Melanin-concentrating hormone-producing neurons in birds.

The peptidergic melanin-concentrating hormone (MCH) system was investigated by immunocytochemistry in several birds. MCH perikarya were found in the periventricular hypothalamic nucleus near the paraventricular organ and in the lateral hypothalamic areas. Immunoreactive fibers were very abundant in the ventral pallidum, in the nucleus of the stria terminalis, and in the septum/diagonal band complex, where immunoreactive pericellular nets were prominent. Many fibers innervated the whole preoptic area, the lateral hypothalamic area, and the infundibular region. Some fibers also reached the dorsal thalamus and the epithalamus. The median eminence contained only sparse projections, and the posterior pituitary was not labeled. Thus, in birds, a neurohormonal role for MCH is not likely. Immunoreactive fibers were observed in other regions, such as the intercollicular nucleus, stratum griseum periventriculare (mesencephalic tectum), central gray, nigral complex (especially the ventral tegmental area), reticular areas, and raphe nuclei. Although no physiological investigation concerning the role of MCH has been performed in birds, the distribution patterns of the immunoreactive perikarya and fibers observed suggest that MCH may be involved in functions similar to those described in rats. In particular, the projections to parts of the limbic system (ventropallidal ganglia, septal complex, hypothalamus, dorsal thalamus, and epithalamus) and to structures concerned with visceral and other sensory information integration suggest that MCH acts as a neuromodulator involved in a wide variety of physiological and behavioral adaptations (arousal) with regard to feeding, drinking, and reproduction.

[Post-embryonic differentiation of a cutaneous electro-receptor]. / Sur la différenciation postembryonnaire d'un électrorécepteur cutané.

In order to decrease the rate of postembryonic development of electroreceptor organs, excisions of epidermis and deafferentations were carried out in the gymnotid fish Eigenmannia virescens. Twenty-five days later, the epidermis showed electroreceptor organs without innervation. Some of these at the beginning of their development consisted of masses of identical cells, whereas others showed presumed sensory cells whose cytoplasm contained rudimentary synaptic structures. The epidermis also showed differentiated tuberous organs with a low number of sensory cells. In all these organs, radioactive thymidine was fixed in the nuclei of the platform accessory cells. Thirty-five-40 days after surgery, tuberous organs were identical to the functional organs, and thymidine was detected in the nuclei of the cavity accessory cells. These results show that the gymnotid electroreceptor organs can develop before any nervous contact occurs, and suggest that they might originate from epidermal cells.

Vasoactive intestinal polypeptide (VIP)-containing neurons: distribution throughout the brain of the chick (Gallus domesticus) with focus upon the lateral septal organ.

The distribution of VIP-like perikarya and fibers was determined throughout the chick brain. The most rostral immunoreactive perikarya were found to be cerebrospinal fluid-contacting neurons in the pars medialis of the lateral septal organ. Additional data were presented supporting the idea that the lateral septal organ is another circumventricular organ within the brain of birds (Kuenzel and van Tienhoven 1982). A large group of immunoreactive perikarya was found in the lateral hypothalamic area and appeared continuous with immunoreactive neurons in the anterior medial and ventromedial hypothalamic nuclei (n). A few perikarya were located in the paraventricular hypothalamic n. A number of immunoreactive neurons were found within and about the infundibular and inferior hypothalamic n., none however was immunoreactive cerebrospinal fluid-contacting neurons. Immunoreactive perikarya were found predominantly in laminae 10-11 of the stratum griseum et fibrosum superficiale. A few scattered perikarya were found ventromedial to the n. tegmenti pedunculo-pontinus pars compacta and locus ceruleus. Some of the immunoreactivity was unusual, being very homogeneous within the cell body with little evidence of the material in the axon or dendrites. Perikarya were found in the central gray, n. intercollicularis, and area ventralis of Tsai. The most caudal structure showing immunoreactive neurons was the n. reticularis paragigantocellularis lateralis. Brain areas containing the most abundant immunoreactive fibers, listed from the rostral-most location, were found in the ventromedial region of the lobus parolfactorius and the lateral septal n. Continuing caudally, there were immunoreactive fibers within the periventricular hypothalamic n.; some of the fibers were found to travel for some distance parallel to the third ventricle. Dense immunoreactive fibers were found in the tractus cortico-habenularis et cortico-septalis, medial habenular n. and posterior and dorsal n. of the archistriatum. A number of areas had what appeared to be baskets of immunoreactive fibers (perhaps immunoreactive terminals) surrounding non-reactive perikarya. Brain areas containing terminals included the piriform cortex, area ventralis of Tsai, interpeduncular n., and specific regions of the stratum griseum et fibrosum superficiale. A very dense immunoreactivity occurred within the external zone of the median eminence, the dorsolateral parabrachial n., and n. tractus solitarii. Vasoactive intestinal polypeptide appears to be a useful peptide for defining the neuroanatomical constituents of the visceral forebrain in birds.

[Development, during ontogenetic development of gymnotids, of substance p expression in tuberous organs (electroreceptors)]. / Evolution, durant le développement ontogénétique des gymnotides, de l'expression de la substance P dans les organes tubéreux (électrorécepteurs).

The evolution of the neuropeptidic expression of Substance P has been investigated with immunohistochemistry in the cutaneous electroreceptors, tuberous organs, during ontogenetic development of Apteronotus leptorhynchus. In the present data, antiSP antiserum has been applied to serial sections of Apteronotus leptorhynchus larvae obtained from several egg layings. Larvae were taken during development from hatching up to one hundred days old. SP immunoreactivity appeared just after hatching, in the epidermal zones which give rise to cutaneous sense organs. Four days after hatching, the tuberous organs are differentiated and immunoreactivity was observed in these organs, in both sensory cells and accessory cells. From day 30 after hatching, there was a regular decrease in the number of tuberous organs showing labelled accessory cells, and one hundred days later only 8% of tuberous organs had immunoreactive accessory cells. The adult accessory cells were no longer labelled with anti-SP antiserum. The results showed that in Apteronotus leptorhynchus, the epidermal structures which give rise to the cutaneous sensory organs were immunoreactive at a very early stage of development; this suggests that SP could have an effect upon their differentiation.

The distribution of gonadotropin-releasing hormone (GnRH) neurons and fibers throughout the chick brain (Gallus domesticus).

Nerve fibers and perikarya containing gonadotropin-releasing hormone (GnRH-like) immunoreactivity were investigated in the brain of the three-week-old chick. Gallus domesticus using the technique of immunocytochemistry. Six major groups of perikarya were found to include the olfactory bulb, olfactory tubercle/lobus parolfactorius, nucleus accumbens, septal preoptic hypothalamic region (three sub-nuclei), lateral anterior thalamic nucleus and in and about the oculomotor complex. The immunostaining was unusual in the latter group, suggesting that the neurons may contain a GnRH-II like material. Immunoreactive fibers for GnRH were found throughout the entire brain extending from the olfactory bulbs to the caudal brainstem. Two anatomical areas, not emphasized in the past literature, which had distinct GnRH-like immunoreactivity, included the lateral anterior thalamic nucleus and the preoptic recess. The former included a group of GnRH perikarya that is also known to be a retino-recipient area while the latter contained neuronal terminals some of which appeared to be contacting the cerebrospinal fluid of the preoptic recess. An attempt was made to list all anatomical structures that contained or were juxta-positioned to sites that displayed immunoreactive perikarya and fibers including circumventricular organs.

The olfactoretinalis system = terminal nerve?

Immunohistochemistry revealed a distinct terminal nerve (nT) from the olfactoretinalis system (ORS) in several gymnotid species. The ganglion (NOR) of the ORS is a cluster of FMRF-amide immunoreactive neurones located on the olfactory nerve and bulb. The NOR projects to the olfactory bulb and optic nerve but not to the olfactory epithelium. In contrast, the nT is a small substance P or GnRH immunoreactive fibre bundle which originates from a ganglion located rostral to the olfactory epithelium, courses caudally with the olfactory nerve and terminates as glomeruli in the olfactory bulb. Peripherally the ganglion projects to the epithelium of the anterior nostril. Thus, the terminal nerve appears to be a cranial nerve, clearly distinct from the ORS which consequently should no longer be considered as the terminal nerve.

[Extra-bulbar primary olfactory projection in teleost fishes]. / Projection olfactive primaire extrabulbaire chez certains poissons téléostéens.

Immunohistochemical investigations with anti-substance P antiserum demonstrate the existence of an extensive extrabulbar primary olfactory projection in several gymnotid teleost fish. This projection, never described before, originates in particular primary olfactory bundles which enter with the olfactory nerve into the olfactory bulb. While the bulk of the olfactory fibers end with glomeruli in the glomerular layer of the olfactory bulb, two particular bundles penetrate into the telencephalon and end, without forming glomeruli, in several telencephalic and diencephalic regions. A few fibers run as far as to the hypothalamus. In the light of these findings, the general notion that the primary olfactory projection is limited to the olfactory bulb and forms only glomeruli-like terminals, should be reconsidered.

Testing of Arg-8-gonadotropin-releasing hormone-directed antisera by immunological and immunocytochemical methods for use in comparative studies.

Three polyclonal antisera raised in rabbits against the mammalian molecular form of gonadotropin-releasing hormone (GnRH) were tested in enzyme-linked immunosorbent assays for crossreactivity with naturally occurring GnRHs and with GnRH analogues. Antisera were then tested immunocytochemically in order (i) to identify amino acids essential for the binding of each antiserum, and (ii) to evaluate the specificity of the immunocytochemical reaction in brain sections from various species of cyclostomes, amphibians, reptiles, and birds. Antiserum GnRH 80/1, recognizing mainly a discontinuous determinant including the NH2- and COOH-termini, crossreacts with GnRHs the molecular bending of which enables the spatial approach of both terminal amino acid residues. Antiserum GnRH 80/2, by requiring the COOH-terminus for binding and not tolerating substitutions by aromatic amino acids in the middle region of the molecule, recognizes chicken I GnRH, however, not the salmon form. The use of this antiserum is appropriate in species synthesizing the mammalian and/or the chicken I form of GnRH. GnRH antiserum 81/1 is specific mostly for mammalian GnRH. The results obtained by ELISAs are confirmed by immunocytochemical studies. A comparison between the results obtained in ELISA and in immunocytochemistry involving mammalian-, chicken I-, chicken II-, salmon-, and lamprey-directed GnRH antisera resulted in the following conclusions: (1) An antiserum recognizing the discontinuous antigen determinant including both NH2- and COOH-termini may be reactive in most vertebrate brain sections thus being appropriate for phylogenetically directed immunocytochemical studies. (2) Moreover, this discontinuous determinant seems to be immunocytochemically reactive in all parts of the neurons in the GnRH system, whereas, in some species, determinants located in the middle region of the molecule(s) tend to become reactive only during the axonal transport. (3) A crossreaction between tissue-bound antigen and antibodies recognizing the above cited discontinuous determinant indicates an appropriate bending of the molecule even in case of severe molecular differences, e.g., in lamprey form of GnRH. (4) It follows that in phylogenetic studies, an immunologically well characterized antiserum can be substituted for a species-directed antiserum.

Changes in body temperature and vasopressin content of brain neurons, in pregnant and non-pregnant guinea pigs, during fevers produced by Poly I:Poly C.

The synthetic polyribonucleotide pyrogen Poly I:Poly C (800 micrograms.kg-1) was injected intramuscularly on alternate days into pregnant and non-pregnant female guinea pigs. Pregnant animals, close to term, had smaller fevers in response to the pyrogen than did non-pregnant animals. Repeated injections of the pyrogen caused sequentially smaller fevers for the first 3-4 injections, particularly in non-pregnant animals, and this appeared to be like the tolerance usually developed to repeated injections of endotoxin. Continued pyrogen injections then caused, in non-pregnant animals, fevers of increasing magnitude until the original fever levels were reached, whereas in pregnant guinea pigs the fever responses remained reduced until parturition. The development of tolerance was associated with an increase in immunoreactivity for arginine vasopressin (AVP) in some neurons in the medial part of the paraventricular nucleus, and in terminals in the lateral septum and amygdala similar to changes found in these areas at term of pregnancy. These observations raise the possibility that AVP in these regions may have a role in the development of tolerance to pyrogens, and further quantitative studies of the AVP content of, and release from, nerve terminals projecting to the limbic system seem warranted.

The ACTH-immunoreactive system in the brain of the white-crowned sparrow, Zonotrichia leucophrys gambelii (Passeriformes, Emberizidae).

The ACTH-immunoreactive (ir) system of the avian brain is particularly conspicuous in the male white-crowned sparrow (Zonotrichia leucophrys gambelii). The irperikaryal population is concentrated mainly within the tuberal region, projecting primarily in a dorsal direction: (i) into the striatum; (ii) into rostral diencephalic, septal, hyperstriatal, and thalamic areas; and (iii) into dorsal and ventral areas of the brain stem. Ir-fibers seemingly contact local non-immunoreactive neurons mainly in the accumbens nucleus, septum, dorsal thalamic nuclei, infundibular and interpeduncular nuclei, and in the rostral diencephalon. Neurohemal zones are not supplied by ACTH-ir terminals. Immunocytochemical problems arising from the complexity of the proopiomelanocortin molecule and its derived peptide components are discussed in relation to phylogenetically directed studies, and contradictory results.

High substance P-like immunoreactivity (SPLI) in the olfactory and electrosensory cells of gymnotid fish.

Substance P has been proposed as a candidate neurotransmitter or neuromodulator in the nociceptive system. Using a light microscopial immunohistochemical peroxidase-anti-peroxidase technique we have detected high substance P-like immunoreactivity (SPLI) in several types of sensory organs of 4 species of gymnotiform teleost fish: olfactory epithelium, vestibular, lateral line and electrosensory organs. The olfactory nerve and its endings within the olfactory bulb and the telencephalon were also strongly labelled. At these sites no SPLI was revealed in other teleosts (Carassius auratus, Gnathonemus petersii). The findings suggest that substance P may be involved in neurotransmission or neuromodulation in these specific sensory systems of these species.

Somatostatin cells in the gastric mucosa of small ruminants. Immunohistochemical study.

The distribution of somatostatin cells was investigated immunohistochemically in the epithelium of the abomasal mucous membrane of sheep and goats. Tissue samples were taken from different areas at the lesser and greater curvature of the glandular stomach. More somatostatin cells per 0.5 mm2 were found in sheep than in goats, predominantly in the pyloric gland region. In the pyloric area of both species the mucosa in the greater curvature contained more immunoreactive cells than the one in the lesser curvature. A few somatostatin cells with basal cytoplasmic processes of the 'open type' were observed over the entire abomasal mucosa.

Gonadotropin-releasing hormone-immunoreactive neuronal structures in the brain and pituitary of the African catfish, Clarias gariepinus (Burchell).

The distribution of gonadotropin-releasing hormone (GnRH) immunoreactivity was studied in the African catfish, Clarias gariepinus, by means of immunofluorescence and immunoperoxidase techniques. Immunoreactive neurons were found throughout the preoptic nucleus (NPO). However, only a portion of the secretory perikarya in the NPO showed a positive reaction by use of an anti-LHRH serum. Numerous immunoreactive fibres were found to enter the pituitary and to terminate in its proximal pars distalis, the site of concentration of the gonadotropic cells. Since GnRH is present in the brain and pituitary of the African catfish, the lack of spontaneous ovulation in captivity is apparently due to an insufficient release of GnRH.

Peptidergic pathways in the avian brain.

The results obtained by topographical studies on the immunoreactive peptide systems in the embryonic and adult avian brain (domestic fowl, domestic mallard, pigeon, Japanese quail, and zebra finch) can be realized only by means of phylogenetical comparisons. The comparative studies mainly demonstrate a fascinating constancy of the immunological properties and the spatial distribution of the neuropeptides. Independent of the development of the neopallium, and the increasing cerebral complexity, the spatial distribution of the neuropeptides, the location of their main perikaryal accumulations which are interconnected by immunoreactive fiber projections (and thereby forming widespread but continuous peptide systems) remain nearly unchanged during vertebrate evolution. The recognition of the neuropeptides as integral parts of the central nervous system is demonstrated by the fact that neuropeptidergic structures connect sensory inputs with central nervous areas as well as with the peripheral endocrinium.

Cerebrospinal fluid-contacting neurons and other somatostatin-immunoreactive perikarya in brains of tadpoles of Xenopus laevis.

In untreated tadpoles of Xenopus laevis, stage 60 (Nieuwkoop and Faber 1956), somatostatin-immunoreactive perikarya and fiber networks are already discernible in the pallium and the septum, as well as in the anterior and posterior hypothalamus. In addition, somatostatin-immunoreactive cerebrospinal fluid (CSF)-contacting neurons were found in the periventricular gray of the most caudal division of the ventral tuber cinereum. The results are discussed with respect to the inhibitory influence of the system of CSF-contacting and other somatostatin-immunoreactive neurons on the anterior pituitary and other target sites, especially during the climax of metamorphosis.

Fever response in the guinea pig before and after parturition.

The febrile response to an intramuscular injection of bacterial endotoxin (E. coli 4 microgram/kg) was tested in guinea pigs at the end of pregnancy in the time period extending from 8 days before until 3 days after parturition. In comparison to non-pregnant female controls both fever height and fever index were reduced in mother guinea pigs one week before parturition. This response was gradually reduced and reached its minimum on the last day before parturition. Immediately after parturition the fever response was still suppressed in mother animals as well as in newborns. Several hours after birth the fever response increased again in both groups of animals. The onset time and duration of fever were, however, shorter than in controls. The full fever responsivity was not reached until several days postpartum. Apparently the guinea pig develops an active antipyresis during the last phase of pregnancy. This resembles the suppression of fever in ewes at term of pregnancy where endogenous arginine-vasopressin has been proposed as an antipyretic agent. The vasopressinergic neuronal systems have therefore been localized by immunohistochemical methods in the brains of the guinea pigs whose responses to bacterial endotoxin were studied. These studies, which are described in detail in a following paper, support the involvement of vasopressin in natural antipyresis in the guinea pig.

Reactivity pattern of vasopressin-containing neurons and its relation to the antipyretic reaction in the pregnant guinea pig.

Changes in the content of vasopressin-immunoreactive material in neurons and their projections were examined in pregnant and nonpregnant guinea pigs as well as in mother and newborn animals. Before sacrifice all animals used in the present study were submitted to a pyrogen test, during which the pregnant animals displayed a reduced fever response to exogenous pyrogen. The unlabeled enzyme-immunoperoxidase method was used in the present study. Light microscopic examination showed that, in comparison to all other groups examined, the pregnant animals exhibited a reduced content of the vasopressin-immunoreactive substance in the supraoptic nucleus (SON), in the neuronal pathways extending between the paraventricular nucleus (PVN) and the SON, as well as in the axons projecting to the neural lobe of the pituitary. An increased amount of vasopressin-immunoreactive material was observed during pregnancy especially in the medial portion of the PVN, in axonal distensions in the external zone of the median eminence and in the extrahypothalamic projection sites of the PVN in the lateral septum and in the amygdala. In the pregnant animals neurovascular contacts of vasopressinergic perikarya and fibers were abundant in the PVN; in the lateral septum and in the amygdala vasopressinergic terminals appeared to contact neurons of other types. It is suggested from the present immunocytochemical results that activation of neurons in the medial portion of the PVN and the increased number of vasopressinergic terminals and preterminals in the lateral septum and in the amygdala might be functionally involved in fever suppression at the term of pregnancy.

The cellular responses in Marisa cornaurietis experimentally infected with Angiostrongylus cantonensis.

The cellular responses in the poorly susceptible ampullariid snail Marisa cornaurietis to Angiostrongylus cantonensis, during a period of 40 days, involved focal and generalized proliferative reactions. The focal reactions appeared around all larvae as accumulations of variable numbers of amoebocytes at 24 h after infection in the loose connective tissues and somewhat later in the dense tissues. This cellular infiltration intensified gradually with time leading to the encapsulation of the parasite. At 18 days after infection a central zone formed mainly of hypertrophic, granular, highly acidophilic amoebocytes and a peripheral zone of fibrous appearance were conspicuous in most capsules. The generalized proliferative reaction appeared at about day 4 after infection as leucocytosis in the vascular system of the snail and in the connective tissue. At 25 days after infection many leucocytes which are markedly hypertrophic, highly eosinophilic, and densely granulated were found in the vascular system. These cells were spread in the connective tissues and contributed to the formation of the capsules at 40 days after infection. The sources of amoebocytes appear to be mainly several hyperactive areas in the loose connective tissue and in the lung, some active foci in the body wall at the head region, and also in an amoebocyte-producing organ which lies in the roof of the lung.