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.

Morphology and distribution of primary afferent fibres expressing alpha-galactose extended oligosaccharides in the spinal cord and brainstem of the rat. Light microscopy.

The light microscopic morphology and distribution of non-substance P-containing small primary afferent fibres were studied. These fibres were labelled using LD2 and LA4 monoclonal antibodies which recognize alpha-galactose extended oligosaccharides expressed by primary afferent neurons. The LD2 and LA4 antibodies immunostained small primary afferent fibres ending mainly in lamina II of the spinal cord dorsal horn and trigeminal subnucleus caudalis of the rat. The lamination pattern of both types of primary afferents was assessed using an image analysis system. The highest density of LD2-immunoreactive fibres was located in a patchy band located in lamina II outer, while LA4-immunoreactive fibres were distributed mainly through lamina II inner. In lateral regions of cervical and lumbar dorsal horn the LA4-immunoreactive band is broader and comprises almost all lamina II. In contrast to substance P-containing primary afferents, a low density of LD2- or LA4-immunoreactive fibres was found in lamina I, and no terminal fields were found in lamina V or lamina X of the spinal cord or in levels of the trigeminal system outside the subnucleus caudalis. Both antibodies also labelled the parent fibres in the white matter fasciles. LD2-immunoreactive fibres were located in the dorsal roots, medial regions of the Lissauer tract, dorsal columns of the spinal cord, outer regions of the spinal trigeminal tract and dorsal to the cuneatus and gracilis nuclei. In contrast, LA4-immunoreactive fibres were restricted to the dorsal roots, medial and lateral regions of the Lissauer tract and the outer regions of the trigeminal tract. Immunostained fibres in the rootlets of the X and IX nerves and immunoreactive terminal arborizations in various subnuclei of the nucleus tractus solitarius were seen using both antibodies. These results show that subpopulations of small primary afferents stained by LD2 and LA4 antibodies have distinct patterns of central distribution and are consistent with a subdivision of small primary afferents into peptide- and non-peptide-containing groups.

Detection of HSV-1 proteins prior to the appearance of infectious virus in mouse trigeminal ganglia during reactivation of latent infection.

Following corneal inoculation of mice HSV-1 produces an acute infection and establishes a latent infection in trigeminal ganglia. The latent virus can be reactivated in vitro by explantation of ganglionic tissue. Viral protein expression was studied in trigeminal ganglia during acute infection of mice and explant reactivation of latent infection. HSV-1 proteins were detectable by immunoprecipitation and immunostaining, in mouse ganglia only from 3-5 days post infection. Although during explant reactivation it has been demonstrated that at 24 h post-explant the trigeminal ganglia are all infectious virus negative (Spivack, O'Boyle II and Fraser (1987) J. Virol. 61, 3288-3291), we have found that three HSV-1 proteins, of 175 kDa, 110 kDa and 90 kDa, are present in latently infected trigeminal ganglia as early as 6-21 h post explantation. Initially, only neuronal cells were positive by immunostaining with anti HSV-1 polyclonal serum for HSV-1 antigens, but at later times HSV-1 antigens were seen in non neuronal cells as well. These proteins may play a role in the initial stages of the reactivation process.

Calcitonin gene-related peptide (CGRP) immunoreactive sensory nerves in the human and guinea pig uvea and cornea.

The presence of calcitonin gene-related peptide (CGRP) immunoreactive nerves in the uvea and cornea of human and guinea pig eyes was evaluated using immunohistochemical techniques. CGRP immunoreactivity was found in thin, varicose nerve fibers in both species. Most of the fibres were localized in the ciliary body, and were mainly associated with blood vessels. In the human ciliary body, a moderate number of CGRP immunoreactive nerves were also seen in the ciliary muscle. In the iris and cornea, CGRP immunoreactive fibres were relatively uncommon. In the iris, they were mostly found associated with blood vessels, while in the cornea they were seen sub-epithelially or as free nerve endings in the epithelium. In the trigeminal ganglion, small sized ganglion cells displayed CGRP immunoreactivity. About 40% of all ganglion cells were immunoreactive nerves in the guinea pig, while sympathetic denervation did not change the staining pattern of CGRP immunoreactivity. The present findings, together with previous physiological data, suggest that CGRP might play a role in the regulation of the blood flow, aqueous humour dynamics, and neurogenic inflammation, not only in experimental animals but also in man.

Effects of lipofuscin on in situ hybridization in human neuronal tissue.

In situ hybridization is a highly sensitive technique for detecting nucleic acid sequences within tissues, and is frequently employed in neurovirology. However, this technique requires many appropriate controls in order to recognize and avoid potential artifactual hybridization. We have encountered abundant reaction to lipofuscin in neurons in human peripheral and central nervous systems, using various DNA probes, which could be misinterpreted as positive signals. This pseudohybridization reaction was resistant to treatment with RNase or DNase and was also present in tissue sections treated with hybridization mixture or nuclear autoradiographic emulsion in the absence of any radioactive probes. Characteristics used to distinguish between authentic in situ hybridization and the reaction to neuronal lipofuscin include cellular localization, color, margins and granular appearance, sensitivity to treatment with nucleases and the effect of exposure time on signal intensity. These guidelines should be used to avoid potential misinterpretation of in situ hybridization results with human tissue.

Evidence for the presence of substance P in cat nasal receptor afferents.

The afferents of the nasal receptors responsible for many upper airway protective reflexes are carried in the ethmoidal branch of the ophthalmic division of the trigeminal nerve. Previous electrophysiological studies indicate that a significant number of ethmoidal afferents respond to noxious stimuli applied to the nose. The objective of the present study was to identify ethmoidal nerve cell bodies within the trigeminal ganglion which demonstrated the presence of the neurotransmitter substance P (SP). SP is believed to be involved in the relay of nociceptive information. A double-labelling technique was employed and involved tracing the afferents to their cell bodies using horseradish peroxidase (HRP) and subsequent identification of SP-immunoreactivity within HRP-filled cells using monoclonal antibody immunohistochemistry. SP-immunoreactive cell bodies constituted 43 per cent-50 per cent of the total number of labelled ethmoidal cell bodies within the trigeminal ganglion. Although ethmoidal cell bodies were much smaller than the overall population of trigeminal ganglion cells, the size of SP-immunoreactive ethmoidal cell bodies was not significantly different from that of ethmoidal cell bodies not exhibiting SP-immunoreactivity.

RNA complementary to herpes simplex virus type 1 ICP0 gene demonstrated in neurons of human trigeminal ganglia.

Recent studies with mice have demonstrated abundant RNA transcripts which are complementary (antisense) to the herpes alpha gene ICP0 in latently infected ganglia. We investigated the situation in unselected human trigeminal ganglia. Strand-specific 2.7-kilobase herpes simplex virus type 1 (HSV-1) ICP0 RNA probes were prepared, and their sense was determined in productively infected cells. Although in situ hybridization demonstrated ICP0 antisense RNA transcripts in the nuclei of neurons in 46% of the ganglia, ICP0 messenger RNA was not found in any of the ganglia. We conclude that HSV-1 antisense ICP0 RNA is present in humans during ganglionic latency.

Localization of proprioceptive neurons innervating the muscle spindles of pig extraocular muscles studied by horseradish peroxidase labelling.

Each muscle of the extraocular muscles, containing abundant muscle spindles, was exposed to horseradish peroxidase (HRP) on 8 young pigs (2-month-old, 20-30 kg in body weight, both sexes). The results obtained are follows: The HRP-labelled neurons innervating the superior rectus muscle were always found in a crescent ventro-medio-dorsal fashion in the most medial position of the contralateral oculomotor nucleus. The HRP-labelled cells for the medial rectus muscle appeared close to the superior rectus group in the ipsilateral nucleus. The labelled cells for the inferior rectus muscle appeared in the ventrolateral position of the ipsilateral nucleus and those for the inferior oblique muscle in the area between the medial rectus and inferior rectus muscle groups. The labelled cells for the superior oblique muscle were found in the contralateral trochlear nucleus and those for the lateral rectus muscle bilaterally in the abducens nuclei, predominantly on the ipsilateral side and poorly on the contralateral side. The HRP-labelled cells were composed of large (alpha) and small (gamma) multipolar cells and of bipolar, oval or round (proprioceptive) cells, all intermingled together within the nucleus. The bipolar cells have been also identified in the 3 nuclei by means of Nissl staining technique. On this basis, they should be considered as the proprioceptive neurons. In the shrew-moles, the cell bodies of the proprioceptive neurons innervating the snout muscle spindles have been found close to the ipsilateral glossopharyngeal ganglion and those of the somatic sensory neurons in the ipsilateral trigeminal ganglion. In the pigs, no HRP-labelled cells were found in the trigeminal mesencephalic tract nucleus, but the HRP-labelled cells were found in the ipsilateral trigeminal and the superior cervical sympathetic ganglia. From the results, it could be emphasized that the proprioceptive neurons innervating the pig extraocular muscle spindles are located within the nuclei of the IIIrd, IVth and VIth cranial nerves.

Distinct substance P and calcitonin gene-related peptide immunoreactive nerves in the guinea pig eye.

Using a double labeling indirect immunofluorescent technique, we studied the guinea pig trigeminal ganglion and eye for co-localization of substance P and calcitonin gene-related peptide. In the trigeminal ganglion, the number of neurons immunoreactive for calcitonin gene-related peptide significantly outnumber those immunoreactive for substance P, but virtually all substance P positive neurons are immunoreactive for calcitonin gene-related peptide. In the eye, a complex pattern of co-localization is present; both peptides co-localize in most immunoreactive nerve fibers. Nerve fibers immunoreactive only for calcitonin gene-related peptide tend to be concentrated in the cornea and posterior ciliary body. Nerve fibers immunoreactive only for substance P are present in relation to both iris muscles. Sensory denervation by intracranial transection of the ophthalmic and maxillary nerves fails to eliminate these substance P positive but CGRP negative iris nerve fibers. These findings indicate an alternative origin for substance P immunoreactive nerves supplying the iris muscles in this species.

Innervation of facial skin but not masticatory muscles or the tongue by trigeminal primary afferents containing somatostatin in the rat.

Using retrograde transport of a fluorescent dye, True blue, the peripheral tissues innervated by trigeminal ganglion neurones containing somatostatin have been investigated. Of ganglion neurones retrogradely labelled from injections of dye into the facial skin, 3.45% were found to be immunoreactive for somatostatin. In contrast, none of the neurones labelled from injections of dye into the tongue or masseter muscle were found to contain this peptide. This demonstration of a restricted distribution of somatostatin-containing primary afferents raises the possibility that somatostatin may be involved in functions which are specific to skin and not to the other tissues examined.

Comparison of the distribution of Na+,K+-ATPase and myelin-associated glycoprotein (MAG) in the optic nerve, spinal cord and trigeminal ganglion of shiverer (shi/shi) and control (+/+) mice.

Na+,K+ ATPase and myelin-associated glycoprotein (MAG) were studied by immunocytochemistry on paraffin sections of the spinal cord, optic nerve and trigeminal ganglion of adult control (+/+) and CNS myelin-deficient shiverer (shi/shi) mice. Immunostaining for Na+, K+-ATPase outlined the periphery of nerve fibers in the spinal cord white matter, optic nerve and trigeminal ganglion of +/+ and shi/shi mice. Immunostaining for Na+,K+-ATPase appeared somewhat denser in the optic nerve and spinal cord lateral funiculi of shi/shi than in +/+ mice. In addition, immunostaining for Na+,K+-ATPase was demonstrated at the plasmalemma of presumed satellite cells situated at the periphery of ganglion cell bodies in the trigeminal ganglion of both species of mice. Immunostaining for MAG was localized along the periphery of nerve fibers in the spinal cord funiculi (with little immunostaining within gray horns), optic nerve and trigeminal ganglion of both +/+ and shi/shi mice. The major differences between shi/shi and +/+ mice were that the number of MAG-immunostained nerve fibers was greatly reduced in the spinal cord funiculi and the density of immunostaining was slightly increased in the optic nerve of shi/shi mice. The numbers of MAG-immunostained nerve fibers in trigeminal ganglion were similar in both species. Also, the cytoplasm of some oligodendrocyte-like cells was found densely immunostained for MAG in the spinal cord and optic nerve of shi/shi mice, but not of +/+ mice. This light microscopic study provides evidence that the defective shiverer gene leads to a decrease in MAG deposition and to aggregations of MAG-like material within perikarya of oligodendrocyte-like cells in regions of the CNS.

Insulin receptors in the peripheral nervous system: a structural and functional analysis.

Although the brain is known to contain specific insulin receptors, there is no information on whether these receptors are also present in the peripheral nervous system (PNS). The present studies sought to provide this information by characterizing insulin binding in bovine autonomic (superior cervical) and sensory (trigeminal) ganglia. It was found that both ganglia contain specific, high-affinity receptors for insulin. Like insulin receptors in other tissue, these receptors could be solubilized and purified on wheat germ agarose columns and were found to have tyrosine-specific kinase activity. SDS-PAGE and autoradiography revealed that the apparent molecular weight (Mr) of the PNS insulin receptor was approximately 133 kDa which is similar to the Mr of hepatic receptors, but is approximately 10 kDa larger than the insulin receptor found in the brain. Because the vasculature of autonomic and sensory ganglia is fenestrated, it is possible that PNS insulin receptors are exposed to blood-borne insulin.

A quantitative correlation of substance P-, calcitonin gene-related peptide- and cholecystokinin-like immunoreactivity with retrogradely labeled trigeminal ganglion cells innervating the eye.

To evaluate the intraganglionic organization of ocular sensory neurons in the guinea pig, we studied the retrograde axonal transport from the eye to the trigeminal ganglion of cholera toxin B subunit and then applied immunohistochemistry for substance P, calcitonin gene-related peptide and cholecystokinin. Retrogradely labeled cells were observed only in the anteromedial portion of the ipsilateral ganglion. We observed no somatotopical organization to trigeminal neurons containing any of these three peptides, either for cells projecting to the eye or for the ganglion as a whole. The relative proportion of neurons immunoreactive for each of these three peptides was similar among the population of neurons retrogradely labeled with cholera toxin B and among the population of neurons without direct projections to the eye.

Detection of bovine herpesvirus type 1 RNA in trigeminal ganglia of latently infected rabbits by in situ hybridization.

At times after conjunctival inoculation with bovine herpesvirus type 1 (BHV-1), representing the acute and latent phases of infection, rabbit trigeminal ganglia were examined for the presence of BHV-1 nucleic acids by in situ hybridization using a 3H-labelled BHV-1 DNA probe. During the acute phase of virus infection, both BHV-1 DNA and RNA were detected in ganglionic neurons and occasionally in adjacent satellite cells. However, during the latent phase of infection only viral RNA was detectable in involved neurons. Viral RNA appeared restricted to the nucleus of latently infected cells and was present in varying amounts in individual cells. These results indicate that the BHV-1 genome is transcriptionally active in ganglionic neurons during latent infection.

Fibrous xanthoma in the gasserian ganglion: case report.

Occurrence of fibrous xanthoma has been reported increasingly in the skull and the central nervous system, but is extremely rare in the gasserian ganglion. We report on the clinical presentation, radiological appearance, surgical treatment, and histological makeup of a fibrous xanthoma arising from the left gasserian ganglion.

The effect of sympathectomy on calcitonin gene-related peptide levels in the rat trigeminovascular system.

The effect of sympathectomy on the calcitonin gene-related peptide (CGRP) level in the rat primary trigeminal sensory neurone was investigated. Six weeks after bilateral removal of the superior cervical ganglion there was a 70% rise in the CGRP content of the iris and the pial arteries, a 34% rise in the concentration in the trigeminal ganglion but no change in the brainstem. The CGRP rise in both end organs suggests that this phenomenon may be common to all peripheral organs receiving combined sensory and sympathetic innervations. The lack of any rise in the brainstem CGRP content raises the possibility that this process spares central terminations. In contrast, the level of neuropeptide Y, a peptide mainly contained in sympathetic terminals, fell to 35% of control values in the iris and pial arteries whilst the trigeminal ganglion and brainstem concentrations remained unchanged. The possible relevance of these observations to the clinical syndrome of postsympathectomy pain (sympathalgia) is discussed. There are similarities between the delayed onset of the human pain state and the delayed rise in sensory peptides after sympathectomy.

Nerve growth factor supply for sensory neurons: site of origin and competition with the sympathetic nervous system.

Nerve growth factor (NGF) levels in the sensory nervous system were measured by a highly sensitive two-site enzyme immunoassay for NGF. Dorsal root ganglia and the adjacent spinal nerves contained 2.8 +/- 0.3 and 1.7 +/- 0.4 ng NGF/g wet wt., respectively, whereas no NGF was detectable in dorsal roots and spinal cord (less than 0.05 ng NGF/g wet wt.). It is concluded that sensory neurons are supplied with NGF exclusively from their peripheral and not from their central field of projection. Two days after treatment with 6-hydroxydopamine, which destroys sympathetic nerve terminals and thereby prevents the removal of NGF by sympathetic neurons, the NGF content of dorsal root ganglia and trigeminal ganglia increased to 285% and 161% of control, respectively. This indicates that in peripheral target tissues sensory and sympathetic neurons compete for NGF.

[Morphologic and histochemical studies of the development of chloroquine-induced storage disease in the semilunar ganglion of the rat]. / Morphologisch-histochemische Untersuchungen zur formalen Genese der Chloroquinspeicherdystrophie im Ganglion semilunare der Ratte.

Intralysosomal storage of polar lipids can be induced in rats by treatment with Chloroquine, which belongs to cationic amphiphilic compounds. The drug accumulates in lysosomes of cells of the peripheral nervous system and interferes with the catabolism of polar lipids. As a consequence the lysosomes are transformed into bodies which are characterized by an internal lamellated pattern. The lipids stored in the neuron cells are phospholipids. The purpose of the present study was to check up the chronological development of the structural and lipid histochemical alterations within neuron cells. A significant enlargement of perikarya was observed after treatment of 94 d. A moderate increase of lysosomes and phospholipids was noticed after 82 d. Some typical foam-cells were seen after a period of 250 d. Moreover, a special mitochondrial alteration was observed, which could express close correlations between phospholipid metabolism and damaged parts of mitochondria.

Detection of herpes simplex virus mRNA in latently infected trigeminal ganglion neurons by in situ hybridization.

Latent infection of the trigeminal ganglion with herpes simplex virus type 2 (HSV-2) was studied in guinea pigs by in situ DNA hybridization. Frozen ganglion sections from animals killed during the period of latent virus infection were studied under nondenaturing conditions. Some sections were treated with deoxyribonuclease (DNase) or ribonuclease (RNase) before incubation with HSV DNA probes. HSV probes consisted of viral DNA nick translated and labeled in vitro with tritiated nucleotides. Bacteriophage lambda DNA, similarly prepared, was used as a control probe. The lambda probe was negative in all situations, including HSV-2-infected monolayer cells in cell culture. HSV-2 probes produced heavy label and, therefore, evidence of hybridization with HSV-2-infected monolayer cells. When HSV-2 probes were incubated with latently infected ganglion sections, hybridization was detected in 71% of guinea pigs and 46% of ganglia. Label was seen only in neurons, and in positive ganglia 0.3 to 5% of neurons were labeled. The amount of label was markedly decreased by pretreatment of ganglion sections with RNase but not DNase, indicating that the DNA probes hybridized to HSV messenger RNA in the latently infected ganglia.