Comparative analysis of Met-enkephalin, galanin and GABA immunoreactivity in the developing trout preoptic-hypophyseal system.

We studied the organization of Met-enkephalin-containing cells and fibers in the developing preoptic-hypophyseal system of the brown trout (Salmo trutta fario) by immunohistochemistry and determined the relationship of these cells and fibers to the galaninergic and GABAergic systems. Met-enkephalin immunoreactivity was observed in cells in the preoptic area, the hypothalamus and the pituitary of late larvae. In the hypophysis, a few Met-enkephalin-containing cells were present in all divisions of the adenohypophysis, and some immunoreactive fibers were present in the interdigitations of the neural lobe with the proximal pars distalis. Concurrently, GABAergic fibers innervated the anterior and posterior neural lobe. Galanin cells coexisted with Met-enkephalin cells in neuronal groups of the preoptic-hypophyseal system. Galaninergic and GABAergic fibers innervated the preoptic and hypothalamic areas, but GABAergic fibers containing galanin were not observed. These results indicate that Met-enkephalin, galanin and GABA may modulate neuroendocrine activities in the preoptic area, hypothalamus and pituitary during the transition from larval to juvenile period. To better know how the development of the trout preoptic-hypophyseal system takes place, we studied the patterns of cell proliferation and expression of Pax6, a conserved transcription factor involved in the hypophysis development. Pax6 expressing cells and proliferating cells were present in the Rathke's pouch, the hypothalamus and the hypophysis of early larvae. In late larvae, Pax6 expression was no longer observed in these areas, and the density of proliferating cells largely decreased throughout development, although they remained in the hypophysis of late larvae and juveniles, suggesting that Pax6 might play an important role in the early regionalization of the pituitary in the trout.

Synthesis of estrogens in progenitor cells of adult fish brain: evolutive novelty or exaggeration of a more general mechanism implicating estrogens in neurogenesis?

In contrast to other vertebrates, in which the adult brain shows limited adult neurogenesis, teleost fishes exhibit an unparalleled capacity to generate new neurons as adults, suggesting that their brains present a highly permissive environment for the maintenance and proliferation of adult progenitors. Here, we examine the hypothesis that one of the factors permitting establishment of this favourable environment is estradiol. Indeed, recent data showed that radial glial cells strongly expressed one of two aromatase duplicated genes. Aromatase is the estrogen-synthesizing enzyme and this observation is of great interest, given that radial glial cells are progenitor cells capable of generating new neurons. Given the well-documented roles of estrogens on cell fate, and notably on cell proliferation, these data suggest that estradiol could be involved in maintaining and/or activating these progenitors. Examination of recent data in birds and mammals suggests that the situation in fish could well be an exaggeration of a more general mechanism implicating estrogens in neurogenesis. Indeed, there is accumulating evidence that estrogens are involved in embryonic, adult or reparative neurogenesis in other vertebrates, notably in mammals.

Distribution of choline acetyltransferase (ChAT) immunoreactivity in the central nervous system of a chondrostean, the siberian sturgeon (Acipenser baeri).

All studies to date of cholinergic systems of bony fishes have been done in teleosts. To gain further insight into the evolution of the cholinergic systems of bony fishes, we have studied the brain of a chondrostean fish, the Siberian sturgeon (Acipenser baeri, Brandt), by using an antibody against choline acetyltransferase (ChAT). This study showed the presence of ChAT-immunoreactive (ChAT-ir) neurons in the preoptic region (parvocellular and magnocellular preoptic nuclei and suprachiasmatic nucleus), the periventricular and tuberal hypothalamus, the saccus vasculosus, the dorsal thalamus, and the habenula. The mesencephalic tegmentum contained ChAT-ir cells in the torus semicircularis and torus lateralis. The isthmus contained several cholinergic populations: the nucleus isthmi, the lateral nucleus of the valvula, the secondary visceral nucleus, and the dorsal tegmental nucleus. The motor neurons of the cranial nerves and the spinal motor column were strongly immunoreactive. The medial (sensory) trigeminal nucleus also contained a ChAT-ir neuronal population. The distribution of ChAT-ir neurons in the sturgeon brain showed some notable differences with that observed in teleosts, such as the absence of cholinergic cells in the telencephalon and the optic tectum. Several brain regions were richly innervated by ChAT-ir fibers, particularly the telencephalon, optic tectum, thalamus, posterior tubercle, and interpeduncular nucleus. The hypothalamo-hypophyseal tract, the tract of the saccus vasculosus, the fasciculus retroflexus, and an isthmo-mesencephalo-thalamic tract were the most conspicuous cholinergic bundles. Comparative analysis of these results suggests that teleosts have conserved most traits of the cholinergic system of the sturgeon, having acquired new cholinergic populations during evolution.

Distribution of serotonin (5HT)-immunoreactive structures in the central nervous system of two chondrostean species (Acipenser baeri and Huso huso).

The distribution of serotonin-immunoreactive (5HT-ir) elements was studied in the brain and rostral spinal cord of two chondrosteans, Acipenser baeri and Huso huso, by using an antibody against serotonin. The distribution of these elements was similar in both sturgeon species. In the telencephalon, 5HT-ir cells were found in the olfactory bulb and in the medioventral wall of the telencephalic ventricle, rostral to the anterior commissure, the latter being cerebrospinal fluid-contacting (CSF-C) neurons. The diencephalon contained the highest number of 5HT-ir cell bodies, most of them of CSF-C type, located in the preoptic recess organ, paraventricular organ, posterior recess nucleus, and in the ventromedial thalamus. 5HT-ir non-CSF-C neurons appeared in the dorsal thalamic nucleus. In the brainstem, 5HT-ir neurons were located in four raphe nuclei (dorsal, superior, medial and inferior raphe nuclei) and four lateral reticular nuclei. The dorsal raphe nucleus contained 5HT-ir CSF-C cells, a type of serotoninergic cell that has not been described before in raphe nuclei of fishes or of other vertebrates. CSF-C and non-CSF-C 5HT-ir cells were observed in the spinal cord. 5HT-ir fibers were also widely distributed in the central nervous system of both sturgeon species. Comparison of these results with the distribution of serotoninergic systems in lampreys and other vertebrates suggests that widespread distribution of 5HT-ir cells is a feature of early vertebrate lines.

Distribution of GABA immunoreactivity in the central and peripheral nervous system of amphioxus (Branchiostoma lanceolatum Pallas).

On the basis of labeling with an anti-gamma-aminobutyric acid (GABA) antibody, we report for the first time the presence and distribution of GABA-immunoreactive cells in the central and peripheral nervous system of amphioxus. In the nerve cord, there is a large dorsorostral group of cerebrospinal-fluid-contacting (CSFc) cells at the caudal end of the brain vesicle that gives rise to a large ventral commissure and neuropilar region. In the middle and caudal region of the brain, numerous commissural and CSFc neurons are situated below the region of large dorsal cells. In the spinal cord, several types of GABA-immunoreactive neurons of different size, appearance, and distribution were observed. In the dorsalmost region, very small commissural cells are scattered regularly along the cord. More ventrally in the cord, GABAergic neurons, both of commissural and CSFc cell types, form segmental groups, but scattered cells are observed throughout. These cells give rise to dense longitudinal fascicles of GABAergic fibers and to scattered commissural fibers. The caudal ampulla lacks GABAergic cells and fibers. Some of the fibers of the most rostral and caudal peripheral (sensory) nerves, as well as some sensory cells of the rostral and caudal epidermis, are GABA immunoreactive. The significance of these results for the understanding of the evolution of GABAergic systems of vertebrates is discussed.

The neuronal system of the saccus vasculosus of trout (Salmo trutta fario and Oncorhynchus mykiss): an immunocytochemical and nerve tracing study.

The neuronal system of the saccus vasculosus of two species of trout was studied with immunocytochemical methods and carboindocyanine-dye (DiI) tract-tracing. The cerebrospinal-fluid-contacting neurons of the saccus were immunoreactive for gamma-aminobutyric acid (GABA), glutamic acid decarboxylase (GAD), and neuropeptide Y (NPY). Immunostaining of alternate sections of the saccus vasculosus of fry with anti-GAD and anti-NPY indicated that these substances were colocalized. The tractus sacci vasculosi and the neuropil of the nucleus sacci vasculosi were also immunoreactive to these substances. The GABA, GAD, and neuropeptide Y immunoreactivity of the saccus vasculosus system appeared early in trout ontogeny. After applying DiI to various levels of the tractus sacci vasculosi of adult trout, we observed massive bilateral saccular projections to the nucleus sacci vasculosi and could follow the course of the sacco-thalamic tract. This tract extended in the subependymal region of the thalamus rostral to the nucleus sacci vasculosi and split into two small tracts that reached the subhabenular-preoptic region. Sacco-thalamic fibers formed extensive periependymal plexuses along their trajectory. Interestingly, no clear evidence of the existence of a saccopetal system was obtained. On the basis of these results, we postulate that the saccus vasculosus system modulates the function of centers of the posterior tubercle and periventricular thalamus.

NADPH-diaphorase histochemistry reveals oligodendrocytes in the rainbow trout (teleosts).

We studied the distribution of NADPH-diaphorase (NADPHd) in the brain of the rainbow trout. Not only neurons but also tanycytes and oligodendrocytes showed NADPHd positivity. Interestingly, staining of oligodendrocytes was delicate and revealed fine characteristics of these cells, comparable to those revealed by classical 'oligodendrocyte methods' in mammals (such as the rich branching of processes and their association with myelin sheaths). NADPHd histochemistry indicated that trout oligodendrocytes are heterogeneous as regards cell size, shape and number of processes, and that there is a positive correlation between the size of axons and that of the associated oligodendrocytes. NADPHd histochemistry thus appears to be a useful method for the study of oligodendrocyte populations in bony fishes. Furthermore, this is the first report of NADPHd activity in oligodendrocytes of any vertebrate.

The electric lobes of the electric ray (Torpedo marmorata) are innervated by GABAergic fibres: immunocytochemical evidence for dual innervation of electromotoneurons.

It is currently thought that the electric lobes of electric rays are innervated by a single neuronal system, the oval nucleus system. In the work reported here, the innervation of the electric lobes was studied with silver staining methods, acetylcholinesterase histochemistry and gamma-aminobutyric acid (GABA) immunocytochemistry. Two types of axon were observed in the lobes: thick GABA-immunonegative fibres, which originated from the oval nucleus, and thin GABAergic fibres of unknown origin, here reported for the first time. Electromotoneurons were strongly acetylcholinesterase-positive. Non-GABAergic and non-cholinergic neurons were observed in the oval nucleus, which is innervated by GABA-immunoreactive fibres. These results suggest that GABA may modulate electric discharge both directly, by GABAergic fibres that project to the lobes, and indirectly, by GABAergic fibres that project to the oval nucleus.

The nitric oxide synthase (NOS)-like immunoreactive extrahypophysial projections of the neurosecretory preoptic nucleus of the electric ray (elasmobranchs) suggest a neuroregulatory role for this nucleus.

The extrahypohysial projections of the neurosecretory preoptic nucleus (PON) of the electric ray were studied with the aid of an antibody against nitric oxide synthase (NOS). PON neurons were the only NOS-like-immunoreactive (NOS-ir) cells in the brain. These neurons gave rise to both hypophysial and extrahypophysial NOS-ir projections. Some fibres coursed from the PON to the neurointermediate lobe in the preoptic-hypophysial tract. Other NOS-ir fibres coursed either rostrally or caudally forming terminal fields in the telencephalon (subpallial region), diencephalon (preoptic nucleus, ventrolateral thalamus and posterior recess nucleus), tuberal region (area tegmentalis ventralis and substantia nigra), mesencephalon (lateral tegmentum), rhombencephalon (isthmal nucleus, vagal viscerosensory column and ventrolateral reticular area) and the spinal cord (intermediate horn). The possible involvement of the extrahypophysial PON projections in neuroregulation of visceral centres is discussed.

GABAergic neuronal circuits in the cerebellum of the dogfish Scyliorhinus canicula (Elasmobranchs): an immunocytochemical study.

The presence of GABAergic cells and fibres in the corpus cerebelli and auricles of a dogfish, Scyliorhinus canicula, was studied with immunocytochemistry. In both locations, two types of GABA-immunoreactive (GABA-ir) neurons were observed: stellate cells in the molecular layer and Golgi cells in the granular layer. Stellate cell axons gave rise to numerous GABA-ir boutons distributed throughout the molecular layer and a smaller number of boutons which contacted Purkinje cell perikarya. No GABA-ir baskets around Purkinje cells were observed. Golgi cells of the granular layer gave rise to numerous GABA-ir boutons which were located around cerebellar glomeruli. Purkinje cell perikarya and their axon terminals in the cerebellar nucleus were not GABA-immunoreactive. These findings are discussed in terms of the phylogeny of cerebellar circuits.