipsilateral) and 17.9 and 54.0 (contralateral). The preferential distribution of labeled RGCs within the nasal YF supports the notion that the thalamofugal visual system in the lateral-eyed pigeon is mainly concerned with viewing in the lateral visual field. Conversely, the relatively low numbers of labeled RGCs observed within the specialized RF indicate that, unlike the case in frontal-eyed bird species and mammals, this system does not appear to be involved in binocular visual processing.

A small contingent of 30–50 of centrifugal visual fibres, showing FMRF-amide-like immunoreactivity, has been identified in C. niloticus; these fibres extend from the chiasmatic region into the retina. They do not take the marginal optic tract, but pass medially to the chiasmatic fascicles, from the preoptic region. The cells of origin of these fibres have not been identified. However, none of the retinopetal neurons of the brainstem [M. Medina, J. Reperant, R. Ward, D. Miceli, Centrifugal visual system of Crocodylus niloticus : a hodological, histochemical and immunocytochemical study, J. Comp. Neurol. 468 (2004) 65–85], labelled by retrograde transport of rhodamine β-isothiocyanate after intraocular injection of this tracer, show FMRF-amide-like immunoreactivity; neither are any of the FMRF-amide-like immunopositive neurons in the crocodile brain, particularly those of the complex involving the terminal nerve and the septo-preoptic region, labelled by rhodamine after its intraocular injection.

The distribution of Kin protein, the vertebrate homologue of the bacterial recA nuclear protein involved in illegitimate recombinant DNA repair and gene regulation, was analysed in the brain of the mouse, quail, turtle and frog by immunocytochemical methods. The protein was expressed in all brains, but not in a uniform manner. Immunoreactivity was absent from major fibre tracts. In the cerebral nuclei, immunolabelling in the various species showed an important variation. A comparative analysis, based on the homologies between different brain structures in these species, showed that this variation was not due to interspecific variation but that of an ancestral pattern of distribution of Kin protein. It is also shown that whatever the species examined, Kin protein is consistently more highly expressed in those regions of the brain with a conservative evolutionary history (e.g. the olfactory and limbic systems, the hypothalamus, the monoaminergic system, the cerebellum, and the nuclei of sensory and motor cranial nerves). The protein is markedly less heavily expressed in the dorsal striatum and the sensory nuclei of the thalamus.

The ultrastructural features of serotonergic fibers, terminals and synaptic contacts were studied with the pre-embedding immunocytochemical method in the isthmo-optic nucleus of the pigeon centrifugal visual system. The 5-HT immunoreactive (-ir) profiles were diffusely distributed and their density was low. The labeled axons were thin and unmyelinated (mean diameter=0.21+/-0.03 microm) though a few larger myelinated axons were observed (mean diameter=0.51+/-0.07 microm). The 5-HT-ir terminals or varicosities were small (diameter=0.71+/-0.54 microm) and contained small agranular synaptic vesicles (diameter=28.5+/-6.9 nm) and large granular vesicles (diameter=102.2+/-19.5 nm). The latter only constituted approximately 1% of the total profiles containing synaptic vesicles in the isthmo-optic nucleus. In single thin sections, only 5% of the 5-HT-ir varicosities exhibited an active asymmetrical zone synapsing upon dendritic profiles of centrifugal visual neurons. Calculations indicated that 17% of these 5-HT-ir varicosities were actually engaged in junctional synaptic relationships, whereas the remaining (83%) were nonjunctional. The data suggest that, within the isthmo-optic nucleus, 5-HT acts both at synaptic junctions (wiring transmission) and at a distance via the extracellular space (volume transmission). These 5-HT afferents could thus modulate the activity of the retinopetal neurons and visual information processing.

A quantitative electron microscopic analysis of glutamate (GLU) immunoreactivity using the post-embedding immunogold technique was carried out within the isthmo-optic nucleus (ION) of the pigeon centrifugal visual system (CVS). Measurements were performed in each of eight different categories of axon terminals, including those that were GABA-immunoreactive (-ir), considered representing control profiles and identified using a single or double-label immunocytochemical procedure. The results demonstrated that the glutamate immunogold particle densities for both mitochondrial and vesicular pools and for total surface area of bouton profiles were significantly higher in P1a, P1b and P2b terminals and not significantly different in P4 and P5 terminals compared to those recorded in control GABA-ir terminals (P2a, P2c, P3). Moreover, the values measured in GLU-ir positive profiles were all significantly higher than in either P4 or P5 terminals. The results suggest that tectal neurons, which provide the main input to the ION cells, are either inhibitory GABA-ir possibly associated with P2c and/or P3 terminals or excitatory GLU-ir via P1a, P1b and P2b terminals. Such differential effects of tectal afferents may be the basis for the modulation of centrifugal activity and consequently of end target retinal ganglion cell responses. The data are relevant to hypotheses implicating the avian CVS in mechanisms of selective enhancement of visual attention to either novel or meaningful stimuli within the visual field.

The retinal projections of Caiman crocodilus and Crocodylus niloticus were investigated by means of the orthograde axonal transport of either rhodamine beta-isothiocyanate or tritiated proline. In these two species, each tracer revealed contralateral retinal projections to three hypothalamic regions (subventricular gray matter, nucleus suprachiasmaticus, and area optica hypothalami lateralis), five thalamic regions (nuclei ovalis, dorsolateralis anterior, ventrolateralis and ventrobasalis, and lateral geniculate complex, of which six subcomponents can be distinguished), six pretectal regions (nuclei posterodorsalis, lentiformis mesencephali, griseus tectalis, geniculatus pretectalis, area optica commissurae posterior and area optica pretectalis lateroventralis), six outermost layers of the optic tectum, and the nucleus opticus tegmenti. Weak ipsilateral retinal projections have been observed in two hypothalamic nuclei and in the nucleus opticus tegmenti. Comparative analysis with other data show that the contralateral retinal projections of crocodiles are considerably more reptilian than avian. Moreover, crocodiles share with birds an extremely poor contingent of ipsilateral retinal projections.

Immunohistochemical techniques were used to determine the distribution of cholecystokinin-8 (CCK8) immunoreactivity in the brain of the chameleon. In the telencephalon, CCK8-immunopositive somata were sparse and observed principally in the olfactory tubercle at the ventromedial edge of the rostral telencephalon and in the medial septum. Immunopositive fibers were observed mainly in the medial septal region and the ventral telencephalon. In the diencephalon, numerous CCK8-reactive fibers were densely concentrated in the periventricular region, the dorsolateral hypothalamus and the external zone of the median eminence. In the thalamus, labelled fibers were restricted to the peri-rotundal nuclei and the lateral part of the habenula. Immunoreactive cell bodies were observed in the medial part of the dorsal lateral geniculate nucleus, in the periventricular, ventral and lateral regions of the hypothalamus. In the mesencephalon, the densest accumulations of immunopositive fibers were observed in the area pretectalis, the periventricular gray matter, the medial tegmentum and the isthmus. Labelled neurons were observed in the deep, and occasionally intermediate, tectal layers and in the laminar nucleus of the torus semicircularis. In the rhombencephalon, labelled fibers were observed at the highest density in the central gray matter and the locus coeruleus; labelled somata were observed only in the nucleus of the tractus solitarius.

The distribution of GABA-immunoreactivity was studied in the brain of the silver eel (Anguilla anguilla) by means of antibodies directed against GABA. Immunoreactive neuronal somata were distributed throughout the brain. Positive perikarya were detected in the internal cellular layer of the olfactory bulb, and in all divisions of the telencephalon, the highest density being observed along the midline. Numerous GABA-reactive cell bodies were found in the diencephalon, particularly in the preoptic and tuberal regions of the hypothalamus, and the dorsolateral, dorsomedial and ventromedial thalamic nuclei. In the optic tectum, the majority of GABApositive cell bodies were located in the periventricular layer. A number of immunolabelled cell bodies were observed in different tegmental structures, notably the torus semicircularis. In the cerebellum, the Purkinje cells were either very intensely or very weakly immunoreactive. In the rhombencephalon, reactive cell bodies were observed in the eminentia granularis, the valvula cerebellaris, the octavolateral nucleus, the lobus vagus and in the vagal and glossopharyngeal motor nuclei. Intensely immunoreactive axons and terminals were observed in the external granular layer and internal cellular layer of the olfactory bulb. In the telencephalon, the highest density of reactive fibres and boutons was found in the fields of the medial wall. Many immunolabelled fibres were seen in the medial and lateral forebrain bundles. In the diencephalon, intense labelling of fibres and terminals were observed in the nuclei situated close to the midline. In the optic tectum the highest density of reactive fibres was seen in the sfgs, the layer to which the retina projects massively. Finally, in the rhombencephalon the strongest labelling of neurites was observed in the nuclei of the raphe, the nucleus octavocellularis magnocellularis and the nuclei of the IXth and Xth cranial nerves. The GABAergic system of the eel, which is well developed, appears to be generally comparable to that described in tetrapod vertebrates.

The retinal projections of two species of flatfish (Scophthalmus maximus, Scophthalmidae; Platichthys flesus, Pleuronectidae) were investigated by autoradiography and by a HRP technique. Contralateral projections to five hypothalamic centres (area optica preoptica ventralis, nucleus opticus preopticus parvocellularis posterior pars lateralis, n. suprachiasmaticus, n. opticus hypothalami ventromedialis and area optica hypothalami posterior), thirteen thalamo-pretectal centres (nucleus opticus dorsolateralis (partes medialis, ventralis and lateralis), n. opticus ventrolateralis, n. opticus commissurae posterioris (partes dorsalis and ventralis), n. opticus accessorius, n. geniculatus lateralis mesencephali, nn. opticus pretectalis dorsalis, medialis and ventralis and n. corticalis), three layers of the optic tectum (stratum opticum pars externa, stratum fibrosum et griseum superficiale, stratum album centrale), and a single target in the tegmentum (n. opticus tegmenti mesencephali dorsalis), were identified in both species. Interspecific variation of the contralateral visual projections is relatively small. Ipsilateral visual projections of fibres which recross the midline in the minor and transverse commissures were also identified; in S. maximus this ipsilateral contingent is poorly developed and concerns principally hypothalamic structures, while in P. flesus the ipsilateral projections are considerably more extensive and involve both hypothalamic and thalamo-pretectal primary visual centres. No differences in the projections from the fixed and from the migrated eye were observed in either species. The findings are discussed in the general context of the existing literature on the visual projections of teleosts, in an attempt to characterize the primary visual system of the Pleuronectiformes in an evolutionary context.