Menek Goldstein

Differential co-existence of neuropeptide Y (NPY)-like immunoreactivity with catecholamines in the central nervous system of the rat

The distribution of neuropeptide Y immunoreactive cell bodies in relation to various types of catecholamine-containing cell bodies in the rat brain was analyzed immunohistochemically using antisera to tyrosine hydroxylase, dopamine beta-hydroxylase and phenylethanolamine N-methyltransferase. Coexistence of the peptide in catecholamine cell bodies was established by using an elution-restaining procedure. Neuropeptide Y-like immunoreactivity was observed in most noradrenergic cell bodies of the Al/Cl cell groups in the ventro lateral medulla oblongata. Similarly this peptide immunoreactivity was also observed in the majority of the adrenergic cell bodies of the C2 group. In the dorsal and dorsal-lateral part of the nucleus of the solitary tract, where a group of small adrenergic cells is present, several small neuropeptide Y immunoreactive cells were also observed. The possibility of coexistence of adrenaline and neuropeptide Y in these cells remains to be established. The majority of the noradrenergic cell bodies of the A2 group, as well as the presumptive dopaminergic cells within its ventromedial part, seemed to lack neuropeptide Y-like immunoreactivity. Many noradrenergic cell bodies of the A6 group in the locus coeruleus proper were neuropeptide Y-immunoreactive, whereas the peptide could not be observed in the subcoeruleus group. Neither the A5 and A7 noradrenergic cells in the pons, nor any of the dopaminergic cell groups in the mesencephalon and forebrain (A8-A15) seemed to contain a neuropeptide Y-like peptide. The findings indicate that central catecholamine neurons can be subdivided into distinct sub-groups based upon the coexistence of a specific peptide.

High levels of neuropeptide Y in peripheral noradrenergic neurons in various mammals including man

Using a highly specific radioimmunoassay for neuropeptide Y (NPY), levels in the peripheral nervous system of guinea-pig, cat, pig and man were measured. In all species very high levels (up to 800 pmol X g-1) were found in sympathetic ganglia and in tissues which receive a dense sympathetic innervation, such as vas deferens, heart atrium, blood vessels and spleen. By immunocytochemistry, NPY-immunoreactive (-IR) principal ganglion cells in sympathetic ganglia and the pelvic plexus were also found to contain dopamine-beta-hydroxylase (DBH) and tyrosine hydroxylase (TH), strongly suggesting that the NPY-IR cells are noradrenergic. NPY- and DBH-IR nerves had a roughly parallel occurrence in the heart, spleen, kidney, respiratory and urogenitary tracts, around blood vessels and within visceral smooth muscle. Considerably more NPY-IR than DBH-IR nerve fibres were seen in the gastrointestinal tract from the oesophagus to the anal sphincter. In addition, NPY-IR local ganglion cells were observed in the submucous and myentric plexuses. NPY-like immunoreactivity was also observed in the adrenal medulla of guinea-pig and cat. NPY thus seems to be a major peptide in the sympathetic nervous system, supporting its proposed role in sympathetic neurotransmission.

Enkephalin-like immunoreactivity in gland cells and nerve terminals of the adrenal medulla.

Abstract Met-enkephalin-like immunoreactivity has been observed in adrenal medullary gland cells of the rat, guinea-pig and cat. There were marked quantitative differences in untreated animals. Most medullary gland cells were positive in the cat, a large proportion in the guinea-pig but only a few in the rat. After sectioning of the splanchnic nerve, however, a large proportion of the gland cells were met-enkephalin positive also in the rat. Occasionally the met-enkephalin-like immunoreactivity had a granular appearance suggesting that the storage sites could be vesicular. In the guinea-pig a moderate number of met-enkephalin immunoreactive nerve fibers were observed in the adrenal medulla. They disappeared to a large extent after sectioning of the splanchnic nerve. A few fibers were also seen in the rat adrenal medulla, and in the cat some fibers could be seen in areas with few immunoreactive cells. The present findings suggest that in the adrenal medulla an enkephalin-like peptide(s) is present both in gland cells and in nerve terminals arising mainly from fibers in the splanchnic nerve. Thus, the possibility exists that in the adrenal medulla opioid peptides may be released from the gland cells into the blood as hormones as well as from nerve terminals to act as a modulator or transmitter. It should, however, be emphasized that the well-documented metabolic instability of met-enkephalin may be somewhat difficult to reconcile with a hormonal role of this peptide.

Coexistence of galanin-like immunoreactivity with catecholamines, 5- hydroxytryptamine, GABA and neuropeptides in the rat CNS

The coexistence of galanin (GAL)-like immunoreactivity (LI) with markers for catecholamines, 5-hydroxytryptamine (5-HT), GABA, or some neuropeptides was mapped in the rat CNS by using adjacent sections, as well as by elution-restaining and double-labeling immunocytochemistry. Many instances of coexistence were observed, but there were also numerous GAL-positive cell body populations displaying distributions similar to those of these markers but without apparent coexistence. In the hypothalamic arcuate nucleus GAL-LI was found in a large proportion of tyrosine hydroxylase (TH)-positive cell bodies (A12 cells), both in the dorsomedial and ventrolateral subdivisions, with a higher number in the latter. GAL-LI coexisted in glutamic acid decarboxylase (GAD)- positive somata in the posterior aspects of the arcuate nucleus and at all rostrocaudal levels in fibers in the external layer of the median eminence. In the anterior hypothalamus, a large population of the cells of the parvocellular and magnocellular paraventricular nuclei contained both GAL-LI and vasopressin-LI. Moreover, somata containing both GAD- and GAL-LI were seen lateral to the mammillary recess in the tuberal and caudal magnocellular nuclei. Some of the neurons of the caudal group were shown to project to the occipital cortex using combined retrograde tracing and immunofluorescence. With regard to mesencephalic and medullary catecholamine neurons, GAL-LI coexisted in a large proportion of the noradrenergic locus coeruleus somata (A6 cell group) and in the A4 group dorsolateral to the fourth ventricle, as well as in the caudal parts of the A2 group in the dorsal vagal complex. However, in more rostral parts of the latter, especially in the medial subdivision of the solitary tract nucleus, a very large population of GAL-IR small cell bodies was seen intermingling with catecholamine neurons, but they did not contain TH-LI. Furthermore, GAL-IR cell bodies coextensive with, but not coexisting in, TH-IR somata were seen in the C1 (epinephrine) horea in the ventrolateral medulla at the level of area postrema and in the most rostral aspects of the C1 group. Finally, 5-HT-positive cell bodies of the mesencephalic and medullary raphe nuclei and a subpopulation of coarse 5-HT nerve fibers in the hippocampus co-contained GAL-LI. The present results demonstrate that a GAL-like peptide is present in many systems containing other neuroactive compounds, including dopamine, norepinephrine, 5-HT, GABA, and vasopressin.(ABSTRACT TRUNCATED AT 400 WORDS)

Locus coeruleus neurons in the rat containing neuropeptide Y, tyrosine hydroxylase or galanin and their efferent projections to the spinal cord, cerebral cortex and hypothalamus.

The efferent projections of locus coeruleus neurons which contain neuropeptide Y-, tyrosine hydroxylase- or galanin-like immunoreactivity were investigated using the indirect immunofluorescence technique combined with the retrograde transport of the fluorescent substance Fast Blue. Four groups of rats received injections of Fast Blue: (1) bilaterally into the mid-thoracic spinal cord (T6-T7); (2) unilaterally into the low cervical spinal cord (C4-C5); (3) unilaterally into the paraventricular, periventricular and dorsomedial hypothalamic nuclei; and (4) unilaterally into five sites in the cerebral cortex (frontal, cingulate and striate cortex). Efferent projections to the spinal cord, hypothalamus and cerebral cortex from neuropeptide Y-, tyrosine hydroxylase- and galanin-containing locus coeruleus cells were observed. A higher percentage of the peptidergic locus coeruleus neurons projected to the hypothalamus than to the spinal cord or cerebral cortex. The distribution and morphology of the neuropeptide Y- and galanin-containing neurons in the locus coeruleus were also investigated. Neuropeptide Y-like immunoreactivity and galanin-like immunoreactivity were found in small, medium and large multipolar neurons, as well as in fusiform locus coeruleus cells. The neuropeptide Y- and galanin-immunoreactive neurons were found throughout the locus coeruleus. In the caudal locus coeruleus, they were primarily located in the dorsal portion. Neuropeptide Y-like immunoreactivity and galanin-like immunoreactivity were only seen in a few tyrosine hydroxylase-positive neurons of the subcoeruleus group. The data show that the peptide-containing locus coeruleus neurons have efferent projections to the spinal cord, hypothalamus and cerebral cortex. The locus coeruleus may be divided into functional subdivisions dependent on the region of the locus coeruleus, the neurotransmitter/neuropeptide(s) contained within the neurons and their efferent projections.

Peptide-monoamine coexistence: studies of the actions of cholecystokinin-like peptide on the electrical activity of midbrain dopamine neurons.

Abstract Recent studies have shown that a cholecystokinin-like peptide coexists in a subpopulation of midbrain dopamine-containing neurons. Using extracellular single unit recording techniques we have found that this peptide increases the activity of some, but not all, dopaminergic neurons (identified on the basis of their electrophysiological features). The responsive neurons were found exclusively in areas which were subsequently shown, using immunocytochemical techniques, to contain both cholecystokinin and the enzyme marker for dopaminergic neurons, tyrosine hydroxylase. When administered intravenously, cholecystokinin increased the firing rate of dopaminergic neurons lying in cholecystokinin-rich areas of the substantia nigra. Cells in cholecystokinin-rich ventral tegmental areas showed more variable responses to comparable doses of the peptide. Iontophoretically-applied cholecystokinin consistently activated dopaminergic cells in the substantia nigra and ventral tegmental area, increasing their firing rate and their bursting activity. Cholecystokinin increased the firing rate of some of those neurons to the extent that they were driven into apparent depolarization inactivation. Furthermore, iontophoresis of cholecystokinin resulted in an activation of a population of normally quiescent dopaminergic cells. These results are discussed in light of a possible functional role of cholecystokinin-like peptides in the brain dopaminergic systems. It is suggested that cells in the dopamine-rich areas of the mesencephalon can be characterized both on the basis of their content of peptide and/or catecholamine and of their responsiveness to cholecystokinin-like peptides.

Biochemical Evidence of Dysfunction of Brain Neurotransmitters in the Lesch-Nyhan Syndrome

Different brain regions were removed post mortem from three patients with the Lesch-Nyhan syndrome and were examined for alterations in hypoxanthine-guanine phosphoribosyl transferase (HGPRT), adenine phosphoribosyl transferase, and biochemical indexes of norepinephrine, dopamine, serotonin, gamma-aminobutyric acid (GABA), and acetylcholine neuron function, as compared with age-matched controls. The level of HGPRT activity in the material from patients with the Lesch-Nyhan syndrome was less than 1 per cent of control levels, whereas adenyl phosphoribosyl transferase was not significantly altered. All biochemical aspects of the function of dopamine-neuron terminals in the striatum (except dihydroxyphenylacetic acid levels) were decreased to 10 to 30 per cent of the control values. Serotonin and 5-hydroxyindoleacetic acid levels were increased, striatal choline acetyltransferase levels were low, and striatal glutamic acid decarboxylase and guanylate cyclase activities were unaltered. The disruption of the balance between the functions of GABA, dopamine, and acetylcholine neurons in the extrapyramidal system probably accounts for some of the symptoms observed in the Lesch-Nyhan syndrome (e.g., choreoathetosis).

The hypothalamic arcuate nucleus-median eminence complex: Immunohistochemistry of transmitters, peptides and DARPP-32 with special reference to coexistence in dopamine neurons

In this paper, we describe the results of a series of experiments which have examined the distribution within the arcuate nucleus of the hypothalamus of neurons containing the following immunoreactivities: TH-LI, GAD-LI, NT-LI, GAL-LI, GRF-LI, Met-ENK-LI, Leu-ENK-LI, Met-ENK-7-LI, Met-ENK-8-LI, metorphamide-LI, DYN-LI, NPY-LI, SOM-LI, FMRFamide-LI, and CLIP-LI and ependymal tanycytes containing DARPP-32-LI. Using elution-restaining and double antibody staining techniques we have established numerous patterns of coexistence of these various neurotransmitters and neuropeptides. Thus, neurons containing TH-LI were, in some instances, also found to contain GAD-LI, NT-LI, GAL-LI, GRF-LI, Met-ENK-8-LI, Leu-ENK-LI, or DYN-LI or combinations of these compounds. For example, some TH-IR neurons also contained GAL-LI and GRF-LI, while other TH-IR. neurons were also seen to contain GRF- and NT-LI. These neurons may, in fact, contain even more compounds. NPY-IR neurons and those containing SOM-LI and CLIP-LI were distinct and separate from those containing TH-LI. The distribution of these different neurochemical types of neurons and their patterns of coexistence are summarized in Fig. 34, while the relative distribution patterns of immunoreactive fibres in the median eminence are summarized in Fig. 35.

Chapter 4 Coexistence of neuronal messengers — an overview

Publisher Summary This chapter discusses results demonstrating that neurons often contain more than one chemical compound. The different types of coexistence situations are described, including (1) a classical transmitter and one or more peptides, (2) more than one classical transmitter, and (3) a classical transmitter, a peptide, and adenosine triphosphate (ATP). The functional significance of these histochemical findings is at present difficult to evaluate, but in studies on the peripheral nervous system evidence has been obtained that classical transmitter and peptide are coreleased and interact in a cooperative way on effector cells. In addition to enhancement, there is evidence that other types of interactions may occur—for example, the peptide may inhibit the release of the classical transmitter. Also in the central nervous system (CNS), indirect evidence is present for similar mechanisms—that is, to strengthen transmission at synaptic (or non-synaptic) sites and for the peptide inhibition of release of a classical transmitter. Multiple messengers may provide the means for increasing the capacity for information transfer in the nervous system.

Human brainstem catecholamine neuronal anatomy as indicated by immunocytochemistry with antibodies to tyrosine hydroxylase.

Immunocytochemistry based on antibodies to tyrosine hydroxylase is used to identify catecholaminergic neurons in the human brain stem. An atlas is provided and the distribution of structures compared with that in other animals and with biochemical and catecholamine fluorescent data from humans. Broad agreement of results increases the confidence with which tyrosine hydroxylase-like immunoreactivity can be used to trace catecholaminergic pathways in human postmortem material. As compared to most studies of other animals there are striking increases in populations of upper pontine and mesencephalic catecholaminergic neurons in the human. Distinct cytoarchitectonic features, consistent differences in tyrosine hydroxylase immunoreactive staining intensity and regional variations in substance P innervation indicate complexity within the substantia nigra. Human catecholaminergic neurons are prominent in the midline of the ventral tegmentum and the upper parts of the central tegmental tracts. A bundle of tyrosine hydroxylase-immunoreactive axons runs between the latter regions and a cluster of smaller catecholaminergic neurons which lie in the oblique band of axons joining ventrolateral and dorsomedial medullary catecholaminergic groups. There are more catecholaminergic neurons within and closely related to the superior cerebellar peduncles than have been described in other species. Anatomically, the central compact nucleus of the locus coeruleus appears to be related to several nearby catecholaminergic cell groups. The data provided are being used as a basis for neuropathologic studies of human neurological diseases.