Comparative Distribution of Neurons Containing FLFQPQRFamide-like (morphine-modulating) Peptide and Related Neuropeptides in the Rat Brain

FLFQPQRF-NH2 (F8Famide; morphine-modulating peptide), isolated from bovine brain, is an FMRFamide-like peptide with opioid analgesia modulating effects. In the rat brain, F8Famide is immunohistochemically localized in neurons of the medial hypothalamus and medulla oblongata. Neuropeptide Y (NPY) is structurally related to F8Famide and the mammalian FMRFamide-like immunoreactivity (LI) was once thought to be due to an NPY-like peptide. We compared the anatomical distribution of F8Famide-LI with the localization of enkephalin- and NPY-LI-containing structures in the rat brain to find out if NPY or enkephalins coexist with F8Famide-LI. Cryostat sections of colchicine-treated Wistar rat brains were incubated with specific antisera against F8Famide, NPY, YGGFMRGL (Met-enkephalin-Arg-Gly-Leu), or YGGFMRF (Met-enkephalin-Arg-Phe) raised in rabbits. The immunoreactivity was visualized by the peroxidase - antiperoxidase or immunofluorescence method. The light microscopic mirror method was applied to study the colocalization of F8Famide and NPY. The F8Famide-immunoreactivity was concentrated in smaller areas of medial hypothalamus and nucleus of the solitary tract than that of enkephalins and NPY. In all brain areas, the distributions of F8Famide-, enkephalin- and NPY-immunoreactive neurons were distinct. F8Famide-, NPY- and enkephalin-LI-containing nerve terminals were seen in the nucleus of the solitary tract and in the lateral parabrachial nucleus. These results show that the neuronal systems containing F8Famide-, enkephalin- or NPY-LI are anatomically separate in all brain regions. However, there are terminal areas in which more than one type of these immunoreactivities are detected. These results have anatomical correlation with pharmacological reports, suggesting modulatory functions for these peptides on regulation of blood pressure, feeding behaviour and endocrine functions.     

Distribution of neuropeptide Y-like immunoreactivity and its relationship to FMRF-amide-like immunoreactivity in the sixth lumbar and first sacral spinal cord segments of the rat

The present study was aimed at describing the distribution of neuropeptide Y (NPY)-like immunoreactivity in the sixth lumbar (L6) and first sacral segments (S1) of the rat spinal cord, comparing this distribution to that of FMRF-amide-like immunoreactivity and determining whether NPY- and FMRF-amide-like immunoreactivities are present in the same neurons in the dorsal gray commissure (DGC) in L6 and S1 of the rat spinal cord. For distribution studies tissue from colchicine-treated animals was processed according to the peroxidase-antiperoxidase technique using anti-NPY as the primary antiserum. For co-localization studies serial 5-micron sections were processed for immunofluorescence. Adjacent sections were incubated with either anti-NPY or anti-FMRF-amide as the primary antiserum. The number of immunoreactive cells per section was counted and each section was photographed. The sections were then restained with the other antiserum (i.e., tissue first stained with anti-NPY was stained with anti-FMRF-amide and vice versa), the number of cells per section was recounted, and the sections were rephotographed. NPY-like immunoreactive cells and fibers were identified in the DGC, sacral parasympathetic nucleus, substantia gelatinosa, marginal zone, nucleus proprius, and ventral horn. Every cell in the DGC that contained NPY-like immunoreactivity was found also to contain FMRF-amide-like immunoreactivity, and the distribution of NPY-like immunoreactive fibers was found to be similar, although denser than FMRF-amide-like immunoreactive fibers. The distribution of NPY-like immunoreactivity in L6 and S1 of the rat spinal cord suggests that an NPY-like peptide may be involved in regulation of pelvic viscera, processing of primary afferent information, and motor regulation of pelvic muscles. The presence of NPY- and FMRF-amide-like immunoreactivities in the same neurons in the DGC together with the lack of bona fide FMRF-amide in the rat central nervous system, the presence of NPY in the rat central nervous system, and the cross-reactivity of anti-FMRF-amide with NPY support the hypothesis that the FMRF-amide antiserum recognizes an NPY-like peptide in the rat spinal cord.     

Androgen receptor-immunoreactivity in the forebrain of the Eastern Fence lizard (Sceloporus undulatus)

Androgen receptor (AR) distribution in the lizard forebrain and optic tectum was examined using PG21 immunohistochemistry. In the male Eastern Fence lizard, AR-immunoreactive (-ir) nuclei were observed in the medial preoptic area, ventromedial and arcuate hypothalamic nuclei, periventricular hypothalamus, premammillary nucleus, bed nucleus of the stria terminalis, and ventral posterior amygdala. Punctate immunostaining of neuronal processes (axons and/or dendrites) was concentrated in the cortex, hypothalamus, and optic tectum. AR-ir nuclei in the female brain were confined to the ventral posterior amygdala and ventromedial hypothalamic nucleus. The AR distribution in the lizard brain is similar to that reported for other vertebrate classes. Sex differences in AR-immunoreactivity may contribute to sex-specific behaviors in the Eastern Fence lizard.     

Characterization of neuropeptide Y2 receptor protein expression in the mouse brain. I. Distribution in cell bodies and nerve terminals

Neuropeptide Y (NPY), a 36-amino-acid peptide, mediates biological effects by activating Y1, Y2, Y5, and y6 receptors. NPY neurons innervate many brain regions, including the hypothalamus, where NPY is involved in regulation of a broad range of homeostatic functions. We examined, by immunohistochemistry with tyramide signal amplification, the expression of the NPY Y2 receptor (Y2R) in the mouse brain with a newly developed rabbit polyclonal antibody. Y2R immunoreactivity was specific with its absence in Y2R knockout (KO) mice and in adjacent sections following preadsorption with the immunogenic peptide (10(-5) M). Y2R-positive processes were located in many brain regions, including the olfactory bulb, some cortical areas, septum, basal forebrain, nucleus accumbens, amygdala, hippocampus, hypothalamus, substantia nigra compacta, locus coeruleus, and solitary tract nucleus. However, colchicine treatment was needed to detect Y2R-like immunoreactivity in cell bodies in many, but not all, areas. The densest distributions of cell bodies were located in the septum basal forebrain, including the bed nucleus, and amygdala, with lower density in the anterior olfactory nucleus, nucleus accumbens, caudal striatum, CA1, CA2, and CA3 hippocampal fields, preoptic nuclei lateral hypothalamus, and A13 DA cells. The widespread distribution of Y2R-positive cell bodies and fibers suggests that NPY signaling through the Y2R is common in the mouse brain. Localization of the Y2R suggests that it is mostly presynaptic, a view supported by its frequent absence in cell bodies in the normal mouse and its dramatic increase in cell bodies of colchicine-treated mice.     

Neuropeptide Y: regional distribution chromatographic characterization and immunohistochemical demonstration in post-mortem human brain

Using a neuropeptide Y (NPY)-directed radioimmunoassay the post-mortem stability of NPY was assessed in both rat and human brain samples. The regional distribution of NPY-like immunoreactivity in human brain was determined. The NPY-like immunoreactivity in human brain separated on Sephadex G-50 columns and ¹⁸C reverse phase at the position expected for NPY. Immunohistochemical staining using the NPY-directed antiserum revealed a characteristic population of cortical and striatal neurons containing NPY-like immunoreactivity.     

Mapping of neuropeptide Y-like immunoreactivity in the human forebrain

By means of immunohistochemistry the distribution of neuropeptide Y (NPY)-containing neurons and fibres was determined in the human forebrain on the basis of frontal paraffin sections from six individuals of different biological age. NPY was located in abundance in telencephalic cortical and subcortical structures like the striatum, the amygdaloid body and the substantia innominata. Variations in the distribution of immunoreactivity were observed and correlated with distinct subdivisions and structural elements of the basal forebrain region identified by Weigert or Nissl-stained neighbouring sections. The diencephalon was characterized by a relative paucity of labeled cells which were mostly confined to the area of the Nc. infundibularis and the median eminence while fibers were widely distributed in high density in most hypothalamic subnuclei except for the supraoptic nucleus. A periventricular zone of high fibre immunoreactivity was observed in the thalamus. NPY distribution in the developing brain was characterized by the finding of numerous labeled perikarya in the subcortical white matter and by far higher densities of labeled cells in the striatum as compared to the adult brain.     

Electron microscope immunocytochemical localization of neuropeptide Y (NPY) in the rat brain

In order to identify the neuronal structures as well as the intracellular organelles which contain the newly discovered brain peptide, neuropeptide Y (NPY), we have localized this peptide by immunoelectron microscopy in brain areas containing high levels of NPY. Ultrastructural studies using both the pre- and post-embedding immunoperoxidase techniques have shown that NPY-like immunoreactivity could be observed in neuronal cell bodies, dendrites and axonal processes. In terminals, the reaction product was associated with the large dense core vesicles. Only a small percentage of the positive terminals were seen making synaptic contacts. These results support the hypothesis that NPY can act as a neurotransmitter or neuromodulator.     

Neuropeptide tyrosine-like immunoreactive system in the brain,olfactory organ and retina of the zebrafish,Danio rerio,during development

The anatomical distribution of neuropeptide tyrosine (NPY)-like immunoreactivity was investigated in the brain, olfactory organ and retina of the zebrafish, Danio rerio, during development and in juvenile specimens, by using the indirect immunofluorescence and the peroxidase-antiperoxidase methods. In 60 h post fertilization (hpf) embryos, NPY-like immunoreactive cell bodies appeared in the hypothalamus, within the posterior periventricular nucleus. Few positive nerve fibers were found in the hypothalamus and in the tegmentum of the mesencephalon. In 72 hpf embryos, a new group of NPY-like immunoreactive cells was found in the olfactory pit. At day 4 of development, NPY-like immunoreactive cell bodies were detected between the olfactory pit and the olfactory organ. In the hypothalamus the location of positive cell bodies was similar to that reported in the previous developmental stages. A few positive nerve fibers appeared in the tegmentum of the rhombencephalon. At days 7 and 15 of development, the distribution of NPY-like immunoreactivity was very similar to that reported at day 4. However, at day 15, NPY-like immunoreactivity appeared for the first time in amacrine cells of the retina and in nerve fibers of the tectum of the mesencephalon. In 1-month/3-month-old animals, additional groups of NPY-like immunoreactive cell bodies appeared in the glomerular layer of the olfactory bulbs, the terminal nerve, the lateral nucleus of the ventral telencephalic area, the entopeduncular nucleus and in the medial region of the reticular formation of the rhombencephalon. These results show that NPY-like immunoreactive structures appear early during ontogeny of zebrafish. The distribution of the immunoreactive system increases during the ontogeny, the juvenile stages, and reaches the complete development in mature animals. The location of NPY-like immunoreactivity indicates that, during development, NPY could be involved in several neuromodulatory functions, including the processing of visual and olfactory information. In 1-month/3-month-old animals, NPY-like immunoreactive nerve fibers are present in the pituitary, suggesting that, from these stages onward, NPY may influence the secretion of pituitary hormones.     

Distribution of neuropeptide Y-like immunoreactivity in the hypothalamus of the adult golden hamster

The distribution of neuropeptide Y (NPY)-like immunoreactivity within the hypothalamus of the adult golden hamster was investigated with conventional immunohistochemical techniques. Neuropeptide Y immunoreactive cell bodies were found in greatest numbers in the arcuate nucleus while a few stained perikarya were seen in the internal and subependymal zones of the median eminence. Isolated perikarya were observed in the anterior commissure and supracommissural portion of the interstitial nucleus of the stria terminalis. Immunoreactive axons were located throughout the hypothalamus with the highest concentrations in the subependymal and internal zones of the median eminence, the interstitial nucleus of the stria terminalis, the medial preoptic area, and in the following nuclei: periventricular, suprachiasmatic, paraventricular, perifornical, median preoptic, and arcuate. Moderate to dense plexuses of immunoreactive fibers were observed in the anterior, lateral, and posterior hypothalamic areas and in the infundibular stalk. The supraoptic nucleus and lateral preoptic area displayed a small number of labeled axons whereas the ventromedial nucleus contained only a few fibers. NPY immunoreactive fibers were present in the optic tract and in the dorsomedial aspect of the optic chiasm. Labeled fibers penetrated the ependymal lining of the third ventricle throughout the ventral aspect of the periventricular zone. Additional fibers were observed in the pia lining the ventral aspect of the hypothalamus. This systematic analysis of hypothalamic NPY immunoreactivity in the adult golden hamster suggests that a portion of the labeled fibers display a distribution that is similar to previously described noradrenergic fibers in the hypothalamus.     

Immunohistochemical localization of a melanin concentrating hormone-like peptide in the rat brain

Using antisera generated in rabbits against salmon melanin concentrating hormone (MCH) coupled to human thyroglobulin, the distribution of MCH-like immunoreactivity was mapped throughout the rat central nervous system. The distribution of MCH-like immunoreactivity in rat brain is unique and different from the distribution of other neuropeptides. MCH-like immunoreactive perikarya and fibers are predominant in the posterior hypothalamic area, mostly in the medial forebrain bundle-lateral hypothalamic area subzona incerta and the perifornical area. Cell bodies are located mainly in the medial forebrain bundle and in proximity to well defined hypothalamic nuclei. Fibers are seen throughout the rat brain in all neocortical areas, the neostriatum and the amygdala, in the diencephalon in most hypothalamic nuclei, the habenula, the mamillary body and very dense in the medial forebrain bundle and just ventral to the zona incerta ("subzona incerta"). In the mesencephalon there are fibers in the central gray; in the pons-medulla fibers are contained in the dorsal and ventral parabrachial nuclei; in the tegmental area ventral to the fourth ventricle; in the spinal trigeminal area, the substantia gelatinosa and the reticular nuclei. In the spinal cord there are more fibers in the dorsal than in the ventral horn. The posterior pituitary also contained few MCH-like fibers. It is suggested that a peptide similar, but not identical, to salmon MCH is present in the rat central nervous system.     

Increases in plasma neuropeptide Y concentrations during sympathetic activation in man

Neuropeptide Y (NPY) coexists with noradrenaline in postganglionic sympathetic neurons. In order to test the hypothesis that NPY may be released along with catecholamines by activation of the sympathoadrenal system we measured plasma NPY-like immunoreactivity (NPY-LI) concentrations during cold pressor test, head up tilt and bicycle exercise in healthy volunteers. All 3 manoeuvres resulted in elevation of blood pressure, heart rate and plasma noradrenaline and adrenaline concentrations. These were accompanied by increases in plasma NPY-LI concentrations on cold pressor test and exercise, but not with head up tilt. The increases in both NPY-LI and catecholamines were greatest with exercise. These findings suggest that NPY is released at the same time as noradrenaline when sympathetic noradrenergic nerves are activated.     

Neuropeptide Y localization in the rat amygdaloid complex

Neuropeptide Y (NPY)-, avian pancreatic polypeptide (APP)-, and molluscan cardioexcitatory peptide (FMRF)-like immunoreactivity in the amygdaloid complex of the rat was investigated immunohistochemically. The distribution of each of these peptides within the amygdala is identical and cross-blocking studies indicate that all three antisera recognize the NPY antigen. Morphologically distinct populations of NPY immunoreactive neurons are differentially distributed in the medial amygdaloid nucleus and at the base of the stria terminalis. Dense plexuses of immunoreactive axons are present in the medial third of the central nucleus and in the dorsal half of the medial nucleus, with light to moderate fiber plexuses present in the lateral and basolateral nuclei and scattered axons present throughout the remainder of the amygdala. The distribution and appearance of NPY immunoreactive plexuses in the amygdala is similar to that described previously for noradrenergic axons arising from brainstem cell groups (Fallon, Koziell, and Moore: J. Comp. Neurol. 180:509-532, '78). However, injections of the noradrenergic neurotoxin 6-hydroxydopamine into the amygdala result in a complete loss of dopamine-beta-hydroxylase (DBH) immunoreactivity in the amygdala and surrounding cortex but leave much of the NPY immunoreactive plexus intact. Similarly, lesions of the locus coeruleus deplete DBH immunoreactivity, leaving NPY-like immunoreactivity in the amygdala unaffected. These results indicate that much of the NPY immunoreactive plexus observed in the amygdala does not arise from brainstem sources in which NPY and noradrenaline are colocalized. Lesions of the stria terminalis or medial nucleus have no observable effect on the density or distribution of NPY immunoreactive terminal fields in the basal forebrain and hypothalamus, suggesting that immunoreactive neurons in the amygdaloid complex do not contribute significantly to this innervation.     

Sex-specific distribution of Neuropeptide Y (NPY) in the brain of the frog,Microhyla ornata

Neuropeptide Y (NPY) is involved in sex-specific behavioural processes in vertebrates. NPY integrates energy balance and reproduction in mammals. However, the relevance of NPY in reproduction of lower vertebrates is understudied. In the present study, we have investigated neuroanatomical distribution and sex-specific differences of NPY in the brain of Microhyla ornata using immunohistochemistry and quantitative real time PCR. NPY is widely distributed throughout the brain of M. ornata. We observed NPY immunoreactivity in the cells of the nucleus accumbens, striatum pars dorsalis, dorsal pallium, medial pallium, ventral pallium, bed nucleus of stria terminalis, preoptic nucleus, infundibular region, median eminence and pituitary gland of adult M. ornata. A higher number of NPY- immunoreactive cells were observed in the preoptic nucleus (p < .01), nucleus infundibularis ventralis (p < .001) and anteroventral tegmental nucleus (p < .001) of the female as compared to that of the male frog. Real-Time PCR revealed higher mRNA levels of NPY in the female as compared to male frogs in the mid-brain region that largely contains the hypothalamus. Sexual dimorphism of NPY expression in M. ornata suggests that NPY may be involved in the reproductive physiology of anurans.     

Neuropeptide Y in male and female brains of Flinders Sensitive Line, a rat model of depression. Effects of electroconvulsive stimuli

Human and animal studies suggest that neuropeptide Y (NPY), a peptide co-localized and co-released with classical neurotransmitters, is involved in the pathogenesis of affective disorders. In addition, lithium, electroconvulsive treatments (ECT in humans and ECS in rodents) and antidepressants affect NPY in a specific temporal- and brain-region fashion. These results have been obtained on healthy male rats; females and/or “depressed” animals have essentially not been studied. Consequently, we studied brain NPY-like immunoreactivity (-LI) under basal conditions and following a series of ECS in both male and female Flinders Sensitive Line (FSL), an animal model of depression, and their controls, the Flinders Resistant Line (FRL) rats. Furthermore, we examined whether the oestrus cycle affects NPY-LI in these strains. Following sacrifice by focused microwave irradiation, the peptides were extracted from dissected brain regions and measured by radioimmunoassay. Hippocampal NPY-LI in both sexes was significantly lower in the “depressed” FSL compared to the control FRL. ECS increased NPY-LI in both male and female rats in both FSL and FRL strains in hippocampus, frontal cortex and occipital cortex. In the hypothalamus, the increase was found only in the FSL rats. In both FSL and control rats, the basal NPY-LI was lower in the hippocampus of female compared to male rats. NPY-LI did not vary during the different phases of the oestrus cycle. These results suggest that the gender differences are not due to NPY-LI variations during the oestrus. The results are consistent with our hypothesis that NPY plays a role in the pathophysiology of depressive disorders and provide further evidence that one of the modes of ECS action is to elevate NPY in the limbic system. Assumption that gender differences in NPY could explain increased rates of depression in women is speculative, but is in line with the findings in the present study.     

Afferent connections of the habenular complex in the lizard Gallotia galloti.

Afferents to the habenular complex were studied by means of in vitro horseradish peroxidase retrograde labeling and anterograde control experiments in the lizard Gallotia galloti. The medial habenular nucleus was found to receive abundant afferent fibers from the nucleus of the posterior pallial commissure and the nucleus septalis impar. More restricted input comes from the nucleus eminentiae thalami and the nucleus of the stria medullaris. The lateral habenular nucleus is innervated by various fiber groups originating from the bed nucleus of the anterior commissure, the diagonal band nucleus, the lateral preoptic area, the anterior entopeduncular nucleus, the lateral hypothalamic and mammillary areas, the nucleus of the stria medullaris, the area tegmentalis ventralis and a scattered neuronal subpopulation in the large-celled dorsolateral nucleus of the dorsal thalamus. Habenulopetal fibers generally follow the stria medullaris, but hypothalamic, entopeduncular and dorsal thalamic afferents course through the dorsal peduncle of the lateral forebrain bundle in a transthalamic route. Mesencephalic ventral tegmental afferents ascend through the tractus retroflexus.     

Immunohistochemical mapping of neuropeptide Y in the tree shrew brain

Day‐active tree shrews are promising animals as research models for a variety of human disorders. Neuropeptide Y (NPY) modulates many behaviors in vertebrates. Here we examined the distribution of NPY in the brain of tree shrews (Tupaia belangeri chinensis) using immunohistochemical techniques. The differential distribution of NPY‐immunoreactive (‐ir) cells and fibers were observed in the rhinencephalon, telencephalon, diencephalon, mesencephalon, metencephalon, and myelencephalon of tree shrews. Most NPY‐ir cells were multipolar or bipolar in shape with triangular, fusiform, and/or globular perikarya. The densest cluster of NPY‐ir cells were found in the mitral cell layer of the main olfactory bulb (MOB), arcuate nucleus of the hypothalamus, and pretectal nucleus of the thalamus. The MOB presented a unique pattern of NPY immunoreactivity. Laminar distribution of NPY‐ir cells was observed in the MOB, neocortex, and hippocampus. Compared to rats, the tree shrews exhibited a particularly robust and widespread distribution of NPY‐ir cells in the MOB, bed nucleus of the stria terminalis, and amygdala as well as the ventral lateral geniculate nucleus and pretectal nucleus of the thalamus. By contrast, a low density of neurons were scattered in the striatum, neocortex, polymorph cell layer of the dentate gyrus, superior colliculus, inferior colliculus, and dorsal tegmental nucleus. These findings provide the first detailed mapping of NPY immunoreactivity in the tree shrew brain and demonstrate species differences in the distribution of this neuropeptide, providing an anatomical basis for the participation of the NPY system in the regulation of numerous physiological and behavioral processes. J. Comp. Neurol. 523:495–529, 2015. © 2014 Wiley Periodicals, Inc.