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.

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 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.

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.

Primary olfactory fibres project to the ventral telencephalon and preoptic region in trout (Salmo trutta): a developmental immunocytochemical study.

We studied the development of the primary olfactory system of a teleost, the brown trout, with the aims of clarifying whether the caudal projection pertains to the olfactory or to the terminal nerve system, of identifying the brain regions receiving this projection, and of investigating its possible functional significance. As olfactory markers (OMs) we used two polyclonal antibodies (to substance P and to alpha-melanocyte-stimulating hormone) that were found to label the olfactory projection strongly after preadsortion of the antibody with the corresponding antigen (OMs), and as a terminal nerve marker we used an antiserum to FMRF-amide peptide. OM labelling was observed in both perikarya and axons of olfactory neurons. In adults, olfactory neurons projected not only to olfactory glomeruli in the olfactory bulb but also, as has been reported previously, to more caudal targets in the forebrain through the medial olfactory tract. Our results show that these targets include the ventral and commissural nuclei of the area ventralis telencephali, the periventricular preoptic region, and the organum vasculosum laminae terminalis. Glomeruli were not observed before hatching, and the extrabulbar olfactory projections appear late in development. Extensive periventricular preoptic olfactory plexuses and olfactory innervation of the organum vasculosum laminae terminalis did not appear until adulthood. The cells of the ganglion nervus terminalis, which form ganglionic groups along the olfactory nerves, were not stained with these olfactory markers at any developmental stage studied, nor was the medial olfactory tract FMRP-amide peptide immunoreactive. Our results thus confirm the existence of primary olfactory projections to extrabulbar targets in trout. The target regions identified in this study are implicated in sexual behaviour: We discuss the related possibility that, in teleosts, these extrabulbar olfactory projections (rather than projections of the terminal nerve, as is widely held) are the primary mediators of neuroendocrine response to pheromones.

Distribution of calcitonin gene-related peptide-like immunoreactivity in the brain of the small-spotted dogfish, Scyliorhinus canicula L.

The distribution of neuropeptides has been useful in comparing neuronal aggregates of elasmobranchs with those in other vertebrates. The distribution of calcitonin gene-related peptide (CGRP)-like immunoreactivity in the brain of the dogfish was examined with an antiserum to rat alpha-CGRP. Western blot analysis confirms that our antiserum recognizes a single peptide in the dogfish brain very similar to mammalian CGRP. CGRP-like immunoreactivity was located in discrete neuronal groups. CGRP-like-immunoreactive (CGRP-ir) neurons were found in the motor nuclei III, IV, V, VI, VII, IX, and X of the brainstem motor column and in the octavolateral efferent neurons. In the isthmal region, two groups of CGRP-ir neurons appeared in the parabrachial region and reticular substance. Three other CGRP-ir cell groups were observed in the mesencephalon: in the ventral tegmental area, in the substantia nigra, and one widely scattered but numerous population in superficial layers of the optic tectum. In the diencephalon, CGRP-ir cells were observed in the magnocellular preoptic nucleus and the organon vasculosum hypothalami. A population of CGRP-ir cells was also observed in the entopeduncular nucleus in the impar telencephalon. CGRP-ir fibers of central origin were widely distributed in the brain, but the most conspicuous areas were found in the ventral telencephalon, the hypothalamus, the mesencephalic lateral reticular area, and the dorsolateral isthmal region. The neurointermediate lobe of the hypophysis was also richly innervated by CGRP-ir fibers. CGRP-ir sensory fibers of cranial nerves IX and X and of dorsal spinal roots formed very conspicuous terminal fields in the lobus vagi and Cajal's nucleus commissuralis and in the dorsal region of the substantia gelatinosa, respectively. Comparison of the distribution of fibers and perikarya in dogfish and other vertebrates suggests that this CGRP-ir system has been well conserved during evolution.

Distribution of choline acetyltransferase (ChAT) immunoreactivity in the brain of the adult trout and tract-tracing observations on the connections of the nuclei of the isthmus.

The distribution of cholinergic neurons and fibers was studied in the brain and rostral spinal cord of the brown trout and rainbow trout by using an antiserum against the enzyme choline acetyltransferase (ChAT). Cholinergic neurons were observed in the ventral telencephalon, preoptic region, habenula, thalamus, hypothalamus, magnocellular superficial pretectal nucleus, optic tectum, isthmus, cranial nerve motor nuclei, and spinal cord. In addition, new cholinergic groups were detected in the vascular organ of the lamina terminalis, the parvocellular and magnocellular parts of the preoptic nucleus, the anterior tuberal nucleus, and a mesencephalic tegmental nucleus. The presence of ChAT in the magnocellular neurosecretory system of trout suggests that acetylcholine is involved in control of hormone release by neurosecretory terminals. In order to characterize the several cholinergic nuclei observed in the isthmus of trout, their projections were studied by application of 1,1;-dioctadecyl-3,3,3;, 3;-tetramethylindocarbocyanine perchlorate (DiI) to selected structures of the brain. The secondary gustatory nucleus projected mainly to the lateral hypothalamic lobes, whereas the nucleus isthmi projected to the optic tectum and parvocellular superficial pretectal nucleus, as previously described in other teleost groups. In addition, other isthmic cholinergic nuclei of trout may be homologs of the mesopontine system of mammals. We conclude that the cholinergic systems of teleosts show many primitive features that have been preserved during evolution, together with characteristics exclusive to the group.

Distribution of substance P-like immunoreactivity in the brain of the elasmobranch Scyliorhinus canicula.

Immunohistochemical methods were used to study the distribution of substance P in the brain of the small-spotted dogfish (Scyliorhinus canicula). Substance P-like immunoreactive (SP-IR) cell bodies and fibers were widely distributed. In the telencephalon, sparse populations of SP-IR neurons are present in the olfactory bulbs, pallium, and subpallium. In the subpallium numerous SP-IR boutons form unusual coats ("pericellular appositions") on SP-immunonegative neurons. In the diencephalon numerous SP-IR cerebrospinal fluid-contacting neurons are present in the preoptic recess organ and organon vasculosum hypothalami. Numerous SP-IR fibers also run in the hypothalamus, although no immunoreactivity was observed in the habenulo-interpeduncular system. A terminal field of SP-IR fibers is present in the median eminence. In the mesencephalic tegmentum, SP-IR neurons were observed in the Edinger-Westphal nucleus. SP-IR fibers are present at high density in the basal tegmentum, forming a conspicuous tract. In the hindbrain, numerous SP-IR fibers were observed in the isthmal region, the trigeminal descending root, the visceral sensory area and commissural nucleus, and the visceromotor column. SP-IR fibers occur at high density in the substantia gelatinosa of the rostral spinal cord.

Distribution of choline acetyltransferase immunoreactivity in the brain of an elasmobranch, the lesser spotted dogfish (Scyliorhinus canicula).

Although the distribution of cholinergic cells is remarkably similar across the vertebrate species, no data are available on more primitive species, such as cartilaginous fishes. To extend the evolutionary analysis of the cholinergic systems, we studied the distribution of cholinergic neurons in the brain and rostral spinal cord of Scyliorhinus canicula by immunocytochemistry using an antibody against the enzyme choline acetyltransferase (ChAT). Western blot analysis of brain extracts of dogfish, sturgeon, trout, and rat showed that this antibody recognized similar bands in the four species. Putative cholinergic neurons were observed in most brain regions, including the telencephalon, diencephalon, cerebellum, and brainstem. In the retrobulbar region and superficial dorsal pallium of the telencephalon, numerous small pallial cells were ChAT-like immunoreactive. In addition, tufted cells of the olfactory bulb and some cells in the lateral pallium showed faint immunoreactivity. In the preoptic-hypothalamic region, ChAT-immunoreactive (ChAT-ir) cells were found in the preoptic nucleus, the vascular organ of the terminal lamina, and a small population in the caudal tuber. In the epithalamus, the pineal photoreceptors were intensely positive. Many cells of the habenula were faintly ChAT-ir, but the neuropil of the interpeduncular nucleus showed intense ChAT immunoreactivity. In the pretectal region, ChAT-ir cells were observed only in the superficial pretectal nucleus. In the brainstem, the somatomotor and branchiomotor nuclei, the octavolateral efferent nucleus, and a cell group just rostral to the Edinger-Westphal (EW) nucleus contained ChAT-ir neurons. In addition, the trigeminal mesencephalic nucleus, the nucleus G of the isthmus, some locus coeruleus cells, and some cell populations of the vestibular nuclei and of the electroreceptive nucleus of the octavolateral region exhibited ChAT immunoreactivity. In the reticular areas of the brainstem, the nucleus of the medial longitudinal fascicle, many reticular neurons of the rhombencephalon, and cells of the nucleus of the lateral funiculus were immunoreactive to this antibody. In the cerebellum, Golgi cells of the granule cell layer and some cells of the cerebellar nucleus were also ChAT-ir. In the rostral spinal cord, ChAT immunoreactivity was observed in cells of the motor column, the dorsal horn, the marginal nucleus (a putative stretch-receptor organ), and in interstitial cells of the ventral funiculus. These results demonstrate for the first time that cholinergic neurons are distributed widely in the central nervous system of elasmobranchs and that their cholinergic systems have evolved several characteristics that are unique to this group.

Distribution of somatostatin-immunoreactivity in the brain of the larval lamprey (Petromyzon marinus).

The detailed distribution of somatostatinergic neurons and fibre tracts in the brain of larval lamprey was studied in serially sectioned material using immunocytochemical techniques. Neurons were found to be arranged in four nuclei: a hypothalamic nucleus consisting of both small cerebrospinal fluid-contacting neurons and larger non-contacting neurons, a thalamomesencephalic nucleus and two isthmotrigeminal reticular nuclei. The hypothalamic nucleus is the first to differentiate. Analysis of young larvae showed that somatostatin-immunoreactivity first appeared in hypothalamic cells (12 mm larvae), while it appeared later in the other nuclei. The different somatostatin-immunoreactive fibre tracts innervate different regions of the brain. In addition, somatostatin-immunoreactive fibres originating from hypothalamic neurons were found in the anterior neurohypophysis, which suggests the presence of a hypothalamohypophysial somatostatinergic system in lampreys.

Differential expression of thymosins beta(4) and beta(10) during rat cerebellum postnatal development.

The beta-thymosins are a family of actin monomer-sequestering proteins widely distributed among vertebrate classes. The most abundant beta-thymosins in mammalian species are thymosin beta(4) (Tbeta(4)) and thymosin beta(10) (Tbeta(10)), two small peptides (43 amino acids) sharing a high degree of sequence homology. In the present work, we have analyzed the distribution of Tbeta(4) and Tbeta(10) in the developing and adult rat cerebellum using in situ hybridization and immunohistochemistry techniques. Our results show that the temporal and cellular patterns of expression of both beta-thymosins are different. In the young (7 and 18 postnatal days) and adult (1 and 4 months old) rat cerebellum, Tbeta(4) was mainly expressed in the glia (microglia, Golgi epithelial cells and oligodendrocytes), neurons (granule cells and Purkinje cells), and in the capillaries. In 14-month-old rats, the Tbeta(4) immunoreactivity was only detected in some microglia cells. In young and adult animals, most of the Tbeta(10) immunoreactivity was localized in several types of neuronal cells including granule cells, Golgi neurons and Purkinje cells. In old animals, a faint Tbeta(10) signal could be detected in a few Purkinje cells. Our results suggest that each beta-thymosin could play a different function in the control of actin dynamics.

Neurons of the olfactory organ projecting to the caudal telencephalon and hypothalamus: a carbocyanine-dye labelling study in the brown trout (Teleostei).

The caudal extrabulbar projections and their neurons of origin in the trout were studied after carbocyanine-dye (DiI) labelling in either the olfactory organ or the caudal telencephalon. DiI application to the caudal telencephalon labelled bipolar neurons in the olfactory epithelium, where they were sparsely distributed throughout the olfactory lamellae. Labelled fibres ran scattered in the olfactory nerve without forming bundles. DiI application to the olfactory organ labelled extrabulbar projections to the ventral telencephalon, preoptic region and tuberal hypothalamus. These results confirm that primary sensory fibres running in the medial olfactory tract of trout have an olfactory origin.

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.

Ontogeny of somatostatin-immunoreactive systems in the brain of the brown trout (Teleostei).

The development of somatostatin-immunoreactive neurons and fibres was studied, using immunocytochemistry, in the brain of the brown trout. Somatostatinergic perikarya were found in many regions including several telencephalic areas, the preoptic nucleus, anterior tuberal and lateral tuberal nuclei, the lateral recess nucleus, dorsal tuberal nucleus, the pre- and pseudoglomerular nuclei, central thalamic nucleus, optic tectum, interpeduncular nucleus, several isthmal and reticular nuclei and the solitary fascicle nucleus. The ventrolateral area of the telencephalon and the nucleus lateralis tuberis are the first immunoreactive nuclei to appear in ontogeny, and cells of some telencephalic areas and of the lateral optic recess nucleus, the latest. Somatostatin-immunoreactive fibre tracts innervate the hypophysis and different regions of the brain. The most richly innervated areas in adults are the dorsolateral telencephalic area and the organon vasculosum laminae terminalis. Two patterns of production of somatostatinergic cells were observed: that of populations in which cell numbers increase over the lifetime of the fish, and that of populations whose cell number is established early in development or even diminishes in adulthood. These results provide interesting contrasts to those previously reported in birds and mammals.

Asymmetric distribution of calbindin-D28K in the ganglia habenulae of an elasmobranch fish.

Using an antibody raised against a purified chick duodenal vitamin D-dependent calcium-binding protein, the presence and distribution of calbindin has been studied immunohistochemically in the habenular ganglia of the dogfish. In the more developed left ganglion, a positive reaction was clearly observed in the neurons of the medial nucleus, whereas in the lateral nucleus, only some scarce, hardly immunostained cells appeared. In the neurons of the right habenula however, no immuno-reactivity was observed. The distribution of vitamin D-dependent calcium-binding protein in the dogfish habenulae is therefore asymmetrical. This may be due to differences in the neuronal activity between the two ganglia.

Aminergic neurons in the hypothalamus of the dogfish, Scyliorhinus canicula L. (Elasmobranch). A histofluorescence study.

Aminergic neurons of the hypothalamus of Scyliorhinus canicula were studied using formaldehyde-induced fluorescence. An aminergic organ of the preoptic recess (PRO) consists of both cerebrospinal fluid (CSF)-contacting and non-contacting green-yellow fluorescent neurons of bipolar or multipolar morphologies. Caudal to the optic chiasma a group of faint fluorescent neurons is located among the fibres of the commissura transversa. In the posterior hypothalamus the infundibular wall has numerous aminergic CSF-contacting neurons along lines of diverticles. Scarce CSF-contacting neurons are also found in the roof and floor of the recesses of the inferior hypothalamic lobes. The folded posterior recess has a very high number of aminergic CSF-contacting neurons of bipolar shape. Its coarse layer of cell bodies is covered by a large amount of fluorescent fibres which form a conspicuous cap-like band. These structures are discussed from a comparative point of view in relation with aminergic hypothalamic systems of other anamniote vertebrates.

A freeze-fracture study of the soma membranes of monoaminergic neurons in the hypothalamic nucleus recessi posterioris of the rainbow trout (Salmo gairdneri Rich.).

Monoaminergic neurons in the hypothalamic nucleus recessi posterioris of the rainbow trout were studied by freeze-fracturing. The size and distribution of intramembrane particles in the nuclear and plasma membranes and the density of nuclear pores in the nuclear envelope were determined. The density of nuclear pores in these cells was significantly greater than in the non-granulated neurons of this nucleus. The density of IMPs in the P face of the inner nuclear membrane was significantly greater than in the P face of the outer nuclear membrane; however in the E faces the density is greater in the outer nuclear membrane but the differences are not statistically significant. In both membranes, the proportion of large particles (greater than 9.6 nm) was greater in the E than in the P faces. The coefficient of partition between the plasma membrane faces was 1.97 +/- 0.24 while in non-granulated neurons was 4.45 +/- 0.51. There are significant differences among these values. Interestingly, the coefficient of partition of the plasma membrane in monoaminergic neurons was lesser than in some other types of neurons. Postsynaptic areas or other membrane specializations were not found on the cell body plasma membrane, which corroborates the results of conventional transmission electron microscopy.

Organization of catecholaminergic systems in the hypothalamus of two elasmobranch species, Raja undulata and Scyliorhinus canicula. A histofluorescence and immunohistochemical study.

We examined the organization of catecholaminergic neurons in the hypothalamus of the painted ray, Raja undulata, and the small-spotted dogfish, Scyliorhinus canicula, with the use of formaldehyde-induced fluorescence (FIF) methods and tyrosine hydroxylase (TH) immunohistochemistry. In both species we identified distinct populations of catecholamine-containing neurons differing in a) their immunoreactivity to antibodies against the enzyme tyrosine hydroxylase (TH), b) their fluorescence in response to FIF methods for the detection of catecholamines, and c) their relationship with the third ventricle. One population is made up of FIF-positive and TH-negative neurons (most of which are CSF [cerebrospinal fluid]-contacting) and located in two circumventricular organs, the preoptic recess organ and the organon vasculosum hypothalami. Another population comprises TH-immunoreactive (TH-IR), FIF negative neurons that are located in the suprachiasmatic nucleus and the posterior tuberculum and are not related to the third ventricle recesses. A third population of TH-IR, CSF-contacting neurons is also present in the organon vasculosum hypothalami. The existence of three catecholaminergic populations suggests differences in the metabolism of catecholamines and/or different functions. The circumventricular neurons are not associated with the hypophysis and appear to accumulate catecholamine (dopamine) obtained from exogenous sources. In both Raja and Scyliorhinus the neurointermediate lobe is innervated by TH-IR fibres originating from dopamine-synthesizing neurons of the second catecholaminergic population.