Cortical cholinergic decline parallels the progression of Borna virus encephalitis.

Borna disease virus (BDV)-induced meningoencephalitis is associated with the dysfunction of the cholinergic system. Temporal development of this cholinergic decline during pre-encephalitic and encephalitic stages of BDV infection remains however elusive. Changes in choline acetyltransferase (ChAT) and acetylcholinesterase (AChE) activities were therefore determined in the cerebral cortex, hippocampus, striatum, amygdala and cholinergic basal forebrain nuclei (ChBFN) of rats infected with BDV. Immunocytochemistry for ChAT and vesicular acetylcholine transporter (VAChT) was employed to identify morphological consequences of BDV infection on cholinergic neurons. Whereas both ChAT and AChE activities changed only slightly under pre-encephalitic conditions, the encephalitic stage was characterized by a significant decrease of ChAT activity in the cerebral cortex, horizontal diagonal band of Broca (hDBB), hippocampus and amygdala concomitant with a marked reduction of AChE activity in the cerebral cortex, hDBB and hippocampus. The striatum and medial septum remained unaffected. ChAT and VAChT immunocytochemistry revealed prominent axonal degeneration in affected cortical and limbic projection areas of ChBFN. In summary, our data indicate progressive deterioration of forebrain cholinergic systems that parallels the progression of BDV encephalitis.

Beta-amyloid(1-42)-induced cholinergic lesions in rat nucleus basalis bidirectionally modulate serotonergic innervation of the basal forebrain and cerebral cortex.

Ample experimental evidence suggests that beta-amyloid (A beta), when injected into the rat magnocellular nucleus basalis (MBN), impels excitotoxic injury of cholinergic projection neurons. Whereas learning and memory dysfunction is a hallmark of A beta-induced cholinergic deficits, anxiety, or hypoactivity under novel conditions cannot be attributed to the loss of cholinergic MBN neurons. As mood-related behavioral parameters are primarily influenced by the central serotonergic system, in the present study we investigated whether A beta(1-42) toxicity in the rat MBN leads to an altered serotonergic innervation pattern in the rat basal forebrain and cerebral cortex 7 days postsurgery. A beta infusion into the MBN elicited significant anxiety in the elevated plus maze. A beta toxicity on cholinergic MBN neurons, expressed as the loss of acetylcholinesterase-positive cortical projections, was accompanied by sprouting of serotonergic projection fibers in the MBN. In contrast, the loss of serotonin-positive fiber projections, decreased concentrations of both serotonin and 5-hydroxyindoleacetic acid, and decline of cortical 5-HT(1A) receptor binding sites indicated reduced serotonergic activity in the somatosensory cortex. In conclusion, the A beta-induced primary cholinergic deficit in the MBN and subsequent cortical cholinergic denervation bidirectionally modulate serotonergic parameters in the rat basal forebrain and cerebral cortex. We assume that enhanced serotonin immunoreactivity in the damaged MBN indicates intrinsic processes facilitating neuronal recovery and cellular repair mechanisms, while diminished cortical serotonergic activity correlates with the loss of the subcortical cholinergic input, thereby maintaining the balance of neurotransmitter concentrations in the cerebral cortex.

Postnatal development of unipolar brush cells in the cerebellar cortex of cat.

The postnatal developmental distribution pattern of metabotropic glutamate receptor (mGluR1a) immunoreactive unipolar brush cells (UBCs) was studied in the cerebellar cortex of kittens. On the day of birth (P0) UBCs are already present in the white matter in lobule X of the vermis, but only a few of these cell seemed to migrate to the deeper region of the internal granular layer. By the end of the first week (P8) UBCs were seen to invade the white matter + internal granular layer of lobules IX, VIII, I, and II of the vermis, and they spread further in the transitory area medio-laterally from the vermis toward the cerebellar hemispheres. By P15, UBCs appeared in lobules III and VII of the vermis, as well as in corresponding lobules of the neocerebellum, with especially high numbers in lobule VII. By P22, UBCs migrated further after their medio-lateral course in the neocerebellum, and began to invade lobules V and VI. At P62 the amount of UBCs in midsagittal planes of early developing vermal lobules (I, II, VII-X) resembled the P132 or adult pattern. The medio-lateral migration and incorporation of UBCs into the late-developing cerebellar lobules V and VI was completed only by P132, when the spatial distribution of UBCs in both the vermal and neocerebellar lobules was comparable to that seen in the 1 year old young adult cat. Although by P132 the postnatal migration of the vast majority of UBCs seemed to be completed, in the cerebellum of adult cats a few migrating UBCs could still be observed in the white matter of the cerebellar lobules, and beneath the ependyma of the fourth ventricle. It is concluded that during ontogenesis the migration course of UBCs follows essentially the developmental sequence of cerebellar lobules, although the incorporation of UBCs into the internal granular layer continues until 4 months postnatally, i.e., much beyond the apparent completion (about two months postnatally) of cytoarchitectonic built up of the cerebellar cortex of kittens.

Immunocytochemical mapping of NPY and VIP neuronal elements in the cat subcortical visual nuclei, with special reference to the pretectum and accessory optic system.

The aim of this study was to describe the distribution patterns of neuropeptide Y (NPY) and vasoactive intestinal polypeptide (VIP)-immunoreactive (ir) neuronal elements in subcortical visual centers of the cat. Numerous NPY-ir neurons were present in the feline nucleus of the optic tract and in the anterior pretectal nucleus. Only a few NPY-ir neurons were found in the posterior, medial and olivary pretectal nuclei and in the accessory optic nuclei. Diffuse and heavily beaded NPY-ir fiber plexuses were observed throughout the superior colliculus, pretectum, and accessory optic system. Extensively arborising NPY-ir fibers were present in the mesencephalon and ventral lateral geniculate nucleus, while the dorsal visual thalamic nuclei contained only a few NPY-ir fibers. VIP-ir cells were present mainly in the accessory optic nuclei, and they were absent in the dorsal visual thalamus. Both NPY- and VIP-ir neurons were multipolar and fusiform in shape in the regions studied. Enucleation did not alter the appearance of NPY- and VIP-containing neuronal elements in the superior colliculus and pretectum while in the thalamus a subset of NPY-ir fiber population disappeared, indicating their retinal origin. Although there is a partial overlap in the topographical localization of the NPY- and VIP-ergic neurons in the pretectum, the colocalization of the two peptides could not be demonstrated. The present observations demonstrate the existence of two different and separate peptidergic (NPY and VIP) neuronal populations in the pretectum.

Metabotrop glutamate receptor type 1a expressing unipolar brush cells in the cerebellar cortex of different species: a comparative quantitative study.

Morphology, distribution and number of unipolar brush cells (UBCs) was studied in the cerebellar vermal lobules I-X of the chicken, rat, guinea pig, cat, and monkey using monoclonal mGluR1a antibody as a marker to visualise these recently described nerve cells (Mugnaini and Floris [1994] J. Comp. Neurol. 339:174-180; Mugnaini et al. [1994] Synapse 16:284-311). The morphological appearance of mGluR1a immunopositive UBCs is similar in all species investigated: they are small cells, having a single, relatively short and thick dendrite, terminating in brush-like dendrioles. Although this, probably excitatory, cell type can be found all over the cerebellar cortex, highest density of UBCs can be seen in the vermal cortex. The present study, therefore, was focused on the quantitative morphology and distribution of UBCs in the 10 lobules of the vermis. Calculating the number of UBCs/l Purkinje cell (PC), we have found differences in this value (average in vermal lobules I-X) from 1.04 in rat, 1.10 in chicken, 1.16 in guinea pig, 2.27 in monkey, and up to 2.44 in cat. The highest density of UBCs was observed in lobules I, IX, and X, whereas the lowest number of UBCs/l PC was found in lobules IV-VI (in the mammals) and in lobules VII-VIII (in the chicken). In mammals, particularly the monkey and cat, an increased presence of UBCs was observed in vermal sub-lobules VIc-VIIb,c, a region defined as the oculomotor vermis because of its role in the control of saccadic eye movement. There is also a basic difference between chicken and mammals in the distribution of UBCs within the lobules: in mammals, the lowest density of these nerve cells was found in the peripheral portion of the lobules, near to the pia, while in the chicken, in contrast, the density of UBCs was the highest subpially with fewer UBCs located in the deepest curvature of the lobules. Finally, the functional significance of the differences in the density and in the distribution pattern of UBCs in the cerebellar vermis between the phylogenetically different species investigated is briefly discussed.

Mossy cells of the rat dentate gyrus are immunoreactive for calcitonin gene-related peptide (CGRP).

The neuropeptide calcitonin gene-related peptide (CGRP) was localized in the hippocampus and dentate gyrus of the rat by immunocytochemistry at the light and electron microscopic levels. Without colchicine treatment only faint neuropil labelling was found in the inner molecular layer of the dentate gyrus. Following colchicine treatment, a large number of neurons with numerous complex spines along the proximal dendrites were visualized in the hilus of the dentate gyrus, particularly in the ventral areas, and, in addition, staining of the inner molecular layer became stronger. Several CA3c pyramidal cells located adjacent to the hilar region in the ventral hippocampus also appeared to be faintly positive, although in most cases only their axon initial segments were labelled. Outside this region, the subicular end of the CA1 subfield contained occasional CGRP-positive non-pyramidal cells. The hilar CGRP-positive neurons were negative for parvalbumin, calretinin, cholecystokinin and somatostatin, whereas most of them were immunoreactive for GluR2/3 (the AMPA-type glutamate receptor known to be expressed largely by principal cells). Correlated electron microscopy showed that the spines along the proximal dendritic shafts indeed correspond to thorny excrescences engulfed by large complex mossy terminals forming asymmetrical synapses. Pre-embedding immunogold staining demonstrated that CGRP immunoreactivity in the inner molecular layer was confined to axon terminals that form asymmetrical synapses, and the labelling was associated with large dense-core vesicles. The present data provide direct evidence that CGRP is present in mossy cells of the dentate gyrus and to a lesser degree in CA3c pyramidal cells of the ventral hippocampus. These CGRP-containing principal cells terminate largely in the inner molecular layer of the dentate gyrus, and may release the neuropeptide in conjunction with their 'classical' neurotransmitter, glutamate.

Light and electron microscopic demonstration of mGluR5 metabotropic glutamate receptor immunoreactive neuronal elements in the rat cerebellar cortex.

The cellular and subcellular localization of the mGluR5 metabotropic glutamate receptor subtype was studied in the rat cerebellar cortex, by using the preembedding immunoperoxidase and immunogold techniques. Light microscopic observations revealed an abundant, intense labeling of neurons in the granular layer as well as in the molecular layer. Lugaro and Golgi cells exhibited an intense mGluR5 immunoreactivity, while only a fraction of the neurons in the molecular layer were found to be mGluR5 immunopositive. In addition to a dense plexus of immunoreactive dendrites in the molecular layer of the cerebellar cortex, the mGluR5 immunopositive Golgi cell dendrites resembling axons at the light microscopic level were also labeled in the granular layer. At the ultrastructural level, mGluR5 immunoreactivity was present in neuronal elements postsynaptic to axon terminals of different morphology. By using a pre-embedding immunogold method, it was found that mGluR5 immunoreactivity is accumulated at the plasma membranes extrasynaptically as well as at the periphery of the postsynaptic specializations, mainly of the parallel fiber synaptic contacts. These findings provide morphological evidence that mGluR5 is expressed by a population of neurons in the cerebellar cortex and can synaptically be activated via the parallel fiber system.

VIP fibers in rat optic chiasm and optic nerve arising from the hypothalamus.

This is the first report showing VIP fibers in the optic chiasm and the optic nerves of intact rats. These fibers form a fan-shaped dorso-medial bundle in the optic nerves. After colchicine injection into the vitreous body VIP fibers could be followed farther in the optic nerve toward the eye when compared to intact rats. After removal of eyes (enucleation) the VIP fiber-bundle became more prominent and VIP immunoreactive perikarya appeared in the supraoptic and para ventricular nuclei. When five-nine months after the enucleation Phaseolus vulgaris leucoagglutinin was administered to the paraventricular or supraoptic area, the anterogradely transported tracer was demonstrated in the optic nerve. These observations suggest the existence of a hypothalamic projection to the eye, which is, at least in part, VIP immunoreactive.

Metabotropic glutamate receptor in GHRH and beta-endorphin neurones of the hypothalamic arcuate nucleus.

Growth hormone-releasing hormone (GHRH) and beta-endorphin are mainly synthesized in neurones of the hypothalamic arcuate nucleus. Arcuate neurones also contain both ionotropic and metabotropic glutamate receptors. The aim of present study was to investigate whether glutamate receptors are present in GHRH and beta-endorphin containing nerve cells of this hypothalamic area. Using double-label immunocytochemistry as well as the mirror technique, we found that almost all GHRH and beta-endorphin immunoreactive arcuate neurones contain the metabotropic glutamate receptor la. The observations provide morphological evidence for the view that glutamate, which appears to be a major excitatory neurotransmitter in the hypothalamus, may directly stimulate GHRH and beta-endorphin neurones of the medial hypothalamus.

New aspects of the neuroendocrine role of PACAP.

The presence of PACAP was revealed in the anterior pituitary with RIA, HPLC, and with the demonstration of its mRNA. The level of PACAP mRNA in the anterior pituitary is the highest during the proestrous LH surge. In our immunohistochemical studies we were able to demonstrate PACAP immunoreactive cells in the anterior pituitary. The shape and the distribution of PACAP immunoreactive cells were very similar to that of the gonadotropes; however, the number of PACAP cells was less than that of LH cells. Additionally, another PACAP-positive cell population with small diameter appeared in the proestrous stage, during pregnancy and lactation. Double labeling revealed that the major part of large PACAP cells exhibited LH immunoreactivity and those with a small diameter contained PRL. It is not clear whether the pituitary- or the hypothalamic-born PACAP, or both, influence pituitary LH and PRL secretion. I.c.v. administration of PACAP just prior to the critical period in the proestrous stage inhibited the expected ovulation and blocked the proestrus LH and PRL surge, although i.v. administration of PACAP had no effect. PACAP antiserum did not interfere with ovulation when i.c.v. or i.v. injection was used. Our results support the view that PACAP has a role in the control of LH and PRL secretion during the estrous cycle, pregnancy, and lactation. The inhibitory effect of PACAP on ovulation is mediated through the hypothalamus.

Different populations of vasoactive intestinal polypeptide-immunoreactive interneurons are specialized to control pyramidal cells or interneurons in the hippocampus.

The postsynaptic targets of three vasoactive intestinal polypeptide-containing GABAergic interneuron types were examined in the rat hippocampus. Two of them showed remarkable target selectivity for other GABAergic neurons, while the third contacted the somata and proximal dendrites of pyramidal cells. Vasoactive intestinal polypeptide-positive interneurons innervating the stratum oriens/alveus border in the CA1 region were shown to establish multiple contacts with horizontal GABAergic interneurons immunoreactive for type 1 metabotropic glutamate receptor. Similarly, identified axons of vasoactive intestinal polypeptide-positive interneurons projecting to stratum radiatum were found to establish symmetrical synapses largely on GABAergic dendrites. The majority of these postsynaptic GABAergic neurons were shown to contain calbindin or vasoactive intestinal polypeptide. In contrast to the first two vasoactive intestinal polypeptide-containing cell populations, vasoactive intestinal polypeptide-positive interneurons arborizing in stratum pyramidale formed baskets around pyramidal cells. These results revealed a new element in cortical microcircuits, interneurons which are specialized to innervate other GABAergic interneurons. The role of this new component may be the synchronization of dendritic inhibition, or an input-specific disinhibition of pyramidal cells in various dendritic domains. In contrast, vasoactive intestinal polypeptide-containing basket cells are likely to be involved in perisomatic inhibition of pyramidal neurons, and represents a new basket cell type different from that containing parvalbumin.

Immunocytochemical visualization of the mGluR1a metabotropic glutamate receptor at synapses of corticothalamic terminals originating from area 17 of the rat.

Pre-embedding immunogold histochemistry was combined with Phaseolus vulgaris leucoagglutinin anterograde tract tracing in order to analyse the relationship between the subcellular localization of the GluR1a metabotropic glutamate receptors and the distribution of corticothalamic synapses in the dorsal lateral geniculate nucleus (dLGN) and the lateral posterior nucleus (LP) of the rat. The injection of the tracer into area 17 labelled two types of corticothalamic terminals: (i) the small boutons constituting the majority of the labelled fibres which form asymmetrical synapses both in the dLGN and LP; and (ii) the giant terminals typically participating in glomerulus-like synaptic arrangements and found exclusively in the lateral posterior nucleus. The small corticothalamic terminals often established synapses with mGluR1a-immunopositive dendrites, with immunometal particles concentrated at the periphery of their postsynaptic membranes. In contrast, the synapses formed by giant boutons in the lateral posterior nucleus were always mGluR1a-immunonegative. We conclude that the corticothalamic fibres forming the small synaptic terminals are the most likely candidates for the postulated mGluR-mediated modulation of visual information flow by corticothalamic feedback mechanisms.

Light and electron microscopic analysis of synaptic input from cortical area 17 to the lateral posterior nucleus in cats.

The morphology and synaptic organization of the corticothalamic (CT) fibres from area 17 were studied in the lateral posterior nucleus (LP) of the thalamus in cats. Injection of the anterograde tracer Phaseolus vulgaris leucoagglutinin (PHAL) into primary visual cortex labelled a band of CT fibres in the LP with terminal field confined to its lateral division "LP1". PHAL-labelled CT axons in the LP1 gave rise to both en passant and terminal boutons. They usually established several synaptic contacts--often in complex glomerulus-like synaptic arrangements--with dendritic shafts of large diameter and presynaptic dendrites containing pleomorphic vesicles. Postsynaptic targets of the PHAL-labelled CT boutons were characterized by postembedding gamma-aminobutyric acid (GABA) immunocytochemistry. It appeared that, in the LP1 of the cat, almost half (44.5%) of the postsynaptic dendrites to CT boutons from area 17 belonged to the GABA-immunopositive interneurons and the majority (41%) of these GABA-immunopositive dendrites were F2 terminals. These results indicate that the CT axons from the striate cortex in the LP of the cat, in addition to a direct excitatory action, exert a powerful feed-forward inhibition on the thalamic principal cells.

Immunohistochemical localization of calcitonin gene-related peptide in the terete nucleus of the rat hypothalamus.

The topographical distribution of calcitonin gene-related peptide (CGRP) containing neuronal elements in the posterior hypothalamus has been re-examined by means of indirect labeled immunohistochemistry in the rat. In contrast to previous findings, no CGRP-like immunoreactive neuronal perikarya were present in the premamillary nuclei, but they form a small group of cells, which is identical to the recently described terete nucleus. In coronal sections, this small and rounded cluster of intermingled CGRP-immunopositive perikarya and fibers is situated in the lateral hypothalamus at the premamillary level, ventrolateral to the fornix. Perikarya, dendrites, presynaptic terminals, as well as nonmyelinated and some myelinated axons were labeled by CGRP-like immunoreactive material within and in the immediate vicinity of the terete nucleus. On some of the CGRP-positive neurons, neuropeptide Y-immunoreactive axon terminals establish synaptic contacts.

Immunocytochemical localization of mGluR1a metabotropic glutamate receptor in inhibitory interneurons of the cerebellar cortex.

Immunocytochemical investigation of metabotrop glutamate receptor la in the cerebellar cortex of rat revealed a presence of this receptor in all cortical inhibitory nerve cells (Purkinje, basket, stellate, Lugaro and Golgi). In addition to the previously described localization of this receptor in Purkinje dendritic spines supplied by parallel and climbing fibres and considered to be responsible for cerebellar long-term depression, we have observed metabotropic glutamate receptor la immunopositivity also in the synapses between parallel fibres and dendrites of basket, stellate and Golgi neurons. The postsynaptic receptor was also present occasionally in inhibitory synapses between stellate cell axons and Purkinje cells as well as between Purkinje axon collaterals and Lugaro cell dendrites. The possibility, that mGluR1a receptors in basket, Golgi and stellate cells might directly or indirectly contribute to cerebellar LTD, is discussed.

PACAP participates in the regulation of the hormonal events preceeding the ovulation.

Pituitary adenylate cyclase activating polypeptide (PACAP) is a member of the secretin family. It was isolated and characterized in 1989. Its neuroendocrine role was demonstrated in vivo and in vitro systems. It seems that in vivo the effect of PACAP on the gonadotrop hormone secretion depends on the route of administration. It was reported that intravenous (i.v.) injection of PACAP elevated, while intra-cerebro-ventricular (i.c.v.) administration depressed plasma LH levels. In the present study it was demonstrated that PACAP, administered i.c.v. before the critical period of the proestrous stage, blocked the ovulation and prevented the proestrous LH surge in rats. The blocking effect of PACAP is not directly mediated by endogenous opioids because the antagonizing effect of Naloxone, an opioid receptor antagonist, was questionable. Under our experimental conditions we could not confirm the stimulating effect of i.v. administered PACAP.

Distribution of metabotropic glutamate receptor 1a in the rat hypothalamus: an immunocytochemical study using monoclonal and polyclonal antibody.

L-glutamate appears to be a major excitatory neurotransmitter in the hypothalamus. Its action is mediated via ionotropic and metabotropic glutamate receptors (mGluR). Eight mGluRs have already been cloned. In the present study the hypothalamic distribution of mGluR1a has been investigated by immunocytochemistry using monoclonal antibodies recently produced by some of the present authors (T. J. G., R. K., T. K.). The observations have been compared with findings obtained with a polyclonal antibody. A widespread and heterogeneous distribution of mGluR1a was found with the monoclonal antibodies. Intense immunolabelling of perikarya and dendrites occurred in several hypothalamic cell groups including the suprachiasmatic, anterior periventricular, anterior hypothalamic (posterior part), paraventricular, supraoptic, arcuate, tuberal magnocellular, dorsomedial and mammillary nuclei (particularly in the medial). It was only the ventromedial nucleus in which several perikarya were stained by the polyclonal antibody but appeared to be negative by the monoclonal antibodies. The findings fit extremely well with the data on the hypothalamic distribution of mGluR1 mRNA with the exception of the ventromedial nucleus. It remains to be elucidated whether alternatively spliced variants of mGluR1 (mGluR1b and 1c) are expressed in this nucleus. Further, they confirm the results of former immunohistochemical studies. In addition, they indicate that a significant part of the neuroendocrine region of the hypothalamus (including the paraventricular, supraoptic and arcuate nuclei) also contains mGluR1 suggesting that this receptor may play a role also in neuroendocrine regulation.

Different ultrastructural localization of VIP and prolactin in anterior pituitary cells of rats chronically treated with estrogen.

In the present study we investigated the effect of a long-term estrogen treatment on the intracellular distribution of VIP immunoreactivity in pituitary prolactin cells using double-labeling immunocytochemistry. With the use of pre-embedding ABC method it was found that VIP immunoreactivity was associated with the outer surface of membrane-bound organelles, and was not found in secretory granules. However, prolactin immunoreactivity demonstrated by postembedding immunogold technique was mainly associated within the secretory granules of the same cells. The discrepancy between our and Hsu et al.'s results (1989), who observed VIP immunoreactivity in secretory granules of human anterior pituitary cells, may be owing to the overstimulation of VIP cells by estrogen. It is possible that estrogen treatment depleted the VIP content of the secretory granules and enhanced the cytosolic VIP. The appearance of an alternative form of VIP in estrogen-treated rats with preferential distribution in the cytosol cannot be excluded.

Immunohistochemical mapping of neuropeptides in the premamillary region of the hypothalamus in rats.

The topographical distribution of neuropeptide-containing cell bodies, fibers and terminals was studied in the premamillary region of the rat hypothalamus using light microscopic immunohistochemistry. Alternate coronal sections through the posterior third of the hypothalamus of normal and colchicine-treated male rats were immunostained for 19 different neuropeptides and their distributions were mapped throughout the following structures: the ventral and dorsal premamillary, the supramamillary, the tuberomamillary and the posterior hypothalamic nuclei, as well as the premamillary portion of the arcuate nucleus and the postinfundibular median eminence. Seventeen of the investigated neuropeptides were present in neuronal perikarya, nerve fibers and terminals while the gonadotropin associated peptide and vasopressin occurred only in fibers and terminals. Growth hormone-releasing hormone-, somatostatin-, alpha-melanocyte stimulating hormone-, adrenocorticotropin-, beta-endorphin- and neuropeptide Y-immunoreactive neurons were seen exclusively in the premamillary portion of the arcuate nucleus. Thyrotropin-releasing hormone-, dynorphin A- and galanin-containing neurons were distributed mainly in the arcuate and the tuberomamillary nuclei. A high number of methionine- and leucine-enkephalin-immunoreactive cells were detected in the arcuate and dorsal premamillary nuclei, as well as in the area ventrolateral to the fornix. Substance P-immunoreactive perikarya were present in very high number within the entire region, in particular in the ventral and dorsal premamillary nuclei. Cell bodies labelled with cholecystokinin- and calcitonin gene-related peptide antisera were found predominantly in the supramamillary and the terete nuclei, respectively. Corticotropin-releasing hormone-, vasoactive intestinal polypeptide- and neurotensin-immunoreactive neurons were scattered randomly in low number, mostly in the arcuate and the ventral and dorsal premamillary nuclei. Peptidergic fibers were distributed unevenly throughout the whole region, with each peptide showing an individual distribution pattern. The highest density of immunoreactive fibers was presented in the ventral half of the region including the arcuate, the ventral premamillary and the tuberomamillary nuclei. The supramamillary nucleus showed moderately dense fiber networks, while the dorsal premamillary and the posterior hypothalamic nuclei were poor in peptidergic fibers.