Upregulation of CXCL10 in human dorsal root ganglia during experimental and natural varicella-zoster virus infection

Varicella-zoster virus (VZV) reactivation causes herpes zoster, which is accompanied by an influx of lymphocytes into affected ganglia, but the stimulus for this infiltrate is not known. We report that VZV infection of ganglia leads to increased CXCL10 production in vitro, in an explant ganglion model and in naturally infected dorsal root ganglia (DRG) during herpes zoster. Lymphocytes expressing the receptor for CXCL10, CXCR3, were also observed throughout naturally infected ganglia during herpes zoster, including immediately adjacent to neurons. This study identifies VZV-induced CXCL10 as a potential driver of T lymphocyte recruitment into DRG during herpes zoster.     

Infection by human varicella-zoster virus confers norepinephrine sensitivity to sensory neurons from rat dorsal root ganglia.

Varicella-zoster virus (VZV) is a widespread human herpes virus causing chicken pox on primary infection and persisting in sensory neurons. Reactivation causes shingles, which are characterized by severe pain and often lead to postherpetic neuralgia. To elucidate the mechanisms of VZV-associated hyperalgesia, we elaborated an in vitro model for the VZV infection of sensory neurons from rat dorsal root ganglia. Between 35 and 50% of the neurons showed strong expression of the immediate-early virus antigens IE62 and IE63 and the late glycoprotein gE. When the intracellular calcium concentration was monitored microfluorometrically for individual cells after infection, the sensitivity to GABA or capsaicin was similar in controls and in VZV-infected neurons. However, the baseline calcium concentration was increased. Neurons became de novo sensitive to adrenergic stimulation after VZV infection. Norepinephrine-responsive neurons were more frequent and calcium responses to norepinephrine were significantly higher after infection with wild-type isolates than with the attenuated vaccine strain OKA. The adrenergic agonists phenylephrine and isoproterenol had similar efficacy. We suggest that the infection with wild-type VZV isolates confers norepinephrine sensitivity to sensory neurons by using alpha(1)- and/or beta(1)-adrenergic receptors providing a model for the pathophysiology of the severe pain associated with the acute reactivation of VZV.     

Productive varicella-zoster virus infection of cultured intact human ganglia

Varicella-zoster virus (VZV) is a species-specific herpesvirus which infects sensory ganglia. We have developed a model of infection of human intact explant dorsal root ganglia (DRG). Following exposure of DRG to VZV, viral antigens were detected in neurons and nonneuronal cells. Enveloped virions were visualized by transmission electron microscopy in neurons and nonneuronal cells and within the extracellular space. Moreover, rather than remaining highly cell associated during infection of cultured cells, such as fibroblasts, cell-free VZV was released from infected DRG. This model enables VZV infection of ganglionic cells to be studied in the context of intact DRG.     

Quantitation of latent varicella-zoster virus and herpes simplex virus genomes in human trigeminal ganglia

Using real-time fluorescence PCR, we quantitated the numbers of copies of latent varicella-zoster virus (VZV) and herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) genomes in 15 human trigeminal ganglia. Eight (53%) and 1 (7%) of 15 ganglia were PCR positive for HSV-1 or -2 glycoprotein G genes, with means of 2,902 +/- 1,082 (standard error of the mean) or 109 genomes/10(5) cells, respectively. Eleven of 14 (79%) to 13 of 15 (87%) of the ganglia were PCR positive for VZV gene 29, 31, or 62. Pooling of the results for the three VZV genes yielded a mean of 258 +/- 38 genomes/10(5) ganglion cells. These levels of latent viral genome loads have implications for virus distribution in and reactivation from human sensory ganglia.     

Ribosomes in myelinated axons of dorsal root ganglia

Summary In the dorsal root ganglia of the rat, ribosomes were found not only in the initial segment, but they were also observed in the axoplasm of intraganglionar myelinated fibres and in the sensory portion of spinal nerves. Axons of seven-days-old rats contained more ribosomes than those of adult animals. The amount of particles decreased gradually from the initial segment trough intraganglionar internodes to the axons of spinal nerves. No ribosomes were found in axons of dorsal roots. In intraganglionar fibres, ribosomal particles were usually observed near the nodes of Ranvier, in the vicinity of Schmidt-Lantermann clefts and in axons near the Schwann cell nuclei. They were arranged in tetrads, pentads or in larger polysomes, and they were often observed adjacent to a group of mitochondria.The particles had invariably a stable size, their average diameters measuring 234 ± 2 × 197 ± 3 Å, which is practically equal to the diameters of 232 ± 2 × 203 ± 3 Å of ribosomes in the Schwann cell cytoplasm. These values fall within the range of diameters of ribosomes isolated from various cells of eukaryotic organisms as given in the literature. Since no other granular component of the cytoplasm has similarly stable dimensions, the measurements are considered to prove that the axonal particles described here are ribosomes.The author wishes to thank Dr. K. Smetana for his valuable suggestions and Mrs. M. Sobotková, Ing. M. Doubek and Mr. H. Kunz for their skillful technical assistance. The investigation was in part supported by a grant-in-aid from the Muscular Dystrophy Associations of America, Inc.     

Differential localization of lectin binding sites and neuropeptides in human dorsal root ganglia

The subpopulations were compared of neurons in human dorsal root ganglia (DRG), as substance P, identified by somatostatin, Glycine max lectin (SBA) specific to terminal N-acetylgalactosamine, and Ulex europaeus I agglutinin (UEA-I) specific to l-fucose. The lectins and neuropeptides all bound to neurons of small diameter. Furthermore, the majority of the SBA binding neurons or somatostatin positive neurons were also UEA-I binding neurons. However, SBA binding neurons were not colocalized with somatostatin or substance P. Less than 20% of substance P positive neurons showed colocalization with l-fucosyl residues, and approximately 10% of l-fucosyl residues showed colocalization with substance P. Our results suggest that both l-fucose and terminal N-acetylgalactosamine containing neurons in the human DRG are subjected to different subpopulations from substance P or somatostatin positive neurons.     

Prenatal development of the rat dorsal root ganglia

Summary This study describes three-dimensional aspects of the development and pseudo-unipolarization of neuroblasts and the maturation of satellite cells in prenatal rat dorsal root ganglia, using scanning electron microscopy, after removal of extracellular connective tissue components by trypsin digestion and HC1 hydrolysis.At 14 days of gestation, the vast majority of neurons are spindle-shaped or bipolar and only 3% are unipolar, while at 16 and 18 days this percentage has increased to 30% and 91%, respectively. The initial portions of the central and peripheral neuronal processes gradually approach each other and form a common initial portion. Finally, the cytoplasm of this common initial portion becomes thinner and elongates to form the stem process of the mature cell.Satellite cells are present from the beginning of the period studied, but intricate networks of branching satellite cell processes only develop after about day 17.     

Quantitation of latent varicella-zoster virus DNA in human trigeminal ganglia by polymerase chain reaction.

Competitive polymerase chain reaction was used to quantitate latent varicella-zoster virus (VZV) DNA in human trigeminal ganglia. Ganglionic DNA from five subjects was amplified with oligonucleotide primers specific for VZV gene 28. Two of the samples were also analyzed with primers specific for VZV gene 62. Our results indicated that there are 6 to 31 copies of the VZV genome in every 100,000 ganglionic cells.     

Adult mouse dorsal root ganglia neurons in cell culture

A method has been developed for the long-term culture of dissociated adult mouse dorsal root ganglia (DRG). Of critical importance to the success of this technique was a three-hour incubation in collagenase which softened the DRG and permitted gentle dissociation. The morphological and electrophysiological features of the dissociated adult DRG were similar to those observed in previous studies of immature (i.e., embryonic and newborn) DRG in culture and also to those of adult DRG in situ. With regard to electrophysiological work, the adult DRG neurons are superior to embryonic and newborn neurons because of their larger size and greatly increased survival in culture (no degeneration for first six days, and thereafter a relatively slow decrease). The adult neurons regenerated nerve fibers to an extent comparable to that of immature neurons. Therefore, the adult DRG cultures might be useful to study factors influencing regeneration in the adult mammalian nervous system. The adult cultures might also be useful to investigate factors influencing the aging process.     

Dopamine as trace amine in the dorsal root ganglia

It has been shown that in the chick dorsal root ganglion (DRG) about 8% of neurons, belonging to both the A and B classes of sensory neurons exhibit a clear dopamine immunoreactivity. In the present study are reported the results of measurements, by mean of HPLC-electrochemical detection (HPLC-ED), of DA and of the DA metabolites dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in the rat DRG and their central nerves. Very low levels of DA, about 10 folds lower than the levels found in the dorsal horn of the spinal cord, were found in the DRG. However the levels of DOPAC and HVA were approximately equivalent to the levels found in the cord. The immunocytochemical study performed in parallel has shown that some dopaminergic-immunoreactive fibers in the DRG are located around the blood vessels. Few dopamine-immunoreactive sensory neurons were identified in the DRG and immunoreactive fibers, not linked to blood vessels, were identified in the dorsal root nerves. The present work indicates that there is a dopaminergic innervation of the blood vessels in the rat DRG but that dopamine may also be, as in the chick, a transmitter of primary afferent fibers.     

Characterization of acute and latent herpes simplex virus infection of dorsal root ganglia in rats.

The characteristics of HSV type-1 infection following subcutaneous inoculation in the dorsum of one hind paw of Sprague-Dawley rats were studied to determine whether infection in rats might more closely parallel the infection in man than is seen in other animals. The serologic and virologic characteristics of acute and latent ganglion infection conformed to those of human infection. Immunohistochemical studies suggested that sensory ganglion infection arose via centripetal axonal migration of virus as is hypothesized in man. In rat, small type B neuronal cell bodies appeared central to the maintenance of latent infection and reactivation observed during cocultivation of lumbar ganglia. Acute and latent lumbar sensory ganglion infection in rats after subcutaneous hind paw injection of HSV-1 appears to be another suitable model of this infection in man.     

Demyelination and cytopathic effects in cultures of mammalian dorsal root ganglia infected with encephalomyocarditis virus.

Replication of encephalomyocarditis virus and its cytopathic effects were studied in myelinated cultures of dorsal root ganglia obtained from newborn mice. Six hours after infection virus progeny was detected in the culture. At 24 h the virus titer reached 2 times 10(6) PFU per culture and remained at this level until 48 h. The first cytopathic alterations began at 24 h and consisted of rounding of Schwann and satellites cells and their detachment from neurons. Later, bead-like swellings of the myelin appeared along the axons followed by splitting and degeneration of lamellae. The cytopathic effect in the neurons started 29 h after infection, reaching complete neuronolysis at 48 h. Virus particles, scattered or arranged in crystal-like aggregates, were first seen in the cytoplasm of glial cells and then in neurons and axons.     

Conditional gene deletion in primary nociceptive neurons of trigeminal ganglia and dorsal root ganglia

The use of Cre-loxP technology for conditional mutagenesis in pain pathways had been restricted by the unavailability of mice expressing Cre recombinase selectively in functionally distinct components of the nociceptive system. Here we describe the generation of transgenic mouse lines which express Cre recombinase selectively in sensory ganglia using promoter elements of the Na(v)1.8 gene (Scn10a). Cre-mediated recombination was greatly evident in all nociceptive and thermoreceptive neurons of the dorsal root ganglia and trigeminal ganglia, but only in a small proportion of proprioceptive neurons. Cre-mediated recombination was not detectable in the brain, spinal cord, or any nonneural tissues and began perinatally after invasion of primary afferents into the developing spinal cord. Thus, these mice enable selective deletion of genes in subsets of sensory neurons and offer a wide scope for studying potential functions of genes in pain perception, independent of secondary effects arising from developmental defects or global gene ablation.