Comparison of the distribution of D-1 and D-2 dopamine receptors in the rat brain

The distribution of dopamine type 1 (D-1) and dopamine type 2 (D-2) receptors in the brain have been compared as assessed by the technique of autoradiography after labelling with highly selective ligands. D-1 receptors, as evidenced by the specific binding of [3H]R-(+)-8-chloro-2,3,4,5-tetrahydro-3-methyl-5-phenyl-IH-3-benzazepine -7- ol (SCH 23390), were found in high concentrations in the caudate-putamen, nucleus accumbens, islands of Calleja, olfactory tubercle and the zona reticulata of the substantia nigra. A similar but distinct distribution was seen for [3H]sulpiride, a ligand which is highly selective for D-2 receptors. Like [3H]SCH 23390, this ligand also labelled the caudate-putamen, nucleus accumbens, islands of Calleja and the olfactory tubercle; however, only a very low density of D-2 receptors could be found in the zona reticulata of the substantia nigra, while a greater degree of binding was present in the zona compacta. Additional brain areas which contained D-1 but not D-2 receptors included the cerebral cortex, accessory olfactory nucleus, amygdala, thalamus, suprachiasmatic nucleus, choroid plexus, claustrum, endopiriform nucleus, zona incerta, dorsal lateral geniculate nucleus and the dentate gyrus. D-2 receptors were also found in areas which appeared to contain only low amounts of D-1 receptors such as the glomerular layer of the olfactory bulb, bed nucleus of the stria terminalis, hypothalamus, habenula, stratum lacunosum moleculare of the hippocampus, intermediate lobe of the pituitary, lateral mammillary nucleus, periaqueductal gray, inferior colliculus, nodulus of the cerebellum and the dorsal horn of the spinal cord. The results show the precise localization of dopamine receptors throughout the brain and provide a means of direct comparison between the distribution of dopamine receptor subtypes. These subtypes are pharmacologically and anatomically distinct entities and their comparison indicates areas where additional biochemical and neuroanatomical studies may be performed to elucidate the roles for these receptor subtypes in the central nervous system.     

Mapping of benzodiazepine-like immunoreactivity in the rat brain as revealed by a monoclonal antibody to benzodiazepines.

A monoclonal antibody against benzodiazepines (21-7F9) was used to study the distribution of benzodiazepine-like immunoreactivity in the rat brain. Immunodensitometry in combination with image analysis were used for quantification. The results showed a ubiquitous distribution of benzodiazepine-like immunoreactivity throughout the brain. Very high levels of benzodiazepine-like immunoreactivity were found in the Purkinje cell layer of the cerebellum, in the primary olfactory cortex, in the stratum pyramidale of the hippocampus and in the mitral cell layer of the olfactory bulb. High densities of benzodiazepine-like immunoreactivity were found in the granule cell layer of the cerebellum, the pyramidal cell layer of the olfactory tubercle, the granule layer of the dentate gyrus, the arcuate nucleus of the hypothalamus, the mammillary bodies, the interstitial nucleus of Cajal and superficial grey layer of superior colliculus. The substantia nigra pars compacta, the islands of Calleja and layers II, III, V and VI of the cerebral cortex had moderate levels of benzodiazepine-like immunoreactivity. Lower densities were found in the internal granular layer and the external plexiform layer of the olfactory bulb, in the molecular layer of the dentate gyrus, in layers I and IV of the cerebral cortex, in the nucleus caudate-putamen and most of the thalamic nuclei. The lowest density of immunoreactivity was found in the globus pallidus, and the strata radiatum, oriens and lacunosum-moleculare of the hippocampus. The distribution of endogenous benzodiazepine-like immunoreactivity was compared with the distribution of the GABA/benzodiazepine receptor by using both immunocytochemistry and receptor autoradiography. Our studies have shown a clear mismatch between the localization of the benzodiazepine-like immunoreactivity and the GABA/benzodiazepine receptors.     

The distribution of m4 muscarinic acetylcholine receptors in the islands of Calleja and striatum of rats and cynomolgus monkeys

The distribution of m4 muscarinic acetylcholine receptors, and their relation to a number other markers, was examined using immunocytochemical techniques. Staining in the dorsal striatum tended to be more pronounced in the striosomal than the matrix compartment of both rats and cynomolgus monkeys. Within the ventral striatum, immunoreactivity was more pronounced within the olfactory tubercle and the shell region of the nucleus accumbens than in the nucleus accumbens core and was especially marked within the lateral striatal stripe. Modest staining was also seen in the external plexiform layer of the olfactory bulb. By far, the most intense staining in the forebrain of both rats and cynomolgus monkeys was found in islands of Calleja, where it appeared to be a selective marker for the core or hilus regions of the islands, or an analogous region found adjacent to them. The core regions of different islands appear to be continuous with each other so as to form a complex three-dimensional structure, which is largely encased by layers of granule cells. The neuronal elements in the islands of Calleja, which express m4 receptors, remain to be identified, but it is unlikely that cholinergic neurons are a major locus of these receptors. Although there are certain similarities between the islands of Calleja and other components of the striatal complex, the current studies emphasize the extent to which the islands are unique in terms of their architecture and chemical anatomy.     

Temporal and spatial relationships between lipopolysaccharide-induced expression of Fos, interleukin-1beta and inducible nitric oxide synthase in rat brain

Interleukin-1beta plays an important role in mediating central components of the host response to peripheral infection such as fever and neuroendocrine activation by acting in the brain. The present study assessed whether interleukin-1beta produced in the brain is relevant to neuronal activation and the fever response induced by intraperitoneal injection of bacterial lipopolysaccharide. The distributions of Fos protein, interleukin-1beta protein and inducible nitric oxide synthase messenger RNA, used as an anatomical indicator of interleukin-1beta bioactivity, were compared in brains of animals killed 2, 4 or 8 h after lipopolysaccharide (250 microg/kg) or saline injection. Saline did not induce interleukin-1beta or Fos immunoreactivity in the brain. Interleukin-1beta positive cells were found 2 h after lipopolysaccharide injection in circumventricular organs. Fos immunoreactivity at this time-point was not found in circumventricular organs, but in parenchymal structures such as the nucleus of the solitary tract, paraventricular hypothalamus and ventromedial preoptic area. Fos expression did occur in circumventricular organs only 8 h after lipopolysaccharide injection. This late pattern of Fos expression coincided with the rise in body temperature and the induction of inducible nitric oxide synthase messenger RNA. These data show that after peripheral lipopolysaccharide administration interleukin-1beta is synthesized and bioactive in circumventricular organs. Interleukin-1beta may activate local neurons that induce fever and neuroendocrine activation via projections to the ventromedial preoptic area and the nucleus of the solitary tract.     

Distribution of [125I]angiotensin II binding sites in the rat brain: a quantitative autoradiographic study

Angiotensin II receptors have been localized by quantitative autoradiography in the rat central nervous system after labeling with [125I]angiotensin II. A highly discrete distribution of these receptors was found throughout the rat brain. The highest density was seen in regions of the medulla, hypothalamus and circumventricular organs where angiotensin II could potentially produce cardiovascular, dipsogenic and neuroendocrine responses. The distribution of angiotensin II receptors correlates relatively well with the previously reported distribution of angiotensin immunoreactive nerve terminals as well as areas determined by various physiological techniques to be sensitive to angiotensin II. Finally, the anatomical localization of angiotensin II receptor populations has revealed several areas of the brain where the effects of this peptide have not been investigated. Many of these nuclei are involved in the transmission and processing of somatic and visceral sensory information. These results suggest a broader role for the central renin-angiotensin system in modulating several types of sensory input.     

Distribution of Serotonin 4(a) Receptors in the juvenile Rat Brain and Spinal Cord

In the last 20 years there have been many studies investigating the distribution of 5-HT₄-receptors in the brain of different species. Most studies are methodically based on RT-PCR or in situ hybridization and have analysed the receptors at the mRNA level.Furthermore there have been some autoradiographic studies using specific 5-HT₄-receptor antagonists like [³H]GR113808, [³H]BIMU-1 or [¹²⁵I]SB207710.This study investigates the topographical distribution of the 5-HT_4(a)-receptor in the juvenile rat brain and spinal cord, which is important for neuromodulation of cellular excitability and could be involved in various developmental processes of the central nervous system. We analysed the 5-HT_4(a)-receptor at protein level with a monospecific polyclonal antibody by using an immunohistochemical staining.We saw an intensive staining in some areas of the cortex, in the olfactory bulb, in most areas of the cerebellum, in hippocampal areas like the dentate gyrus and in several different areas of the brainstem, especially in the motor nuclei. Overall we have shown comparable results in accordance with the results of other studies investigating the distribution of 5-HT_4(a)-receptors.Some areas like the islands of Calleja, the preoptic nucleus or the medial habenular nucleus showed a lower intensity of 5-HT_4(a)-receptors in comparison with the results of other studies. As a novel result we found a higher intensity of 5-HT_4(a)-receptor in several brain areas associated with motor function than was shown by other studies, especially in the motor cortex, in different areas of the cerebellum, in the red nucleus, in the motor nuclei of the brainstem or in the ventral horn cells of the spinal cord.We conclude that the 5-HT_4(a)-receptor may play a more prominent role in the modulation of motor cortico-ponto-cerebellar, cortico-spinal, rubro-spinal, vestibulo-spinal and cortico-nuclear tracts during juvenile development.     

Angiotensin converting enzyme in the monkey (Macaca fascicularis) brain visualized by in vitro autoradiography.

Angiotensin converting enzyme is localized in the monkey (Macaca fascicularis) brain by in vitro autoradiography using the radiolabelled inhibitor, [125I]351A. This radioligand binds with high affinity and specificity to monkey cortical sections. Specific inhibitors of converting enzyme, lisinopril and perindoprilat complete for the radioligand binding to monkey cortex sections with inhibitory constants of 10 nM. High concentrations of angiotensin converting enzyme occur in most components of the basal ganglia including the caudate nucleus, putamen, the internal and external globus pallidus, nucleus accumbens, ventral pallidum and the reticular part of the substantia nigra. The distribution of converting enzyme in the caudate nucleus and putamen is heterogeneous, with prominent patches of higher activity. The patches in the caudate nucleus correspond closely with the acetylcholinesterase-poor striosomes. In the hypothalamus, very high levels of angiotensin converting enzyme occur in the median eminence and the pituitary stalk and high concentrations occur in the supraoptic and suprachiasmatic nuclei. Moderate, diffuse binding is observed in the median preoptic nucleus, the medial preoptic area, and in the anterior, lateral, ventromedial, posterior and arcuate nuclei. In the circumventricular organs, the subcommissural and subfornical organs exhibit high levels of angiotensin converting enzyme. The organum vasculosum of the lamina terminalis and the pineal body display moderate enzyme activities whereas the area postrema is devoid of labelling. The interpeduncular nucleus and, in the hippocampal formation, the molecular layer of the dentate gyrus are also intensely labelled. High levels of angiotensin converting enzyme activity are also detected throughout the cerebral cortex with laminations of higher activity corresponding to cell dense layers of the cortex. Layered binding is also present in the cerebellar cortex, with the most intense labelling in the molecular layer. Moderate concentrations of converting enzyme also occur in the paraventricular, medial habenula, lateral habenula and central median nuclei of the thalamus, the amygdala, the central gray, the locus coeruleus, the parabrachial nucleus and dorsal tegmental nucleus. The dorsal vagal complex, inferior olivary nucleus and the caudal subnucleus of the spinal trigeminal nucleus all display high levels of binding. Moderate, diffuse labelling is found throughout the reticular region and is also present in the gracile and cuneate nuclei. Although the overall distribution of angiotensin converting enzyme in the monkey brain resembles that in the rat, there are some striking differences. These include the high levels of binding throughout the monkey cerebral cortex and in the interpeduncular and suprachiasmatic nuclei.     

A detailed mapping of histamine H1-receptors in guinea-pig central nervous system established by autoradiography with [125I]iodobolpyramine

[125I]Iodobolpyramine, a potent and selective histamine H1-receptor antagonist derived from mepyramine, was used to generate light microscopic autoradiograms on sections of guinea-pig brain and spinal cord. Histamine H1-receptors were labelled with high sensitivity over a low background as determined using mianserin or other H1-receptor antagonists as competing agents. An atlas of H1-receptors was established using five sagittal sections and 39 frontal sections, the latter serially prepared at 50 micron intervals. Labelled areas were identified by comparison with corresponding, classically stained sections and their density was rated according to an arbitrary scale. Autoradiographic grains were detected in a large variety of gray matter areas whereas they were generally absent from white matter areas. In the cerebral cortex, H1-receptors are present in all areas and layers with a higher density in lamina IV. In the hippocampal formation, H1-receptors display a laminated pattern of distribution and are the most abundant in the dentate gyrus (hilus and molecular layer) and in several areas of the subiculum and commissural complex. In the amygdaloid complex, the highest densities are found in the medial group of nuclei. In the basal forebrain, the striatum is moderately labelled whereas the nucleus accumbens, islands of Calleja and most septal nuclei are highly labelled. In the thalamus, H1-receptors are present in high density, particularly in the anterior, median and lateral groups of nuclei. In the hypothalamus the labelling is highly heterogeneous with high densities in, for example, medial preoptic area, dorsomedial, ventromedial and most posterior nuclei, including the tuberomammillary complex in which histaminergic perikarya and short axons are present.(ABSTRACT TRUNCATED AT 250 WORDS)     

Production of antisera to acidic fibroblast growth factor and their application to immunohistochemical study in rat brain

Antisera against acidic fibroblast growth factor purified from bovine brain were produced in rabbits and used for immunohistochemical study of the rat brain. When examined in an immunospot assay using a nitrocellulose membrane, the best antibody was capable of detecting 80 fmol of acidic fibroblast growth factor but failed to react even with up to 5 pmol of basic fibroblast growth factor.

Using this antiserum, the immunohistochemical distribution of acidic fibroblast growth factor was examined in rat brain. Acidic fibroblast growth factor-like immunoreactivity was localized mainly in a subpopulation of ependymal cells and tanycytes, as well as in some glial cells. Positive ependymal cells were observed throughout the walls of ventricles, including the third ventricle and cerebral aqueduct. Immunoreactive processes of tanycytes were found extending from the ventral wall of the third ventricle to the brain parenchyma and surface. The most intense immunostaining was observed in circumventricular organs such as the organum vasculosum laminae terminalis and the subfornical organ. Particularly in the latter organ, there was an extremely dense plexus of immunoreactive fibers and processes around the wall of capillaries.

The present results suggest that the effects of acidic fibroblast growth factor on brain functions may be exerted through the circumventricular organs and/or ependymal cells.     

D1 and D2 dopamine receptor mRNA expression in whole hemisphere sections of the human brain

Understanding dopamine signaling in human behavior requires knowledge of the distribution of all molecular components involved in dopamine pathways throughout the human brain. In the present study, the relative distributions of D1 and D2 dopamine receptor mRNAs were determined by in situ hybridization histochemistry in whole hemisphere sections from normal human post mortem brains. The findings confirmed information documented from single structure examination that the highest expression of both the D1 and D2 mRNAs were localized to the striatum. The cerebral cortex expressed moderate D1 mRNA in all regions with the highest signal in the medial orbital frontal area (Brodmann areas 11, 14), the paraterminal gyrus (Brodmann area 32) and the insular cortex (Brodmann areas 13-16), whereas the D2 mRNA expression had very low cortical expression. The bed nucleus of the stria terminalis and islands of Calleja had high expression of the D1 mRNA and moderate D2 mRNA levels. Moderate to high expression of the D2 mRNA was evident in the hippocampal formation, parafascicular and paraventricular thalamic nuclei, geniculate bodies, subthalamic nucleus, and pineal gland, all of which were devoid of, or showed only faint, D1 mRNA expression. Brainstem regions, e.g. substantia nigra, red nucleus, inferior colliculus, medial lemniscus, and pontine nuclei expressed D2, but not D1, mRNA. These results emphasize the differential anatomical localization of D1 and D2 dopamine receptor mRNA neuronal populations in the human brain. The restricted expression of the D1 mRNA to the cortical mantle and to a few forebrain structures indicates a strong involvement of the D1 system in cognitive function.     

Neuronal expression of cAMP-specific phosphodiesterase 7B mRNA in the rat brain

cAMP plays an important role as second messenger molecule controlling multiple cellular processes in the brain. cAMP levels depend critically on the phosphodiesterases (PDE) activity, enzymes responsible for the clearance of intracellular cAMP. We have examined the regional distribution and cellular localization of mRNA coding for the cAMP-specific phosphodiesterase 7B (PDE7B) in rat brain by in situ hybridization histochemistry. PDE7B mRNA is specifically distributed in rat brain, preferentially in neuronal cell populations. The highest levels of hybridization are observed in olfactory tubercle, islands of Calleja, dentate gyrus, caudate-putamen and some thalamic nuclei. Positive hybridization signals are also detected in other areas, such as cerebral cortex, Purkinje cells of the cerebellum and area postrema. By double in situ hybridization histochemistry, we found that 74% and 79% of the cells expressing PDE7B mRNA in striatum and olfactory tubercle, respectively, were GABAergic cells (expressing glutamic acid decarboxylase mRNA), in contrast with the lack of expression in the few cholinergic cells (expressing choline acetyltransferase mRNA) present in those two areas (around 0.4% in olfactory tubercle). In the thalamic nuclei, a majority of cells containing PDE7B mRNA also expresses a glutamatergic marker (76.7% express vesicular glutamate transporter vGluT1 and 76% express vGluT2 mRNAs). Almost all PDE7B expressing cells in dentate gyrus (93%) were glutamatergic. These results offer a neuroanatomical and neurochemical base that will support the search for specific functions for cAMP dependent PDEs and for the development of specific PDE7 inhibitors.     

Muscarinic M2 receptor mRNA expression and receptor binding in cholinergic and non-cholinergic cells in the rat brain: a correlative study using in situ hybridization histochemistry and receptor autoradiography.

The goal of the present study was to identify the cells containing mRNA coding for the m2 subtype of muscarinic cholinergic receptors in the rat brain. In situ hybridization histochemistry was used, with oligonucleotides as hybridization probes. The distribution of cholinergic cells was examined in consecutive sections with probes complementary to choline acetyltransferase mRNA. Furthermore, the microscopic distribution of muscarinic cholinergic binding sites was examined with a non-selective ligand ([3H]N-methylscopolamine) and with ligands proposed to be M1-selective ([3H]pirenzepine) or M2-selective ([3H]oxotremorine-M). The majority of choline acetyltransferase mRNA-rich (i.e. cholinergic) cell groups (medial septum-diagonal band complex, nucleus basalis, pedunculopontine and laterodorsal tegmental nuclei, nucleus parabigeminalis, several motor nuclei of the brainstem, motoneurons of the spinal cord), also contained m2 mRNA, strongly suggesting that at least a fraction of these receptors may be presynaptic autoreceptors. A few groups of cholinergic cells were an exception to this fact: the medial habenula and some cranial nerve nuclei (principal oculomotor, trochlear, abducens, dorsal motor nucleus of the vagus). Furthermore, m2 mRNA was not restricted to cholinergic cells but was also present in many other cells throughout the rat brain. The distribution of m2 mRNA was in good, although not complete, agreement with that of binding sites for the M2 preferential agonist [3H]oxotremorine-M, but not with [3H]pirenzepine binding sites. Regions where the presence of [3H]oxotremorine-M binding sites was not correlated with that of m2 mRNA are the caudate-putamen, nucleus accumbens, olfactory tubercle and islands of Calleja. The present results strongly suggest that the M2 receptor is expressed by a majority of cholinergic cells, where it probably plays a role as autoreceptor. However, many non-cholinergic neurons also express this receptor, which would be, presumably, postsynaptically located. Finally, comparison between the distribution of m2 mRNA and that of the proposed M2-selective ligand [3H]oxotremorine-M indicates that this ligand, in addition to M2 receptors, may also recognize in certain brain areas other muscarinic receptor populations, particularly M4.     

Cellular distribution of transferrin immunoreactivity in the developing rat brain

The cellular distribution of immunoreactive transferrin has previously been studied in adult rat brain with the conclusion that transferrin is largely confined to oligodendrocytes, although studies in the developing rodent brain indicated some staining of neurons. Detailed investigations of a wider range of ages than studied earlier have shown that in the developing rat brain, transferrin is not only present in oligodendrocytes but also in other glial cell types and in neurons and endothelial cells. The preponderant cell type which is positive for transferrin and its regional distribution are, however, related to the age of the animal except in most circumventricular organs in which strong transferrin immunoreactivity was seen throughout the developmental period studied.     

Immunohistochemical colocalization of delta sleep-inducing peptide and luteinizing hormone-releasing hormone in rabbit brain neurons

The anatomical distributions of luteinizing hormone-releasing hormone and delta sleep-inducing peptide immunoreactivity in the rabbit brain were studied by indirect immunofluorescence technique. The comparison of adjacent serial sections, one being immunolabeled with an antiserum to luteinizing hormone-releasing hormone, the other with an antiserum to delta sleep-inducing peptide, showed that the respective distribution patterns of immunoreactivity exhibited a remarkable overlap through the basal forebrain and hypothalamic regions. A sequential double-immunolabelling (elution-restaining method) clearly indicated that all the luteinizing hormone-releasing hormone-immunoreactive cell bodies displayed delta sleep-inducing peptide immunoreactivity. These cell bodies were sparse and mainly located throughout the septal-preoptico-suprachiasmatic region and the ventrolateral hypothalamus. The colocalization of luteinizing hormone-releasing hormone and delta sleep-inducing peptide immunoreactivity was also observed in many fibres supplying all these brain regions and terminal areas such as the organum vasculosum of the lamina terminalis, the subfornical organ, the median eminence and the pituitary stalk. These neuroanatomical findings are suggestive of interaction between delta sleep-inducing peptide and luteinizing hormone-releasing hormone in various brain areas including some circumventricular organs.     

Distribution of apelin-synthesizing neurons in the adult rat brain

The peptide apelin originating from a larger precursor preproapelin molecule has been recently isolated and identified as the endogenous ligand of the human orphan G protein-coupled receptor, APJ (putative receptor protein related to the angiotensin receptor AT(1)). We have shown recently that apelin and apelin receptor mRNA are expressed in brain and that the centrally injected apelin fragment K17F (Lys(1)-Phe-Arg-Arg-Gln-Arg-Pro-Arg-Leu-Ser-His-Lys-Gly-Pro-Met-Pro-Phe(17)) decreased vasopressin release and altered drinking behavior. Using a specific polyclonal antiserum against K17F for immunohistochemistry, the aim of the present study was to establish the precise topographical distribution of apelin immunoreactivity in colchicine-treated adult rat brain. Immunoreactivity was essentially detected in neuronal cell bodies and fibers throughout the entire neuroaxis in different densities. Cells bodies have been visualized in the preoptic region, the hypothalamic supraoptic and paraventricular nuclei and in the highest density, in the arcuate nucleus. Apelin immunoreactive cell bodies were also seen in the pons and the medulla oblongata. Apelin nerve fibers appear more widely distributed than neuronal apelin cell bodies. The hypothalamus represented, by far, the major site of apelin-positive nerve fibers which were found in the suprachiasmatic, periventricular, dorsomedial, ventromedial nuclei and in the retrochiasmatic area, with the highest density in the internal layer of the median eminence. Fibers were also found innervating other circumventricular organs such as the vascular organ of the lamina terminalis, the subfornical and the subcommissural organs and the area postrema. Apelin was also detected in the septum and the amygdala and in high density in the paraventricular thalamic nucleus, the periaqueductal central gray matter and dorsal raphe nucleus, the parabrachial and Barrington nuclei in the pons and in the nucleus of the solitary tract, lateral reticular, prepositus hypoglossal and spinal trigeminal nuclei.The topographical distribution of apelinergic neurons in the brain suggests multiple roles for apelin especially in the central control of ingestive behaviors, pituitary hormone release and circadian rhythms.     

Expression of the neuropeptide Y Y1 receptor in the CNS of rat and of wild-type and Y1 receptor knock-out mice. Focus on immunohistochemical localization

The distribution of neuropeptide Y (NPY) Y1 receptor-like immunoreactivity (Y1R-LI) has been studied in detail in the CNS of rat using a rabbit polyclonal antibody against the C-terminal 13 amino acids of the rat receptor protein. The indirect immunofluorescence technique with tyramide signal amplification has been employed. For specificity and comparative reasons Y1 knock-out mice and wild-type controls were analyzed. The distribution of Y1R mRNA was also studied using in situ hybridization. A limited comparison between Y1R-LI and NPY-LI was carried out.A widespread and abundant distribution of Y1R-LI, predominantly in processes but also in cell bodies, was observed. In fact, Y1R-LI was found in most regions of the CNS with a similar distribution pattern between rat and wild-type mouse. This staining was specific in the sense that it was absent in adjacent sections following preadsorption of the antibody with 10(-5) M of the antigenic peptide, and that it could not be observed in sections of the Y1 KO mouse. In contrast, the staining obtained with an N-terminally directed Y1R antiserum did not disappear, strongly suggesting unspecificity. In brief, very high levels of Y1R-LI were seen in the islands of Calleja, the anterior olfactory nucleus, the molecular layer of the dentate gyrus, parts of the habenula, the interpeduncular nucleus, the mammillary body, the spinal nucleus of the trigeminal, caudal part, the paratrigeminal nucleus, and superficial layers of the dorsal horn. High levels were found in most cortical areas, many thalamic nuclei, some subnuclei of the amygdaloid complex, the hypothalamus and the nucleus of the stria terminalis, the nucleus of the solitary tract, the parabrachial nucleus, and the inferior olive. Moderate levels of Y1R-LI were detected in the cornu Ammonis and the subicular complex, many septal, some thalamic and many brainstem regions. Y1R staining of processes, often fiber and/or dot-like, and occasional cell bodies was also seen in tracts, such as the lateral lemniscus, the rubrospinal tract and the spinal tract of the trigeminal. There was in general a good overlap between Y1R-LI and NPY-LI, but some exceptions were found. Thus, some areas had NPY innervation but apparently lacked Y1Rs, whereas in other regions Y1R-LI, but no or only few NPY-positive nerve endings could be detected.Our results demonstrate that NPY signalling through the Y1R is common in the rat (and mouse) CNS. Mostly the Y1R is postsynaptic but there are also presynaptic Y1Rs. Mostly there is a good match between NPY-releasing nerve endings and Y1Rs, but 'volume transmission' may be 'needed' in some regions. Finally, the importance of using proper control experiments for immunohistochemical studies on seven-transmembrane receptors is stressed.     

Atrial natriuretic polypeptide: topographical distribution in the rat brain by radioimmunoassay and immunohistochemistry

The widespread distribution of neurons containing alpha-atrial natriuretic polypeptide-like immunoreactivity in the rat brain was demonstrated using radioimmunoassay and immunohistochemistry in conjunction with specific antisera. The highest concentrations of alpha-atrial natriuretic polypeptide-like immunoreactivity were in the hypothalamus and septum, with low but still appreciable concentrations in the mesencephalon, cerebral cortex, olfactory bulb and thalamus by radioimmunoassay. Immunohistochemical studies clearly showed that the perikarya of immunoreactive neurons are most prevalent in the ventral part of the lateral septal nucleus, periventricular preoptic nucleus, bed nucleus of the stria terminalis, periventricular and dorsal parts of the paraventricular hypothalamic nucleus, ventromedial nucleus, dorsomedial nucleus, arcuate nucleus, median mamillary nucleus, supramamillary nucleus, zona incerta, medial habenular nucleus and the periaqueductal grey matter. Scattered neurons were seen in the cingulate cortex, endopiriform nucleus, lateral hypothalamic area, and pretectal and dorsal thalamic areas. In addition to the areas mentioned above, high concentrations of immunoreactive varicose fibers were seen in the glomerular layer of the olfactory bulb, external layer of the median eminence, central to paramedian parts of the interpeduncular nucleus and the paraventricular hypothalamic nucleus. The globus pallidus, medial and central amygdaloid nuclei, dorsal raphe, dorsal parabrachial nucleus, locus coeruleus, vagal dorsal motor nucleus, solitary nucleus and some circumventricular organs, including the subfornical organ and organum vasculosum laminae terminalis, contained considerable numbers of immunoreactive varicose fibers. In dehydrated rats and homozygous Brattleboro rats, the pattern of alpha-atrial natriuretic polypeptide-immunoreactive neurons and varicose fibers was qualitatively similar to that seen in normal conditioned rats. This study gives an atlas of the distribution of the alpha-atrial natriuretic polypeptide-containing neuronal system in the rat brain and provides the groundwork for studying the influence of this new peptide on various brain functions.     

Signaling pathways of interleukin-1 actions in the brain: anatomical distribution of phospho-ERK1/2 in the brain of rat treated systemically with interleukin-1beta

Interleukin-1beta is released at the periphery during infection and acts on the nervous system to induce fever, neuroendocrine activation, and behavioral changes. These effects are mediated by brain type I IL-1 receptors. In vitro studies have shown the ability of interleukin-1beta to activate mitogen-activated protein kinase signaling pathways including p38, c-Jun N-terminal kinase and extracellular signal-regulated protein kinase 1 and 2 (ERK1/2). In contrast to other mitogen-activated protein kinases, little is known about ERK1/2 activation in the rat brain in response to interleukin-1beta. The aim of the present study was therefore to investigate spatial and temporal activation of ERK1/2 in the rat brain after peripheral administration of interleukin-1beta using immunohistochemistry to detect the phosphorylated form of the kinase. In non-stimulated conditions, phosphorylated ERK1/2 immunoreactivity was observed in neurons throughout the brain. Administration of interleukin-1beta (60 microg/kg, i.p.) induced the phosphorylation of ERK1/2 in areas at the interface between brain and blood or cerebrospinal fluid: meninges, circumventricular organs, endothelial like cells of the blood vessels, and in brain nuclei involved in behavioral depression, fever and neuroendocrine activation: paraventricular nucleus of the hypothalamus, supraoptic nucleus, central amygdala and arcuate nucleus. Double labeling of phosphorylated ERK1/2 and cell markers revealed the expression of phosphorylated ERK1/2 in neurons, astrocytes and microglia. Since phosphorylated ERK1/2 was found in structures in which type I IL-1 receptor has already been identified as well as in structures lacking this receptor, activation of ERK1/2 is likely to occur in response to both direct and indirect action of interleukin-1beta on its target cells.     

The hippocampal continuation (indusium griseum): its connectivity in the hedgehog tenrec and its status within the hippocampal formation of higher vertebrates

The indusium griseum and its precallosal extension are usually considered poorly differentiated portions of the hippocampus. The connections of this so-called 'hippocampal continuation' (HCt) have only been analyzed so far in rodents, which show one of the least-developed HCt among mammals. In this study we have investigated the relatively well differentiated HCt of the small Madagascan hedgehog tenrec (Afrotheria) using histochemical and axonal transport techniques. The tenrec's HCt shows associative and commissural connections. It receives laminar specific afferents from the entorhinal cortex (collaterals from neurons projecting to the dentate area), the anterior and posterior piriform cortices as well as the supramammillary region. A few fibers also originate in the olfactory bulb and the dentate hilus. Among these input areas only the dentate hilus receives a significant reciprocal projection from the HCt. Additional HCt efferents are directed to the subcallosal septum (presumed septohippocampal nucleus), the olfactory tubercle and the islands of Calleja. With the exception of the supramammillary afferents and possible efferents to the supraoptic nucleus we failed, however, to demonstrate distinct thalamic and hypothalamic connections. A comparison of the connections of the HCt with those of the hippocampal subdivisions reveal some similarity between the HCt and the dentate area, but the overall pattern of connectivity does not permit a correlation of the HCt with the dentate area, let alone the cornu ammonis and the subiculum. This view is supported by histochemical findings in the tenrec (immunoreactivity to calcium binding proteins) as well as the rat (data taken from the literature). The HCt is therefore considered a region in its own right within the hippocampal formation. It may be tentatively correlated with the medial cortex of reptiles, while the dentate area and the cornu ammonis may have evolved de novo in mammals.