Central projections of the nervus terminalis in the bichir,Polypterus palmas

Summary Central pathways of the nervus terminalis (n.t.) in the bichir, Polypterus palmas, were studied with the use of tracing techniques. After application of horseradish peroxidase to the unilateral olfactory mucosa labeled n.t. fibers were traced in seven distinct bundles through the subpallium. Projection areas are found in the precommissural ventral nucleus of the area ventralis telencephali ipsilaterally, the anterior commissure and commissural parts of the periventricular preoptic nucleus bilaterally; few n.t.-fibers cross via the anterior commissure to the contralateral side; no fibers were observed to turn rostrally to the contralateral olfactory bulb. Major targets of the n.t. include a restricted ventral part of the periventricular preoptic nucleus at the level of the optic chiasma bilaterally, and the periventricular nuclei located between the thalamic nuclei and the hypothalamus bilaterally. N.t. fibers continue their course through the ipsilateral hypothalamus and are traced as far as the mesencephalic tegmentum ipsilaterally. N.t. terminations are found consistently within the boundaries of periventricular cell nuclei, suggesting axosomatic synaptic contacts. We propose a differentiation of the n.t. ganglion cells into a distal (mucosal) and proximal (bulbar) type regarding the peripheral cell processes. Our findings are compared with those of other reports on the n.t. system.     

The nervus terminalis also exists in cyclostomes and birds

Summary The terminal nerve has been described in all vertebrate classes, with the exception of cyclostomes and birds. With regard to this question, we have examined representatives of these two classes using tracer techniques, and found a terminal nerve in larval lampreys and young domestic mallards. Horseradish peroxidase or cobaltous lysine was injected into the olfactory mucosa, which is known to be innervated by peripheral branches of the terminal nerve. The brains were then searched for labeled, centrally directed fibers of the terminal nerve that project further caudally than the glomerular layer of the olfactory bulb. In larval lampreys, centrally projecting fibers of the terminal nerve were found in the tel-, diand mesencephalon. Termination of labeled fibers was observed in the hypothalamus. Some fibers of the terminal nerve cross to the contralateral side via the commissure of the posterior tuberculum. In young ducks, the terminal nerve projects ipsilaterally along the medial edge of the telencephalon.     

Central connections of the olfactory bulb in the goldfish,Carassius auratus

Summary The central connections of the goldfish olfactory bulb were studied with the use of horseradish peroxidase methods. The olfactory bulb projects bilaterally to ventral and dorsolateral areas of the telencephalon; further targets include the nucleus praeopticus periventricularis and a caudal olfactory nucleus near the nucleus posterior tuberis in the diencephalon, bilaterally. The contralateral bulb and the anterior commissure also receive an input from the olfactory bulb. Contralateral projections cross in rostral and caudal portions of the anterior commissure and in the habenular commissure. Retrogradely labeled neurons are found in the contralateral bulb and in three nuclei in the telencephalon bilaterally; the neurons projecting to the olfactory bulb are far more numerous on the ipsilateral side than in the contralateral hemisphere. Afferents to the olfactory bulb are found to run almost entirely through the lateral part of the medial olfactory tract, while the bulb efferents are mediated by the medial part of the medial olfactory tract and the lateral olfactory tract. Selective tracing of olfactory sub-tracts reveals different pathways and targets of the three major tract components. Reciprocal connections between olfactory bulb and posterior terminal field suggest a laminated structure in the dorsolateral telencephalon.     

A light- and electron-microscopic study of mesencephalic neurons projecting to the ganglion of the nervus terminalis in the goldfish

Summary After application of various neuronal tracers (horseradish peroxidase, cobalt-chloride lysine, true blue) to the ganglion of the nervus terminalis a small number of neurons was retrogradely labeled in the mesencephalon. As revealed by combined horseradish peroxidase and catecholamine-fluorescence techniques these neurons are located in the isthmic area immediately rostral to, but not within the locus coeruleus. Cobalt-labeled axons of the mesencephalic neurons were traced individually in serial sections. Neurons projecting contralaterally cross in the horizontal commissure. Tracing of single fibers provided no evidence for axon collaterals within this pathway. Retrograde labeling reveals two different types of isthmic neurons afferent to the ganglion of the nervus terminalis: One smaller-sized type is located bilaterally and consists of four to six neurons; another type possessing many dendritic processes was consistently found as only one single cell located contralateral to the side of injection. The existence of two types of neurons was confirmed by their cytological differences: The small-sized type receives only sparse perisomatic input, while the large-sized type shows heavy somatic and dendritic, probably monoaminergic innervation.     

Olfactory and vomeronasal projections and the pathway of the nervus terminalis in ten species of salamanders

Summary Primary olfactory and vomeronasal projections as well as the pathway of the nervus terminalis were studied in 10 representative species of salamandrid and plethodontid salamanders by means of injections of horseradish peroxidase and examination of whole-mount preparations. Olfactory projections are very similar in the different urodeles, but vomeronasal projections differ in shape and number of termination fields. Whereas the direct-developing Plethodontini and Bolitoglossini reveal only one or two fields, the salamandrid species and the members of the plethodontid tribes Desmognathinae and Hemidactyliini, all possessing an aquatic larval stage, exhibit several vomeronasal projection fields. In all species examined centrifugal axons of the nervus terminalis leave the olfactory projection area ventrocaudally and terminate in the preoptic region and the hypothalamus.Abbreviations COM. ANT commissura anterior - DGL displaced glomeruli - HY hypophysis - HYTH hypothalamus - LF lateral fibers of the nervus terminalis - ME medulla oblongata - MF medial fibers of the nervus terminalis - Nt nervus terminalis - Npo nucleus praeopticus     

Substance P-, F8Famide-, and A18Famide-like immunoreactivity in the nervus terminalis and retina of the goldfish Carassius auratus

We re-investigated the occurrence of substance P-like immunoreactivity in the retina of the goldfish Carassius auratus using antisera to substance P and other tachykinins. Most antisera labelled a previously described single class of mono-stratified amacrine cells arborizing in layer 3 of the inner plexiform layer. Preabsorption experiments showed that these amacrine cells contained at least one tachykinin-like peptide. One antiserum (INC 353) to substance P labelled not only these amacrine cells but also fibres in layer 1 of the inner plexiform layer and fibres in the optic nerve. These fibres were identified as retinopetal projections of the nervus terminalis, in part because of colocalized labelling with antisera against gonadotropin-releasing hormone and FMRFamide. Preabsorption experiments showed that the substance P-immunoreactive material in the nervus terminalis was not substance P or any other typical tachykinin. Labelling of the nervus terminalis with INC 353 was blocked by preabsorption with two bovine FMRFamide-like peptides, F8Famide and A18Famide, which contain a substance P(4–7)-like region. Antisera to F8Famide and A18Famide strongly labelled ganglia of the nervus terminalis and retinopetal fibres. We suggest that labelling of the nervus terminalis by antisera to substance P and FMRFamide occurs because of homologies between these antigens and a non-tachykinin, endogenous peptide that is similar to F8Famide and A18Famide.     

Peripheral projections of nervus terminalis LHRH-containing neurons in the tiger salamander,Ambystoma tigrinum

The peripheral projections of the nervus terminalis (NT) have been difficult to examine due to the weak immunoreactivity of the processes to various antibodies. We performed two experimental manipulations in the tiger salamander in an attempt to increase the luteinizing hormone-releasing hormone-immunoreactive (LHRH-ir) labelling in the peripheral processes of the NT: 1) the NT was sectioned centrally, or 2) a 100 mg melatonin pellet was embedded subcutaneously for 3 days prior to sacriffice. Following these manipulations, animals were sacrifficed and tissue was processed with standard immunocytochemical techniques for the analysis of the distribution of LHRH-ir processes. In the nasal cavity, LHRH-ir fibers were observed projecting 1) into the rostral olfactory epithelium, 2) to Bowman's glands in the lamina propria of the rostromedial olfactory mucosa and ventrolateral mucosa between the main nasal cavity and Jacobson's organ, 3) into the naris constrictor muscle, and 4) along the palatine nerves and ganglia. These lesion and hormone manipulations have enabled the detection of peripheral projections of the NT not observed previously with immunocytochemical procedures alone. The wide distribution of LHRH-ir NT processes in the nasal cavity and cranium suggests that this nerve may influence many different cranial structures during appropriate pheromonal or neuroendocrine events.     

Connections of the olfactory bulb in the piranha (Serrasalmus nattereri)

Summary The connections of the olfactory bulb were studied in the piranha using the Nauta and horseradish-peroxidase methods. Three olfactory tracts project to seven terminal fields in the telencephalon and one in the diencephalon, all of them bilaterally. The contralateral olfactory bulb also receives a small input. All contralateral projections decussate in the anterior commissure and are relatively weak compared to the ipsilateral projections. HRP-containing cells were found in all of the ipsilateral telencephalic aggregates receiving an olfactory tract projection; the contralateral side was free of labeled cell bodies. Although only about one fourth of the entire telencephalon receives a direct olfactory input, the high degree of differentiation of the olfactory system suggests that the piranha depends substantially on the sense of olfaction and that this species may be a good model for further studies on olfactory mechanisms.     

Central connections of the olfactory bulb in the bichir,Polypterus palmas,reexamined

Summary Central connections of the olfactory bulb of Polypterus palmas were studied with the use of horseradish peroxidase and cobalt-tracing techniques. The olfactory bulb projects to subpallial and palliai areas in the ipsilateral telencephalon; a projection to the contralateral subpallium is noted via the habenular commissure. A further target of secondary olfactory fibers is a caudal olfactory projection area in the ipsilateral hypothalamus. No labeling was seen in the anterior commissure and in the contralateral olfactory bulb. The medial and the lateral pallium receive secondary olfactory fibers in distinct areas. Neurons projecting to the bulb are found in the ipsilateral subpallium, mainly in one dorsal longitudinal nucleus. The main connection with the tel- and diencephalon is mediated via the medial olfactory tract. This tract also contains fibers to the contralateral telencephalon, and to the hypothalamus. The smaller lateral olfactory tract mediates fibers to the lateral pallium. The organization of pathways of secondary olfactory fibers in the telencephalon is described. The present findings are compared to those obtained in species possessing an inverted forebrain.This investigation was supported by grants from the Deutsche Forschungsgemeinschaft to DLM     

Distribution of FMRFamide-like immunoreactivity in the brain,retina and nervus terminalis of the sockeye salmon parr,Oncorhynchus nerka

Summary Neurons displaying FMRFamide(Phe-Met-Arg-Phe-NH₂)-like immunoreactivity have recently been implicated in neural plasticity in salmon. We now extend these findings by describing the extent of the FMRF-like immunoreactive (FMRF-IR) system in the brain, retina and olfactory system of sockeye salmon parr using the indirect peroxidase anti-peroxidase technique. FMRF-IR perikarya were found in the periventricular hypothalamus, mesencephalic laminar nucleus, nucleus nervi terminalis and retina (presumed amacrine cells), and along the olfactory nerves. FMRF-IR fibers were distributed throughout the brain with highest densities in the ventral area of the telencephalon, in the medial forebrain bundle, and at the borders between layers III/IV and IV/V in the optic tectum. High densities of immunoreactive fibers were also observed in the area around the torus semicircularis, in the medial hypothalamus, median raphe, ventromedial tegmentum, and central gray. In the retina, immunopositive fibers were localized to the inner plexiform layer, but several fiber elements were also found in the outer plexiform layer. The olfactory system displayed FMRF-IR fibers in the epithelium and along the olfactory nerves. These findings differ from those reported in other species as follows: (i) FMRF-IR cells in the retina have not previously been reported in teleosts; (ii) the presence of FMRF-IR fibers in the outer plexiform layer of the retina is a new finding for any species; (iii) the occurrence of immunopositive cells in the mesencephalic laminar nucleus has to our knowledge not been demonstrated previously.     

Pheromone detection and olfactory pathways in the brain of the female African catfish,Clarias gariepinus

Summary The olfactory tract of the African catfish, Clarias gariepinus, consists of two tracts, the medial and lateral olfactory tract. Ovulated female catfish are attracted by male steroidal pheromones. Attraction tests with catfish in which the medial and lateral olfactory tract have been selectively lesioned show that the effects of these pheromones are mediated by the medial olfactory tract. The central connections of the medial and lateral olfactory tract have been studied by retro- and anterograde transport techniques using horseradish peroxidase as a tracer. Upon entering the forebrain, the medial olfactory tract innervates the posterior pars ventralis and pars supracommissuralis of the area ventralis telencephali and the nucleus preopticus periventricularis, the nucleus preopticus and the nucleus recessus posterioris. Application of horseradish peroxidase to the olfactory epithelium shows that part of the innervation of the area ventralis telencephali and the nucleus preopticus periventricularis can be attributed to the nervus terminalis, which appears to be embedded in the medial olfactory tract. The lateral olfactory tract sends projections to the same brain areas but also innervates the nucleus habenularis and a large terminal field in the area dorsalis telencephali pars lateralis ventralis. Furthermore, the medial olfactory tract carries numerous axons from groups of perikarya localized in the area dorsalis telencephali. Contralateral connections have been observed in the olfactory bulb, telencephalon, diencephalon and mesencephalon. It is suggested that processes of the medial olfactory tract innervating the preoptic region may influence the gonadotropin-releasing hormone system and in doing so may lead to behavioral and physiological changes related to spawning.     

Electron microscopic study of the termination of the centrifugal fibers in the goldfish olfactory bulb

Summary The terminals of centrifugal fibers to the olfactory bulbs of goldfish were studied by electron microscopy after transection of the medial, lateral or entire olfactory tract. The centrifugal fibers originate in the telencephalic hemisphere, pass through both the medial and the lateral olfactory tract, and form synaptic contacts with dendrites in the granule cell layer.     

A novel population of neuronal cells expressing the olfactory marker protein (OMP) in the anterior/dorsal region of the nasal cavity

The olfactory marker protein (OMP) is expressed in mature chemosensory neurons in the nasal neuroepithelium. Here, we report the identification of a novel population of OMP-expressing neurons located bilaterally in the anterior/dorsal region of each nasal cavity at the septum. These cells are clearly separated from the regio olfactoria, harboring the olfactory sensory neurons. During mouse development, the arrangement of the anterior OMP-cells undergoes considerable change. They appear at about stage E13 and are localized in the nasal epithelium during early stages; by epithelial budding, ganglion-shaped clusters are formed in the mesenchyme during the perinatal phase, and a filiform layer directly underneath the nasal epithelium is established in adults. The anterior OMP-cells extend long axonal processes which form bundles and project towards the brain. The data suggest that the newly discovered group of OMP-cells in the anterior region of the nasal cavity may serve a distinct sensory function.     

Ultrastructural identification of catecholaminergic fibers in the goldfish pituitary

Summary The monoaminergic innervation of the goldfish pituitary gland was studied by means of light- and electronmicroscopic radioautography after in vitro administration of ³H-dopamine. The tracer was specifically incorporated and retained by part of the type-B fibers innervating the different lobes of the pituitary. In the rostral pars distalis labeled fibers were most frequently observed in contact with the basement membrane separating the neurohypophysis and the adenohypophysis. In the proximal pars distalis and the pars intermedia, labeled profiles were detected in the neural tissue and in direct contact with the different types of secretory cells.According to the previous data concerning the uptake and retention of tritiated catecholamines in the central nervous system, it is assumed that the labeled fibers are mainly catecholaminergic (principally dopaminergic). This study provides morphological evidence for a neuroendocrine function of catecholamines in the goldfish.     

An interplexiform cell in the goldfish retina: light-evked response pattern and intracellular staining with horseradish peroxidase

Summary The light-evoked response pattern and morphology of one interplexiform cell were studied in the goldfish retina by intracellular recording and staining. The membrane potential of the cell spontaneously oscillated in the dark. In response to a brief light stimulus, the membrane potential initially gave a slow transient depolarization. During maintained light, the oscillations showed a tendency to be suppressed; the response of the cell to the offset of the stimulus was not so prominent. The perikaryon of the interplexiform cell was positioned at the proximal boundary of the inner nuclear layer. The cell had two broad layers of dendrites; one was diffuse in the inner plexiform layer, the other was more sparse in the outer plexiform layer. The morphological and electrophysiological characteristics of the cell are discussed in relation to dopaminergic interplexiform cells and the light-evoked release pattern of dopamine in the teleost retina.     

Immunocytochemical distribution of FMRFamide-like substance in the brain of the cloudy dogfish,Scyliorhinus torazame

Summary The distribution of the molluscan cardioexcitatory tetrapeptide FMRFamide (Phe-Met-Arg-Phe-NH₂) in the brain of the cloudy dogfish, Scyliorhinus torazame, was examined by immunocytochemistry. FMRFamide-like immunoreactivity was demonstrated to occur extensively in various regions of the dogfish brain, except for the corpus cerebelli. Immunoreactive neuronal perikarya were located in the ganglion of the nervus terminalis, the preoptic area, and the hypothalamic periventricular gray matter consisting of the nucleus medius hypothalamicus, the nucleus lateralis tuberis, and the nucleus lobi lateralis. some of the immunoreactive cells in the hypothalamus were identified as cerebrospinal fluid-contacting neurons. The bulk of the immunostained fibers in the nervus terminalis penetrated into the midventral portion of the telencephalon and ran dorsocaudally toward the basal telencephalon and hypothalamus, showing radial projections or ramifications. The labeled fibers were abundant in the midbasal part of the telencephalon and in the hypothalamus, where some fibers were found in loose networks around the cell bodies of the nucleus septi and hypothalamic periventricular nuclei. The fibers demonstrated in the hypothalamus terminated around the vascular wall of the primary capillary plexus of the median eminence or penetrated deeply into the pars intermedia of the hypophysis. These results suggest that, in the dogfish, an FMRFamide-like substance participates in the regulation of adenohypophysial function. This molecule may have a role as a neurotransmitter and/or neuromodulator in the central nervous system.     

Distribution of neuropeptide Y-like immunoreactivity in the brain and hypophysis of the cloudy dogfish,Scyliorhinus torazame

Summary Using a specific antiserum raised against synthetic neuropeptide Y, we examined the localization of immunoreactivity in the brain and hypophysis of the cloudy dogfish, Scyliorhinus torazame, by the peroxidase-antiperoxidase method. Immunoreactive perikarya were demonstrated in the ganglion of the nervus terminalis, the dorsocaudal portions of the pallium dorsale, the basal telencephalon, and the nucleus lateralis tuberis and the nucleus lobi lateralis in the hypothalamus. Labeled perikarya were also found in the tegmentum mesencephali, the corpus cerebelli, and the medulla oblongata. Some of the immunoreactive neurons in the hypothalamus were of the CSF-contacting type. The bulk of the labeled fibers in the nervus terminalis ran toward the basal telencephalon, showing radial projections and ramifications. Large numbers of these fibers coursed into the nucleus septi caudoventralis and the nucleus interstitialis commissurae anterioris, where they became varicose and occasionally formed fine networks or invested immunonegative perikarya. In the diencephalon, immunoreactive fibers were observed throughout the hypothalamus, e.g., in the pars neurointermedia of the hypophysis, the subependymal layer of the lobus inferior hypothalami, and in the neuropil of the posterior (mammillary) recess organ. Labeled fibers were scattered throughout the rest of the brain stem and were also seen in the granular layer of the cerebellum. These results suggest that, in the dogfish brain, neuropeptide Y or a related substance is involved in a variety of physiological processes in the brain, including the neuroendocrine control of the hypophysis.     

Origin of the pituitary innervation in the goldfish

Despite the large number of studies devoted to the pituitary of teleosts, the origin of the direct pituitary innervation is still largely unknown. Although such a model is ideal for applying retrograde transport techniques, these methods involve the difficult in vivo injection of tracers into the pituitary and have never been applied. Recently, a lipophilic fluorescent dye (1-1-dioctadecyl-3,3,3,3-tetramethylindocarbocyanin; DiI) has been introduced and shown to have the capacity of being transported by the membranes of paraformaldehydefixed tissues. Microcrystals of DiI were implanted via a ventral approach into the pituitary of goldfish previously fixed by intracardiac perfusion of paraformaldehyde. The goldfish heads were kept immersed in paraformaldehyde for various periods of time (2–6 weeks). The brains were then dissected and cut transversally using a Vibratome. The results demonstrate that hypophysiotrophic areas are essentially restricted to the preoptic region, the mediobasal hypothalamus and the nucleus dorsolateralis thalami. In addition, cell bodies probably containing gonadotrophin releasing-hormone were also retrogradely stained along a pathway that can be traced up to the olfactory bulbs. The results also confirm that cell bodies, located around the ventral aspect of the preoptic recess and probably corresponding to dopaminergic neurons, project to the pituitary. Large neurons have also been observed in the rostral dorsal midbrain tegmentum just caudal to the posterior commissure. Most neurons of the so-called paraventricular organ remain unstained. Finally, a fiber tract originating from an undetermined territory of the posterior brain has been observed. The results are discussed in relation to the possible chemical nature of the hypophysiotropic neurons.     

Feeding-induced changes of melanin-concentrating hormone (MCH)-like immunoreactivity in goldfish brain

Intracerebroventricular (ICV) injection of melanin-concentrating hormone (MCH) influences feeding behavior in the goldfish and exerts an anorexigenic action in goldfish brain, unlike its orexigenic action in mammals. Despite a growing body of knowledge concerning MCH function in mammals, the role of MCH in appetite has not yet been well studied in fish. The aim of the present study was to investigate the involvement of endogenous MCH in the feeding behavior of the goldfish. We examined the distribution of MCH-like immunoreactivity (MCH-LI) in the goldfish brain and the effect of feeding status upon this distribution. Neuronal cell bodies containing MCH-LI were localized specifically to four areas of the hypothalamus. Nerve fibers with MCH-LI were found mainly in the neurohypophysis, with a few in the telencephalon, mesencephalon, and diencephalon. The number of neuronal cell bodies containing MCH-LI in the dorsal area adjoining the lateral recess of the third ventricle in the posterior and inferior lobes of the hypothalamus showed a significant decrease in fasted fish compared with that in normally fed fish, although other areas showed no evident differences. We also administered an antiserum against fish MCH (anti-MCH serum) by ICV injection and examined its immunoneutralizing effect on food intake by using an automatic monitoring system. Cumulative food intake was significantly increased by ICV injection of the anti-MCH serum. These results indicate that MCH potentially functions as an anorexigenic neuropeptide in the goldfish brain, and that the further study of the evolutionary background of the MCH system and its role in appetite is warranted. This work was supported by Grants-in-Aid from the Ministry of Education, Culture, Sports, Science, and Technology of Japan (K.M. and A.T.) and by a research grant from the Toyama Marine Biotechnology Association (K.M.).