Human Trigeminal Ganglia Research Articles

Capsaicin receptor immunoreactivity in the human trigeminal ganglion

The cloned capsaicin receptor, also known as vanilloid receptor subtype 1 (VR1) receptor, has been demonstrated to be an integral membrane protein with homology to a family of putative store-operated calcium channels. The VR1 receptor is activated not only by capsaicin but also by noxious heat and protons, and therefore it is suggested as a molecular integrator of chemical and physical stimuli that elicit pain. In the present study, indirect immunofluorescence detected a small number of neurons that are VR1 receptor immunoreactive (ir) (171 versus 1038 or 16% of all neuronal cell bodies) in the human trigeminal ganglion (TG). In addition, RT-PCR confirmed the presence of VR1 mRNA in the human TG. It has been hypothesized that TG neuronal cell bodies are the source of capsaicin-stimulated release of calcitonin gene-related peptide (CGRP), and hence co-localization experiments were performed. Around 10% of the VR1 receptor-ir is expressed on neurons that contain CGRP-ir (ten among 74) in the human TG, indicating that capsaicin may act through the VR1 receptor to modulate the release of CGRP and in turn to modulate pain. We observed that 8% of the VR1 receptor-ir neuronal cell bodies contain substance P-ir and 5% nitric oxide synthase. Capsaicin can release nitric oxide, CGRP and substance P from sensory nerves and contribute to central sensitization.

Does nociceptin play a role in pain disorders in man?

Nociceptin-immunoreactive cellbodies were detected in the human trigeminal ganglion, while no such fibers were identified in the temporal artery or in dermal tissue from the neck region. In four healthy subjects receiving nociceptin into the temporal muscle in an open labeled design no pain was detected. In 10 healthy subjects who received 200 pmol of nociceptin into tender non-dominant trapezius muscles in a placebo-controlled, randomized, balanced, and double-blinded design local tenderness increased ( P=0.025) while no pain was noted. Thus, the action of nociceptin should be searched for in the trigeminal ganglion and/or in the central nervous system (CNS).

Sensory nerves in the human cerebral circulation and trigeminal ganglion: role in primary headaches

Our knowledge of the nervous control of cerebral circulation has increased by the use of denervations and retrograde tracing in combination with immunohistochemical techniques. We have demonstrated that cerebral vessels are supplied with sensory nerve fibers containing a multiplicity of transmitter substances originating in the trigeminal ganglion. The majority of these transmitters are neuropeptides, but the gaseous signal substance, nitric oxide (NO), is also included. In primar headaches, calcitonin generelated peptide (CGRP) is released in parallel with the headache, while the parasympathetic nerve transmitter vasoactive intestinal peptide (VIP) is released in parallel with facial symptoms. Thus, the perivascular nerves participate in the pathogenesis of primary headaches. Current migraine drugs, e. g. triptans, act in part by inhibiting the release of CGRP from the sensory nerves.

Fractionation of neurons and satellite cells from human sensory ganglia in order to study herpesvirus latency

A method is described for fractionating human trigeminal ganglia into highly purified populations of neurons and satellite cells in order to study α-herpesvirus latency. The method was validated by microscopy of the separated populations and by the observation that only the neuronal population, not the satellite cells, contained herpes simplex virus (HSV) DNA. The frequency of detecting HSV in neurons from ganglia was 3% (43 of 1440 neurons). HSV DNA was not detected in approximately 17,500 satellite cells. The HSV DNA genome copy number in single cells ranged from 2 to 50. These data on the frequency and cellular location of latent HSV indicate that our mechanical fractionation of cell types results in low levels of cross-contamination and provides samples from which cells infected latently can be studied at the single cell level.

AMPA-type glutamate receptor subunits 2/3 in the human trigeminal sensory ganglion and subnucleus caudalis from prenatal ages to adulthood

Using immunohistochemistry, the occurrence and distribution of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) glutamate receptor subunits GluR2/3 is shown in the human trigeminal ganglion and subnucleus caudalis from 20 weeks of gestation to adulthood. In the trigeminal ganglion a subpopulation of GluR2/3-like immunoreactive (LI) primary sensory neurons occurred at all examined ages, amounting to about 20% of all ganglion cells in the earliest pre-term newborn and in the adult, to about 30% at 24 and 32 weeks of gestation, and peaking to about 40% in the neonate. At all ages examined, GluR2/3-LI neurons were heterogeneous in size, although in the adult most of the labeled perikarya were large-sized, with a mean cell diameter above 35 μm. In the trigeminal subnucleus caudalis, positive elements could be first detected at 30 weeks of gestation and persisted at all other examined ages. At pre- and perinatal ages, the immunoreactivity was restricted to neuronal perikarya in the superficial layers and in the marginal zone of the nucleus. In the adult tissue, the subnucleus caudalis harbored a loose meshwork of varicose thread- and dot-like elements in the superficial layers and numerous immunoreactive neurons, distributed in lamina I, substantia gelatinosa, and in the superficial zone of the magnocellular region.

Key issues in varicella-zoster virus latency.

The molecular mechanisms by which varicella-zoster virus (VZV) causes a latent infection in human trigeminal and spinal ganglia are not well understood. It is known that VZV establishes latency in ganglia following the primary infection causing varicella (chickenpox), and that the virus may reactivate after years of dormancy to produce herpes zoster (shingles). Two key issues have been the cell-type localization of latent VZV in human ganglia, and the nature and extent of VZV gene expression during latency. Although the cell specificity of latent VZV has been controversial for almost a decade, it is now widely accepted that the virus is mainly latent in neuronal cells, with only a small proportion of non-neuronal cells infected. All of the studies carried out so far have indicated that VZV gene expression is highly restricted during ganglionic latency. Although at least four VZV genes have been identified as being expressed, the possibility that latent gene expression is significantly greater than this cannot yet be excluded. There is also evidence for VZV gene-encoded proteins being expressed during latency, although the precise extent of this is unclear. Advances in this difficult field may be expected to arise from both newly developed molecular technology and more refined animal models of VZV latency.

Atomic Force Microscopy Imaging of the Human Trigeminal Ganglion

Thispaper describes an investigation of gangliocytes via imaging semithin sections of two human trigeminal ganglia with an atomic force microscope (AFM). Whereas semithin sections are usually employed for transmission electron microscopy, we adopted this special type of sample preparation for our AFM studies to extract topographical data from the gangliocyte itself and from the nucleus, the nucleolus, the crystal-arranged lipofuscin granules, and the cell-surrounding mantle cells; simultaneously we characterized the samples with error signal mode. This AFM-related technique revealed no information concerning friction force and elasticity due to the presence of the embedding material (epoxy), but it gave additional topographical contrast. These are the first images of the human trigeminal ganglion by AFM.

Prevalence of HSV-1 LAT in human trigeminal, geniculate, and vestibular ganglia and its implication for cranial nerve syndromes.

Herpes simplex virus type 1 (HSV-1) enters sensory neurons and can remain latent there until reactivation. During latency restricted HSV-1 gene expression takes place in the form of latency-associated transcripts (LAT). LAT has been demonstrated to be important not only for latency but also for reactivation, which may cause cranial nerve disorders. Tissue sections of the trigeminal ganglia (TG), geniculate ganglia (GG), and the vestibular ganglia (VG) from seven subjects were examined for the presence of LAT using the in situ hybridization technique. LAT was found on both sides in allTG (100%), on both sides of five subjects (70%) in the GG, and in none of the VG. Using a second more sensitive detection method (RT-PCR), we found LAT in the VG of seven of ten other persons (70%). This is the first study to demonstrate viral latency in the VG, a finding that supports the hypothesis that vestibular neuritis is caused by HSV-1 reactivation. The distribution of LAT in the cranial nerve ganglia indicates that primary infection occurs in the TG and GG and subsequently spreads along the faciovestibular anastomosis to the VG.

5-HT 1B and 5-HT 1D receptors in the human trigeminal ganglion: co-localization with calcitonin gene-related peptide, substance P and nitric oxide synthase

5-Hydroxytryptamine (5-HT) is implicated in migraine and agonist directed aganist 5-HT 1B and 5-HT 1D receptors are commonly used as effective therapies. The antimigraine mechanisms involve the inhibition of intracranial sensory neuropeptide release. In order to determine which 5-HT 1 receptor subtypes are involved we have by immunocytochemistry examined the distribution of 5-HT 1B and 5-HT 1D receptors in the human trigeminal ganglia, and addressed which of them colocalize with calcitonin gene-related peptide (CGRP), substance P (SP) or nitric oxide synthase (NOS). We detected that 5-HT 1D receptor immunoreactivity (i.r.) was predominantly expressed in medium-sized cells (86% of positive cells, 30–60 μm). About 9% of the 5-HT 1D receptor i.r. cells were large in size (>60 μm) and 5% were small in size (<30 μm). In a similar pattern, 5-HT 1B receptor i.r. was mainly expressed in medium-sized cells (81% in 30–60 μm, 15% in >60 μm and 4% in <30 μm). Double immunostaining was used to determine whether the 5-HT 1B or 5-HT 1D receptor immunoreactive cells co-localized with either CGRP, SP or NOS. Thus, 89% of the CGRP i.r. cells expressed 5-HT 1D receptor i.r. and 65% of the CGRP positive cells were 5-HT 1B receptor positive. Most of the 5-HT 1D (95%) and the 5-HT 1B (94%) receptor i.r. cells showed SP immunostaining and 83% of 5-HT 1D receptor and 86% of 5-HT 1B receptor i.r. cells contained NOS. In conclusion, both 5-HT 1B and 5-HT 1D receptors are expressed in the human trigeminal ganglion and they are mainly localized in medium-sized cells and they seem to colocalize with CGRP, SP and NOS.

5-HT 7 receptor mRNA expression in human trigeminal ganglia

The present study investigated, by using the method of reverse transcriptase polymerase chain reaction, whether the 5-HT 7 receptor mRNA is expressed in human trigeminal ganglia. Trigeminal ganglia were excised post mortem from five human subjects. Oligonucleotide primers were selected based upon unique regions of complementary DNA sequence for the cloned human 5-HT 7 receptor. Sequence analysis revealed the presence of a single message that matched the sequence of the cloned human 5-HT 7 receptor. The present results may direct future efforts to determine the potential excitatory role of the 5-HT 7 receptor in the neuropathological events that develop in migraine headache.

Immunohistochemical localization of adenosine A 1 receptors in human brain regions

Adenosine exerts its physiological actions by binding to G-protein coupled receptors, four of which have been identified and cloned to date (A 1, A 2a, A 2b and A 3). Here we report the development of anti-human adenosine A 1, receptor anti-peptide polyclonal antibodies and their use to define the distribution of A 1, receptors in human brain regions, spinal cord and trigeminal ganglia by an immunohistochemical approach. Although the distribution of adenosine A 1, receptor and its mRNA in the human brain has been investigated in the past by autoradiography and in situ hybridization, this is the first demonstration of localization of the A 1, receptors by immunohistochemical means. Our localization data broadly agree with immunohistochemical data published for the human brain obtained using other experimental approaches. Furthermore, we have demonstrated the novel finding that abundant expression of the adenosine A 1, receptor protein occurs in the trigeminal ganglia, which may be suggestive of a role of this receptor in analgesia.

Varicella-zoster virus gene expression in latently infected and explanted human ganglia.

A consistent feature of varicella-zoster virus (VZV) latency is the restricted pattern of viral gene expression in human ganglionic tissues. To understand further the significance of this gene restriction, we used in situ hybridization (ISH) to detect the frequency of RNA expression for nine VZV genes in trigeminal ganglia (TG) from 35 human subjects, including 18 who were human immunodeficiency virus (HIV) positive. RNA for VZV gene 21 was detected in 7 of 11 normal and 6 of 10 HIV-positive subjects, RNA for gene 29 was detected in 5 of 14 normal and 11 of 11 HIV-positive subjects, RNA for gene 62 was detected in 4 of 10 normal and 6 of 9 HIV-positive subjects, and RNA for gene 63 was detected in 8 of 17 normal and 12 of 15 HIV-positive subjects. RNA for VZV gene 4 was detected in 2 of 13 normal and 4 of 9 HIV-positive subjects, and RNA for gene 18 was detected in 4 of 15 normal and 5 of 15 HIV-positive subjects. By contrast, RNAs for VZV genes 28, 40, and 61 were rarely or never detected. In addition, immunocytochemical analysis detected the presence of VZV gene 63-encoded protein in five normal and four HIV-positive subjects. VZV RNA was also analyzed in explanted fresh human TG and dorsal root ganglia from five normal human subjects over a period of up to 11 days in culture. We found a very different pattern of gene expression in these explants, with transcripts for VZV genes 18, 28, 29, 40, and 63 all frequently detected, presumably as a result of viral reactivation. Taken together, these data provide further support for the notion of significant and restricted viral gene expression in VZV latency.

Numbers of neurons and non-neuronal cells in human trigeminal ganglia

We analyzed both trigeminal ganglia of eight post-mortem human subjects for their content of neurons and non-neuronal cells using dissociated cell suspension techniques. Neuronal counts in each of the 16 ganglia ranged from 20,000 to 35,400, with an average of 27,400± 4800, and the estimated ratio of non-neuronal to neuronal cells was 100 to 1. Our numbers are comparable to the lower ranges obtained using different techniques in previous studies. [Neurol Res 2000; 22: 565-566]

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.

Functional calcitonin gene-related peptide type 1 and adrenomedullin receptors in human trigeminal ganglia, brain vessels, and cerebromicrovascular or astroglial cells in culture.

Calcitonin gene-related peptide (CGRP) and adrenomedullin (ADM) are potent dilators of human brain arteries, and they have been implicated in the neurogenic inflammation underlying migraine headache and in the evolution of stroke, respectively. However, little is known about the presynaptic and postsynaptic distribution of their respective receptors in the human cerebrovascular bed and trigeminovascular system. In the current study, the expression of mRNA for ADM and the two cloned human CGRP1 receptors (identified here as A-CGRP1 receptors [Aiyar et al., 1996] and K-CGRP1 receptors) [Kapas and Clark, 1995] were evaluated in human brain vessels and trigeminal ganglia. Further, the ability of CGRP and ADM to activate adenylate cyclase in cerebromicrovascular and astroglial cell cultures was determined, and the receptors involved were characterized pharmacologically. Isolated human pial vessels, intracortical microvessels, and capillaries, as well as cultures of brain endothelial (EC), smooth muscle (SMC), and astroglial (AST) cells, all expressed mRNA for the two cloned CGRP1 receptors; however, message for the K-CGRP1 receptor was barely detectable in microvascular tissues and cells. In contrast, only isolated capillaries and cultured AST exhibited message for the ADM receptor. In human trigeminal ganglia, mRNA for ADM and the two CGRP1 receptors was systematically present. The CGRP dose-dependently increased (up to 50-fold) cAMP formation in cell cultures, an effect significantly blocked by 0.1 to 10 micromol/L of the CGRP1 receptor antagonist CGRP8-37. The ADM receptor agonist, ADM13-52 (1 micromol/L), similarly increased cAMP production in all cell types, and this response was virtually abolished by 1 micromol/L CGRP8-37. Low concentrations (1 to 10 micromol/L) of the ADM receptor antagonist ADM22-52 blocked the ADM13-52-induced cAMP formation in AST (26% at 10 micromol/L, P < 0.05), whereas they potentiated this response in brain EC and SMC (40% and 100%, P < 0.001, respectively). Even at a higher dose (50 micromol/L), ADM22-52 inhibited the ADM13-52 effect in vascular cells (45%) much less effectively than in AST (95%). These results indicate that both CGRP and ADM can affect human brain vessels through a CGRP1 receptor, and they further suggest the presence of functional ADM receptors in human astroglial cells.

Glial cell line-derived neurotrophic factor-like immunoreactivity in human trigeminal ganglion and nucleus

Glial cell line-derived neurotrophic factor (GDNF) is shown by immunohistochemistry in human trigeminal sensory system from 22 weeks of gestation to adulthood. In the trigeminal ganglion, a distinct subpopulation of GDNF-positive neurones is observed, which amounts to about 15% at early pre-term and adult ages and peaks to around 30% at perinatal ages. Labelled neurones are mostly small- and medium-sized. Occasionally, Schwann and satellite cells are stained. GDNF/substance P (SP) and GDNF/calcitonin gene-related peptide (CGRP) double stained neurones occur at all ages examined, whereas GDNF/trkA coexistence can be observed in pre- and full-term newborns only. Centrally, GDNF-immunostained fibers and terminal-like structures are mainly restricted to the spinal trigeminal nucleus, where they are codistributed with SP and CGRP. In the subnucleus caudalis, positive neurones can also be observed both in the superficial laminae and in the magnocellular part, with higher frequency in adults. These results suggest that GDNF may play a functional role in human trigeminal primary sensory neurones throughout life and provide indication for its possible involvement in the regulation of pain-related neuronal circuits in human trigeminal sensory system.

Prevalence of varicella-zoster virus DNA in dissociated human trigeminal ganglion neurons and nonneuronal cells.

Previous analyses using in situ hybridization alone or together with PCR have yielded conflicting results regarding the cell type in which latent varicella-zoster virus (VZV) resides. We separated human trigeminal ganglia (TG) into neuronal and nonneuronal fractions, followed by primary and nested PCR to quantitate VZV DNA at the single cell level. Both TG from each of eight cadavers were dissociated and separated into neuronal and nonneuronal cell suspensions by differential filtration. Analysis of the neuron fraction (5,000 neurons per sample) revealed VZV DNA in 9 of 16 samples, with copy numbers ranging from 1 to 12, whereas only 2 of 16 nonneuronal cell samples were positive for VZV DNA, with 1 copy each. Further analysis of 10 samples of 100 neurons and the corresponding nonneuronal cell fractions from each TG of a single subject revealed VZV DNA in 3 of 10 samples of the left TG (range, 2 to 5 copies) and in 1 of 10 samples of the right TG (2 copies) but in none of the 20 nonneuronal cell fractions. These data indicate that latent VZV DNA is present primarily, if not exclusively, in neurons, at a frequency of two to five copies per latently infected neuron.

Messenger molecules and receptor mRNA in the human trigeminal ganglion

The presence and distribution of neuromessenger molecules and receptor mRNA in human trigeminal ganglion was studied with immunocytochemical, in situ hybridisation and RT-PCR techniques. Immunofluorescence staining revealed that calcitonin gene-related peptide (CGRP) immunoreactive (-ir) neurons occurred in high numbers, constituting 36–40% of all nerve cell bodies in the ganglion. Accordingly, in situ hybridisation demonstrated CGRP mRNA in a large portion of the trigeminal neurons. A small number of the nerve cell bodies showed substance P (SP)-ir, (18%), nitric oxide synthase (NOS)-ir (15%), and pituitary adenylate cyclase activating peptide (PACAP)-ir (20%). Double immunostaining revealed that only few CGRP-ir neurons also were NOS-ir (less than 5%). The C-terminal flanking peptide of neuropeptide Y, C-PON, was not visible in any of the nerve cell bodies studied. Agarose gel electrophoresis of the RT-PCR products from the ganglia demonstrated the presence of mRNA corresponding to CGRP 1, NPY Y 1 and Y 2, and VIP 1 receptors. These results suggest both sympathetic and parasympathetic influence on the activity in the trigeminal ganglion.

Presence of opioid receptor-like (ORL1) receptor mRNA splice variants in peripheral sensory and sympathetic neuronal ganglia

The expression of ORL1 receptor mRNA splice variants is determined in peripheral sensory and sympathetic ganglia and compared to mRNA expression for the three classic opioid receptor subtypes (mu, delta, and kappa) using the method of reverse transcription-polymerase chain reaction. ORL1, mu, delta and kappa receptor subtype mRNAs are present in human dorsal root ganglia (DRG) and trigeminal ganglia and rat DRG. ORL1, mu and delta receptor subtype mRNAs are present in rat superior cervical ganglia and only ORL1 and delta receptor mRNAs are present in rat lumbar sympathetic ganglia. Both the ORL1 mRNA splice variants are present in sensory and sympathetic ganglia, however, expression of the shorter ORL1 receptor mRNA dominates over expression of the longer splice variant in rat brain and DRG, whereas, expression of the longer splice variant is dominant in sympathetic ganglia.

Expression of calcitonin gene-related peptide-1 receptor mRNA in human tooth pulp and trigeminal ganglion

Numerous nerve fibres containing calcitonin gene-related peptide (CGRP) were found by immunocytochemistry in human molar pulp. These nerves were often seen around small blood vessels and as free endings without vascular contact. In the trigeminal ganglion a large number of CGRP-immunoreactive nerve-cell bodies, mostly of small to medium size, was encountered. Reverse transcriptase-polymerase chain reaction, using specific sense and antisense primers, detected mRNA expression of the human CGRP 1 receptor in the pulp tissue and the trigeminal ganglion. Thus, both CGRP-containing nerve fibres and CGRP 1 receptor mRNA are present in human tooth pulp, where they may be involved in the regulation of vascular tone and other local reactions to injury.