Latent varicella-zoster viral DNA in human trigeminal and thoracic ganglia.

BACKGROUND: Some human herpesviruses become latent in dorsal-root ganglia. Primary infection with the varicella-zoster virus causes chickenpox, followed by latency, and subsequent reactivation leading to shingles (zoster), but the frequency and distribution of latent virus have not been established. METHODS: Using the polymerase chain reaction, we performed postmortem examinations of trigeminal and thoracic ganglia of 23 subjects 33 to 88 years old who had not recently had chickenpox or shingles to identify the presence of latent varicella-zoster viral DNA. Oligonucleotide primers representing the origin of replication of the varicella-zoster virus and varicella-zoster virus gene 29 were used for amplification. RESULTS: Among the 22 subjects seropositive for the antibody to the virus, both the viral origin-of-replication and gene-29 sequences were detected in 13 of 15 subjects (87 percent) in whom trigeminal ganglia were examined and in 9 of 17 (53 percent) in whom thoracic ganglia were examined. Viral DNA was not detected in brain or mononuclear cells from the seropositive subjects. None of three thoracic ganglia from the one seronegative subject contained varicella-zoster viral DNA. CONCLUSIONS: These findings indicate that after primary infection with varicella-zoster virus (varicella), the virus becomes latent in many ganglia--more often in the trigeminal ganglia than in any thoracic ganglion--and that more than one region of the viral genome is present during latency.

Galanin is co-localized with noradrenaline and neuropeptide Y in dog pancreas and celiac ganglion.

To visualize the localization and potential co-localization of noradrenaline and the putative pancreatic sympathetic neurotransmitters, galanin and neuropeptide Y (NPY), immunofluorescent staining for galanin, NPY and tyrosine hydroxylase (TH) was performed on sections of canine pancreas and celiac ganglion. In the pancreas, galanin-immuno-fluorescent nerve fibers were confirmed as densely and preferentially innervating the islets, whereas numerous NPY-positive nerve fibers were found in the exocrine parenchyma, the surrounding of the blood vessels and within the islets. Double-staining for the peptides and TH indicated that most galanin-positive nerve fibers were adrenergic, most NPY-positive nerve fibers were adrenergic, and many islet nerves contained both galanin and NPY, although some galanin-positive nerve fibers appeared to lack NPY. In the celiac ganglion, virtually all cell bodies were positive for both galanin and TH; a large subpopulation of these cells were also positive for NPY. Radioimmunoassay (RIA) of galanin in extracts of dog celiac ganglion revealed a very high content (256 +/- 33 pmol/g wet weight) of galanin-like immunoreactivity (GLIR), consistent with the dense staining observed. This GLIR behaved in a similar manner to synthetic porcine galanin in the RIA. In addition, the majority of the GLIR in ganglion extracts co-eluted with the synthetic peptide upon gel filtration, although a minor peak of a larger apparent molecular weight was also observed, observations consistent with the presence of a precursor peptide. These findings suggest that galanin is a sympathetic post-ganglionic neurotransmitter in the canine endocrine pancreas and that NPY might serve a similar function.

Localization of sulfated glucuronyl glycolipids in human dorsal root and sympathetic ganglia.

Sulfated glucuronyl glycolipids (SGGLs) in human dorsal root ganglion (DRG) and sympathetic ganglion (SG) were analyzed biochemically and immunohistochemically. SGGLs were enriched in human DRG (1.02 +/- 0.23 micrograms/mg protein), whereas much lower concentrations of these glycolipids (0.043 +/- 0.23 micrograms/mg protein) were detected in SG. Myelin within DRG and SG was immunostained by anti-SGGL antiserum, although only a few myelinated fibers were seen in SG. Nerve cell bodies or unmyelinated fibers were not immunostained. Subcellular fractionation study of human DRG demonstrated that these glycolipids were not only enriched in myelin but also in the axolemma-enriched fraction. These data are consistent with the view that SGGLs may be expressed on myelinated fibers in myelin and axolemma, suggesting that these compounds may play an important role in regulating myelinogenesis.

Localization of neuropeptide Y in human sympathetic ganglia: correlation with met-enkephalin, tyrosine hydroxylase and acetylcholinesterase.

The relationships of immunoreactive neuropeptide Y, enkephalin and tyrosine hydroxylase, on the one hand, and acetylcholinesterase histochemical activity, on the other, were studied in human lumbar sympathetic ganglia. Two thirds of the ganglion cells contained immunoreactive neuropeptide Y. Electron microscopically the immunoreaction was localized in the Golgi apparatus and in large dense-cored vesicles in the nerve endings. Most of the neuropeptide-containing neurons and nerve fibres were also reactive for tyrosine hydroxylase. Nerve fibres reactive for neuropeptide Y were found around ganglion cells regardless of their transmitter contents, whereas enkephalin-reactive nerve terminals surrounded only acetylcholinesterase-containing neurons. The results demonstrate that neuropeptide Y is colocalized with noradrenaline in most of the human sympathetic neurons and that the nerve fibres may innervate selectively the noradrenergic and cholinergic subpopulations of ganglion cells depending on the transmitters of the nerves.

Ultrastructural and cytochemical studies of nucleolus-like bodies in neurons of rat sympathetic ganglia.

Intracytoplasmic fibrillar inclusions, generally referred to as nucleolus-like bodies (NLBs) were studied by means of ultrastructural cytochemistry. The structure of these bodies was visualized by several different staining procedures: conventional electron microscopy and preferential staining methods for localization of various proteins including ribonucleoproteins, basic proteins, glycoproteins and phosphorylated proteins. The results of the cytochemical tests indicate that NLBs have an essentially proteinaceous nature. They consist of ribonucleoproteins, basic proteins and glycoproteins but do not contain phosphorylated proteins. These findings suggest that NLBs are, at least partially, of the same nature as nucleoli and coiled bodies. The origin of NLBs and their possible functional role is briefly discussed.

Immunohistochemical localization of Met5-enkephalin-Arg6-Gly7-Leu8 in the female genital organs and in the paracervical ganglion of the pig.

Indirect immunofluorescence method was used to study the localization and distribution of the proenkephalin A-derived octapeptide, Met5-enkephalin-Arg6-Gly7-Leu8 (MEAGL), in the paracervical ganglion and in the female genital organs of the pig. In the paracervical ganglion, a subpopulation of principal neurons and nerve fibers contained MEAGL immunoreactivity. In the vagina, numerous MEAGL-immunoreactive nerve fibers were localized in the muscular membrane, under the serous membrane and in the submucous layer. The uterine cervix contained a great number of immunoreactive nerve fibers in muscular membrane and in submucous and subserous layers. The pattern of distribution of MEAGL-immunoreactive nerve fibers in the uterine horns was similar to that of cervix, but their number in the uterine horns was lower. MEAGL-immunoreactive fibers were also observed through different oviductal layers. In the ovary a low number of immunoreactive fibers were seen in the medullary and cortical parts of the organ. The results of this study indicate that the female genital organs, particularly the uterus and vagina, of the pig receive dense innervation by nerve fibers containing the proenkephalin A-derived octapeptide MEAGL. The presence of MEAGL in principal neurons and fibers of the paracervical ganglion suggests that a large proportion of them originate from neurons of the paracervical ganglion.

Neuronal pathways in the guinea-pig lumbar sympathetic ganglia as revealed by immunohistochemistry.

Tyrosine hydroxylase (TH)- and peptide-immunoreactivity of postganglionic neurons and of nerve fibres in guinea pig lumbar paravertebral sympathetic ganglia 2-4 after transection of the communicating rami and the visceral branches, respectively, were investigated by single- and double-labelling techniques. Six subpopulations of postganglionic neurons were discriminated immunohistochemically: two cell types, which were immunoreactive to only one of the applied antisera - TH, and vasoactive intestinal polypeptide (VIP); and four cell types in which immunoreactivity was colocalized - TH/neuropeptide Y (NPY), NPY/VIP, dynorphin/alpha-neoendorphin and dynorphin (alpha-neoendorphin)/NPY. Small intensely fluorescent (SIF) cells dependent on their location exhibited differential immunobehaviour to NPY-/dynorphin-(alpha-neoendorphin-) and TH-antisera. Immunoreactivity to substance P (SP), calcitonin gene-related peptide (CGRP), met-enkephalin-arg-phe (MEAP) and leu-enkephalin was present in nerve fibres but not in postganglionic neurons with frequent colocalization of SP/CGRP- and MEAP/leu-enkephalin- and, sometimes leu-enkephalin/SP- and dynorphin/SP-immunoreactivity. TH-immunoreactive intraganglionic nerve fibres were numerically more increased after cutting the visceral branches, than after transection of the communicating rami. Vice versa, NPY-, VIP-, dynorphin- and alpha-neoendorphin-immunoreactive nerve fibres were particularly increased in number after cutting the communicating rami. Many but not all of the nerve fibres exhibited colocalization of two of these peptides. SP-, CGRP-, and enkephalin-immunoreactive nerve fibres were not visibly affected by cutting the visceral branches but virtually disappeared after lesioning the communicating rami.

Isobestic wavelength determination for succinic dehydrogenase-nitroblue tetrazolium-derived formazans in autonomic neurons.

Two methods have been used to determine the isobestic (equiconcentration) wavelength of formazans derived from nitroblue tetrazolium-succinate dehydrogenase (EC 1.3.99.1) activity in cryostat sections of the rat superior cervical ganglion prior to microdensitometric measurements. Both methods indicate that maximal absorbance of the final reaction product is at a wavelength of 540-550 nm. This wavelength differs by 35-45 nm from that used to measure the same reaction product in sections of other rat tissues such as liver and implies that it may be unwise to adopt a "standard" wavelength for a particular reaction product when making microdensitometric measurements in relation to quantitative enzyme histochemistry.

Projections of neurons with neuromedin U-like immunoreactivity in the small intestine of the guinea-pig.

Neuromedin U immunoreactivity was located histochemically in the guinea-pig small intestine. Projections of immunoreactive neurons were determined by analysing patterns of degeneration following nerve lesions. The co-localization of neuromedin U immunoreactivity with immunoreactivity for substance P, neuropeptide Y, vasoactive intestinal peptide and calbindin was also investigated. Neuromedin U immunoreactivity was found in nerve cells in the myenteric and submucous plexuses and in nerve fibres in these ganglionated plexuses, around submucous arterioles and in the mucosa. Reactive fibres did not supply the muscle layers. Most reactive nerve cells in the myenteric ganglia had Dogiel type-II morphology and in many there was co-localization of calbindin, although some Dogiel type-II neuromedin U neurons were calbindin negative. Lesion studies suggest that these myenteric neurons project circumferentially to local myenteric ganglia. Projections from myenteric neurons also run anally in the myenteric plexus, while other projections extend to submucous ganglia, and still further projections run from the intestine to provide terminals in the coeliac ganglia. In the submucous ganglia neuromedin U was co-localized in three populations of nerve cells: (i) those with vasoactive intestinal peptide immunoreactivity, (ii) neurons containing neuropeptide Y, and (iii) neurons containing substance P. Each of these populations sends nerve fibres to the mucosa. Neuromedin U immunoreactivity is thus located in a variety of neurons serving different functions in the intestine and therefore probably does not have a single role in intestinal physiology.

Nerve growth factor receptor immunoreactivity in the neuronal perikarya of human sensory and sympathetic nerve ganglia.

We examined immunohistochemically the dorsal root ganglia, sympathetic ganglia, spinal cord, ventral and dorsal roots, and sciatic nerves obtained at autopsy from adult humans, using a monoclonal antibody against the human nerve growth factor receptor. We observed labelling in a granular pattern in the neuronal perikarya of dorsal root and sympathetic nerve ganglia. Ventral horn cells and axons were not labelled.

Quantification of herpes simplex virus infection in cervical ganglia of mice.

Mice were inoculated with herpes simplex virus (HSV) type 1 in the skin of the neck. The extent of primary and latent infection in the second and third cervical ganglia was investigated. Immunoperoxidase staining of ganglia during primary infection demonstrated HSV antigens initially in a restricted area of the ganglion. By the 5th day after infection, antigen was more widespread. Such a change in the staining pattern is explicable in terms of the zosteriform spread of virus from neurons innervating the site of infection to others supplying other areas of the dermatome. A maximum of approximately 10% of neurons became infected. By the 7th day staining was limited to a few cells. During latent infection, enzymic disaggregation of ganglia followed by immunoperoxidase staining or infectious centre assay indicated that virus reactivation began within 30 h of removal of ganglia and occurred in approximately 1% of viable neurons.

Distribution of peptidergic terminals of enteric origin in the rat celiac ganglion.

We examined the distribution of enterofugal nerve terminals of bombesin-, cholecystokinin- and vasoactive intestinal polypeptide-like immunoreactivity in the rat celiac-superior mesenteric ganglion complex. The majority of these nerve terminals were concentrated in the mesenteric side of the ganglion. The present findings suggest that some functional specialization occurs in the celiac ganglion of the rat.

Effects of aldose reductase inhibitor sorbinil on neuroaxonal dystrophy and levels of myo-inositol and sorbitol in sympathetic autonomic ganglia of streptozocin-induced diabetic rats.

Biochemical and ultrastructural effects of the aldose reductase inhibitor sorbinil were examined in two experimental rat models of chronic diabetic neuropathy: rats with streptozocin-induced diabetes (STZ-D) and rats fed a galactose-enriched diet. The frequency of neuroaxonal dystrophy in the superior mesenteric sympathetic ganglia of rats with untreated 8-mo STZ-D increased sevenfold compared with that in age-matched controls. Animals chronically maintained on a diet containing 50% galactose, however, did not develop neuroaxonal dystrophy in excess of that found in untreated age-matched control rats. Institution of sorbinil therapy at the time of induction of STZ-D decreased, but did not completely normalize, the frequency of neuroaxonal dystrophy without altering the severity of diabetes; this finding is based on measurements of plasma glucose, body weight, food consumption, 24-h urine volume, and levels of glycosylated hemoglobin. Sorbitol levels in the superior cervical sympathetic ganglia (SCG) of untreated 8-mo-diabetic animals increased three- to fourfold compared with levels in controls. The increase in sorbitol content of diabetic SCG was completely prevented by early institution of dietary sorbinil therapy. The myo-inositol content of 8-mo-diabetic SCG was modestly decreased compared with controls. Sorbinil administration improved but did not completely normalize diabetic SCG myo-inositol. The sorbitol content of the SCG, superior mesenteric and celiac sympathetic ganglia, and a major trunk of the superior mesenteric nerve of short-term (2.5-mo)-diabetic rats increased comparably, but only the diabetic SCG showed a decrease in myo-inositol.

Immunohistochemical evidence for different opioid systems in the rat superior cervical ganglion as revealed by imipramine treatment and receptor blockade.

The distribution pattern of opioid-immunoreactive nerve cell bodies and varicose fibres in the rat superior cervical ganglion after chronic administration of the tricyclic antidepressant imipramine, various receptor blockades (muscarinic antagonist, atropine sulphate; opiate antagonist, naloxone; kappa-antagonist, MR2266BS), and denervation was investigated immunohistochemically using a biotin-streptavidin-peroxydase complex method. Antisera to four peptides derived from two different precursors of the opioid family were used. In control superior cervical ganglia sparsely scattered nerve fibres and no neuronal cell bodies were immunoreactive when antisera to dynorphin A (1-17) or alpha-neo-endorphin (cleavage products of prodynorphin) were applied. A moderate number of nerve fibres and neuronal perikarya were immunoreactive to antisera directed against met-enkephalin-arg-phe (cleavage product of proenkephalin) and leu-enkephalin (cleavage product of prodynorphin and proenkephalin); non-identical cell bodies contained met-enkephalin-arg-phe- or leu-enkephalin-immunoreactivity. After drug treatment specific changes in the immunoreactivity of the investigated peptides in the superior cervical ganglion were demonstrated. (a) Treatment with imipramine resulted in an increase of nerve fibres demonstrating immunoreactivity to antisera against dynorphin A and alpha-neoendorphin. In contrast, no alteration in the numbers of nerve fibers but a numerical increase of postganglionic cell bodies immunoreactive to either met-enkephalin-arg-phe or leu-enkephalin antisera was demonstrated. Moreover, some perikarya exhibited immunoreactivity to both these opioids. (b) Receptor blockade with the muscarinic antagonist atropine sulphate or the general opiate antagonist naloxone had no effect on the number and distribution of dynorphin A or alpha-neoendorphin immunoreactive fibres, whereas both met-enkephalin-arg-phe and leu-enkephalin-immunoreactive fibres and postganglionic perikarya were increased in number. (c) After the kappa antagonist (MR2266BS), an increase of fibres with prodynorphin-derived opioid immunoreactivity as well as those with met-enkephalin-arg-phe- or leu-enkephalin-immunolabelling was visible and the met-enkephalin-arg-phe and leu-enkephalin-immunoreactive cell bodies were increased in number. The preganglionic origin of the investigated fibres with prodynorphin cleavage products was concluded from the complete disappearance of such fibres after preganglionic denervation. Denervation also resulted in an increase of met-enkephalin-arg-phe- and leu-enkephalin-immunoreactive perikarya. Small intensely fluorescent (SIF) cells, which in controls were nonrea

Cloning of a putative neuronal nicotinic acetylcholine receptor subunit.

A cDNA clone was isolated from a rat superior cervical ganglion cDNA library with an oligonucleotide that hybridized to muscle-like nicotinic acetylcholine receptor (nAChR) subunit cDNA. The deduced amino acid sequence possesses characteristics expected of a nAChR subunit that does not bind acetylcholine, in addition to distinctive features such as unique cysteine residues and N-linked glycosylation sites.

Evidence for GABAergic fibers entering the superior cervical ganglion of rat from the preganglionic nerve trunk.

The origin of gamma-aminobutyric acid immunoreactive (GABA-IR) nerve fibers present in the superior cervical ganglion (SCG) of rat was investigated. With immunocytochemical techniques many nerve fibers showed GABA-like positivity in the cervical sympathetic trunk, whereas similar staining could not be revealed in the internal carotid nerve or in the external carotid nerve. Ligation of the cervical sympathetic trunk for 24 h resulted a dramatic reduction in the staining density in the ganglion and in the cervical sympathetic trunk distal to the ligature. After transection of the preganglionic nerve fibers for eleven days or more, very few fibers staining for GABA were seen in the ganglion. The immunohistochemical results suggest that a major source of GABA within the SCG is a population of GABAergic axons entering from the preganglionic trunk.

Pronase treatment increases the staining intensity of GABA-immunoreactive structures in the paravertebral sympathetic ganglia.

A novel tissue preparation technique for improving gamma-aminobutyric acid (GABA) immunocytochemistry has been developed. The influence of the glutaraldehyde concentration in the fixative and the effect of pronase treatment on the GABA immunostaining were tested. This method includes fixation with a high concentration of glutaraldehyde, gelatin embedding and treatment of the sections with pronase. In sympathetic (paravertebral) ganglia and their connectives, the most intense and specific immunoreaction was obtained with the following procedure: immersion fixation in 5% glutaraldehyde, infiltration and embedding in 15% gelatin, secondary fixation of the samples with 4% formaldehyde, floating frozen sections and digestion with 0.1% pronase for 15-20 min. With this technique, the GABA-containing structures (cells and nerve fibers with varicosities forming basket-like networks around some principal neurons) were selectively labeled. The data presented suggest that (1) a high concentration (5%) of glutaraldehyde in the primary fixative is necessary to preserve a large proportion of the GABA content; (2) this glutaraldehyde fixation partly masks the GABA immunoreactivity; and (3) this masking may be overcome by a proteolytic treatment preceding the immunostaining. This method has been extensively tested for the light microscopic visualization of GABA-containing tissue components in the sympathetic ganglion chain, but it may probably also be used for the immunocytochemical detection of other small molecules in other parts of the nervous system.