Yuan, Hongtu; Zhang, Weiwei; Li, Hao; Chen, Cheng; Liu, Huaxiang; Li, Zhenzhong
Studies have established that ethanol (EtOH) consumption results in damage to the peripheral nervous systems. Although the pathobiological mechanism is still unclear, oxidative stress is known to play an important role in EtOH-induced neurotoxicity. Because resveratrol (Res) is attracting increased attention due to its antioxidative properties, we investigated the neuroprotective efficacy of Res in ethanol-treated embryonic dorsal root ganglion (DRG) neurons in vitro. Organotypic DRG explants and a dispersed cell culture model were used to evaluate the effects of Res on EtOH-induced neurotoxicity. Res increased the number of extended nerve fibers and neurons that migrated from the DRG explants. Hoechst 33342 staining and terminal deoxynucleotidyl-transferase-mediated dUTP nick-end-labeling analysis showed that the EtOH-induced apoptosis was inhibited by Res. The effects of Res were blocked by the 5'-adenosine monophosphate-activated protein kinase inhibitor Compound C and the sirtuin 1 inhibitor nicotinamide. The elevation of oxidative/nitrosative stress, as measured by the amount of reactive oxygen species, malondialdehyde, nitrite, glutathione and superoxide dismutase activity, was also attenuated by Res. The data from the present study indicate that Res protects DRG neurons from EtOH-induced neurotoxicity. Res and its derivative may be effective for the treatment of diseases characterized by axonopathy and neuron loss induced by EtOH. Copyright © 2013 Elsevier Ltd. All rights reserved.
Lange, Miranda; Zeng, Yan; Knight, Andrew; Windebank, Anthony; Trushina, Eugenia
Changes in mitochondrial dynamics and function contribute to progression of multiple neurodegenerative diseases including peripheral neuropathies. The Seahorse Extracellular Flux XF24 analyzer provides a comprehensive assessment of the relative state of glycolytic and aerobic metabolism in live cells making this method instrumental in assessing mitochondrial function. One of the most important steps in the analysis of mitochondrial respiration using the Seahorse XF24 analyzer is plating a uniform monolayer of firmly attached cells. However, culturing of primary dorsal root ganglion (DRG) neurons is associated with multiple challenges, including their propensity to form clumps and detach from the culture plate. This could significantly interfere with proper analysis and interpretation of data. We have tested multiple cell culture parameters including coating substrates, culture medium, XF24 microplate plastics, and plating techniques in order to optimize plating conditions. Here we describe a highly reproducible method to obtain neuron-enriched monolayers of securely attached dissociated primary embryonic (E15) rat DRG neurons suitable for analysis with the Seahorse XF24 platform. PMID:23248613
Lange, Miranda; Zeng, Yan; Knight, Andrew; Windebank, Anthony; Trushina, Eugenia
Changes in mitochondrial dynamics and function contribute to progression of multiple neurodegenerative diseases including peripheral neuropathies. The Seahorse Extracellular Flux XF24 analyzer provides a comprehensive assessment of the relative state of glycolytic and aerobic metabolism in live cells making this method instrumental in assessing mitochondrial function. One of the most important steps in the analysis of mitochondrial respiration using the Seahorse XF24 analyzer is plating a uniform monolayer of firmly attached cells. However, culturing of primary dorsal root ganglion (DRG) neurons is associated with multiple challenges, including their propensity to form clumps and detach from the culture plate. This could significantly interfere with proper analysis and interpretation of data. We have tested multiple cell culture parameters including coating substrates, culture medium, XF24 microplate plastics, and plating techniques in order to optimize plating conditions. Here we describe a highly reproducible method to obtain neuron-enriched monolayers of securely attached dissociated primary embryonic (E15) rat DRG neurons suitable for analysis with the Seahorse XF24 platform.
Lüscher, C; Streit, J; Lipp, P; Lüscher, H R
1. The reliability of the propagation of action potentials (AP) through dorsal root ganglion (DRG) cells in embryonic slice cultures was investigated during repetitive stimulation at 1-20 Hz. Membrane potentials of DRG cells were recorded intracellularly while the axons were stimulated by an extracellular electrode. 2. In analogy to the double-pulse experiments reported previously, either one or two types of propagation failures were recorded during repetitive stimulation, depending on the cell morphology. In contrast to the double-pulse experiments, the failures appeared at longer interpulse intervals and usually only after several tens of stimuli with reliable propagation. 3. In the period with reliable propagation before the failures, a decrease in the conduction velocity and in the amplitude of the afterhyperpolarization (AHP), an increase in the total membrane conductance, and the disappearance of the action potential "shoulder" were observed. 4. The reliability of conduction during repetitive stimulation was improved by lowering the extracellular calcium concentration or by replacing the extracellular calcium by strontium. The reliability of conduction decreased by the application of cadmium, a calcium channel blocker, 4-amino pyridine, a fast potassium channel blocker, or apamin or muscarine, the blockers of calcium-dependent potassium channels. The reliability of conduction was not effected by blocking the sodium potassium pump with ouabain or by replacing extracellular sodium with lithium. 5. In the period with reliable propagation cadmium, apamin, and muscarine reduced the amplitude of the AHP. The shoulder of the action potential was more pronounced and not sensitive to repetitive stimulation when extracellular calcium was replaced by strontium. It disappeared when cadmium was applied. 6. In DRG somata changes of the intracellular Ca2+ concentration were monitored by measuring the fluorescence of the Ca2+ indicator Fluo-3 with a laser-scanning confocal
Marrs, Glen S; Spirou, George A
During early development, peripheral sensory systems generate physiological activity prior to exposure to normal environmental stimuli. This activity is thought to facilitate maturation of these neurons and their connections, perhaps even promoting efficacy or modifying downstream circuitry. In the mammalian auditory system, initial connections form at embryonic ages, but the functional characteristics of these early neural connections have not been assayed. We investigated processes of embryonic auditory development using a whole-head slice preparation that preserved connectivity between peripheral and brainstem stations of the auditory pathway. Transgenic mice expressing fluorescent protein provided observation of spiral ganglion and cochlear nucleus neurons to facilitate targeted electrophysiological recording. Here we demonstrate an apparent peripheral-to-central order for circuit maturation. Spiral ganglion cells acquire action potential-generating capacity at embryonic day 14 (E14), the earliest age tested, and action potential waveforms begin to mature in advance of comparable states for neurons of the ventral cochlear nucleus (VCN) and medial nucleus of the trapezoid body (MNTB). In accordance, auditory nerve synapses in the VCN are functional at E15, prior to VCN connectivity with the MNTB, which occurs at least 1 day later. Spiral ganglion neurons exhibit spontaneous activity at least by E14 and are able to drive third-order auditory brainstem neurons by E17. This activity precedes cochlear-generated wave activity by 4 days and ear canal opening by at least 2 weeks. Together, these findings reveal a previously unknown initial developmental phase for auditory maturation, and further implicate the spiral ganglion as a potential controlling centre in this process. PMID:22371481
Yin, Kathleen; Baillie, Gregory J
Background Dorsal root ganglion neuron-derived immortal cell lines including ND7/23 and F-11 cells have been used extensively as in vitro model systems of native peripheral sensory neurons. However, while it is clear that some sensory neuron-specific receptors and ion channels are present in these cell lines, a systematic comparison of the molecular targets expressed by these cell lines with those expressed in intact peripheral neurons is lacking. Results In this study, we examined the expression of RNA transcripts in the human neuroblastoma-derived cell line, SH-SY5Y, and two dorsal root ganglion hybridoma cell lines, F-11 and ND7/23, using Illumina next-generation sequencing, and compared the results with native whole murine dorsal root ganglions. The gene expression profiles of these three cell lines did not resemble any specific defined dorsal root ganglion subclass. The cell lines lacked many markers for nociceptive sensory neurons, such as the Transient receptor potential V1 gene, but expressed markers for both myelinated and unmyelinated neurons. Global gene ontology analysis on whole dorsal root ganglions and cell lines showed similar enrichment of biological process terms across all samples. Conclusions This paper provides insights into the receptor repertoire expressed in common dorsal root ganglion neuron-derived cell lines compared with whole murine dorsal root ganglions, and illustrates the limits and potentials of these cell lines as tools for neuropharmacological exploration. PMID:27130590
Jones, T A; Jones, S M; Paggett, K C
This study examined the nature of spontaneous discharge patterns in cochlear ganglion cells in embryonic day 13 (E13) to early E17 chicken embryos (stages 39-43). Neural recordings were made with glass micropipettes. No sound-driven activity was seen for the youngest embryos (maximum intensity 107 dB sound pressure level). Ganglion cells were labeled with biotinylated dextran amine in four embryos. In two animals, primary afferents projected to hair cells in the middle region along the length of the basilar papilla in which, in one cell, the terminals occupied a neural transverse position and, in the other, a more abneural location. Statoacoustic ganglion cells showing no spontaneous activity were seen for the first time in the chicken. The proportion of "silent" cells was largest at the youngest stages (stage 39, 67%). In active cells, mean spontaneous discharge rates [9.4 +/- 10.4 spikes (Sp)/sec; n = 44] were lower than rates for older embryos (19 +/- 17 Sp/sec) (Jones and Jones, 2000). Embryos at stages 39-41 evidenced even lower rates (4.2 +/- 5.0 Sp/sec). The most salient feature of spontaneous activity for stages 39-43 was a bursting discharge pattern in >75% of active neurons (33 of 44). Moreover, in 55% of these cells, there was a clear, slow, rhythmic bursting pattern. The proportion of cells showing rhythmic bursting was greatest at the youngest stages (39-42) and decreased to <30% at stage 43. Rate of bursting ranged from 1 to 54 bursts per minute. The presence of rhythmic bursting in cochlear ganglion cells at E13-E17 provides an explanation for the existence of such patterns in central auditory relays. The bursting patterns may serve as a patterning signal for central synaptic refinements in the auditory system during development.
Yu, Lingli; Ding, Yindi; Spencer, Ambre; Ma, Ji; Lu, Ruisheng; Rudkin, Brian B.; Yuan, Chonggang
This study examined the isolation and differentiation of dorsal root ganglion progenitor cells for therapeutic use in neurodegenerative diseases. Rat embryonic dorsal root ganglia progenitors were isolated and purified using the differential adhesion method combined with cytosine arabinoside treatment. After culture in serum-free medium supplemented with B27, basic fibroblast growth factor and epidermal growth factor, these cells remained viable and survived for more than 18 months in vitro. Most cells differentiated to neurons that were immunoreactive for gamma-aminobutyric acid and choline acetyltransferase as detected by immunohistochemical staining. In addition, nerve growth factor and neurotrophic tyrosine kinase receptor expression were also observed in dorsal root ganglion progenitors and differentiated cells. K252a, an inhibitor that blocks nerve growth factor-induced signaling, inhibited cell survival, suggesting the possible existence of a nerve growth factor autocrine loop in these proliferating cells. PMID:25745432
Gill, Katherine P.; Hung, Sandy S. C.; Sharov, Alexei; Lo, Camden Y.; Needham, Karina; Lidgerwood, Grace E.; Jackson, Stacey; Crombie, Duncan E.; Nayagam, Bryony A.; Cook, Anthony L.; Hewitt, Alex W.; Pébay, Alice; Wong, Raymond C. B.
Optic neuropathies are characterised by a loss of retinal ganglion cells (RGCs) that lead to vision impairment. Development of cell therapy requires a better understanding of the signals that direct stem cells into RGCs. Human embryonic stem cells (hESCs) represent an unlimited cellular source for generation of human RGCs in vitro. In this study, we present a 45-day protocol that utilises magnetic activated cell sorting to generate enriched population of RGCs via stepwise retinal differentiation using hESCs. We performed an extensive characterization of these stem cell-derived RGCs by examining the gene and protein expressions of a panel of neural/RGC markers. Furthermore, whole transcriptome analysis demonstrated similarity of the hESC-derived RGCs to human adult RGCs. The enriched hESC-RGCs possess long axons, functional electrophysiological profiles and axonal transport of mitochondria, suggestive of maturity. In summary, this RGC differentiation protocol can generate an enriched population of functional RGCs from hESCs, allowing future studies on disease modeling of optic neuropathies and development of cell therapies. PMID:27506453
Fan, Ni; Sikand, Parul; Donnelly, David F; Ma, Chao; Lamotte, Robert H
We investigated the effects of chronic compression (CCD) of the L3 and L4 dorsal root ganglion (DRG) on pain behavior in the mouse and on the electrophysiological properties of the small-diameter neuronal cell bodies in the intact ganglion. CCD is a model of human radicular pain produced by intraforaminal stenosis and other disorders affecting the DRG, spinal nerve, or root. On days 1, 3, 5, and 7 after the onset of compression, there was a significant decrease from preoperative values in the threshold mechanical force required to elicit a withdrawal of the foot ipsilateral to the CCD (tactile allodynia). Whole cell patch-clamp recordings were obtained, in vitro, from small-sized somata and, for the first time, in the intact DRG. Under current clamp, CCD neurons exhibited a significantly lower rheobase compared with controls. A few CCD but no control neurons exhibited spontaneous action potentials. CCD neurons showed an increase in the density of TTX-resistant and TTX-sensitive Na(+) current. CCD neurons also exhibited an enhanced density of voltage-dependent K(+) current, due to an increase in delayed rectifier K(+) current, without a change in the transient or "A" current. We conclude that CCD in the mouse produces a model of radicular pain, as we have previously demonstrated in the rat. While the role of enhanced K(+) current remains to be elucidated, we speculate that it represents a compensatory neuronal response to reduce ectopic or aberrant levels of neuronal activity produced by the injury.
Puil, E; Gimbarzevsky, B; Miura, R M
1. Membrane potentials of trigeminal root ganglion neurons were varied systematically by intracellular injections of long-lasting step currents to determine the voltage dependence of their membrane electrical properties. The complex impedance and impedance magnitude functions were first determined using oscillatory input currents superimposed on these step currents. 2. Systematic step variations in the membrane potential led to qualitative changes in the impedance magnitude functions. Depolarization of neurons exhibiting resonance at their initial resting membrane potentials resulted in a reduction in the resonance behavior. Hyperpolarization of these neurons to membrane potentials of about -80 to -90 mV led to a disappearance of the resonant peak but increased the maximum of the impedance magnitude. 3. The complex impedance data were fitted with a neuronal model derived from linearized Hodgkin-Huxley-like equations, yielding estimates for the membrane properties. The four parameters of the model were 1) a time invariant, resting membrane conductance, Gr, 2) a voltage- and time-dependent conductance, GL, 3) a time constant, tau u, for the unknown ionic channels that are activated by the 2- to 5-mV oscillatory perturbation of the stepped membrane potential, and 4) Ci, the input capacitance. 4. The results of the curve-fitting procedures suggested that all parameters depended on membrane voltage. The most voltage-dependent parameters were GL and tau u throughout a 25- to 30-mV range that was subthreshold to the production of action potentials. Both Gr and GL increased with subthreshold depolarization. 5. These impedance data suggest the very important role of the membrane potential of the trigeminal root ganglion neurons on their abilities to synthesize and filter inputted electrical signals.
Grava, André Luiz de Souza; Ferrari, Luiz Fernando; Parada, Carlos Amílcar; Defino, Helton Luiz Aparecido
To evaluate the hyperalgesia and histological abnormalities induced by contact between the dorsal root ganglion and the nucleus pulposus. Methods: Twenty Wistar rats were used, divided into two experimental groups. In one of the groups, a fragment of autologous nucleus pulposus was removed from the sacrococcygeal region and deposited on the L5 dorsal root ganglia. In the other group (control), a fragment of adipose tissue was deposited on the L5 dorsal root ganglia. Mechanical and thermal hyperalgesia was evaluated on the third day and the first, third, fifth and seventh weeks after the operation. A L5 dorsal root ganglion was removed in the first, third, fifth and seventh weeks after the operation for histological study using HE staining and histochemical study using specific labeling for iNOS. Results: Higher intensity of mechanical and thermal hyperalgesia was observed in the group of animals in which the nucleus pulposus was placed in contact with the dorsal root ganglion. In this group, the histological study showed abnormalities of the dorsal root ganglion tissue, characterized by an inflammatory process and axonal degeneration. The histopathological abnormalities of the dorsal root ganglion tissue presented increasing intensity with increasing length of observation, and there was a correlation with maintenance of the hyperalgesia observed in the behavioral assessment. Immunohistochemistry using specific labeling for iNOS in the group of animals in which the nucleus pulposus was placed in contact with the dorsal root ganglion showed higher expression of this enzyme in the nuclei of the inflammatory cells (glial cells) surrounding the neurons. Conclusion: Contact between the nucleus pulposus and the dorsal root ganglion induced mechanical and thermal hyperalgesia and caused histological abnormalities in the dorsal root ganglion components. These abnormalities were characterized by an inflammatory and degenerative process in the structures of the dorsal root
Yoshikawa, Masaaki; Murakami, Yuuki; Senzaki, Kouji; Masuda, Tomoyuki; Ozaki, Shigeru; Ito, Yoshiaki; Shiga, Takashi
Runt-related transcription factors (Runx) regulate the development of various cells. It has been reported that Runx1 and Runx3 are expressed in distinct subpopulations of primary sensory neurons in the dorsal root ganglion (DRG), and play important roles in the differentiation of nociceptive and proprioceptive neurons, respectively. In the present study, we examined the developmental changes of the expression of Runx1 and Runx3 in the mouse DRG during embryonic and postnatal stages. We found that the expression of Runx3 preceded that of Runx1, but dramatically decreased before birth, whereas the Runx1 expression was maintained during postnatal periods. These results suggest that roles of Runx1 and Runx3 may change dynamically in the differentiation and maturation of DRG neurons. In addition, several DRG neurons expressed both Runx1 and Runx3 throughout embryonic and postnatal stages and many Runx3-expressing DRG neurons coexpressed Runx1 at postnatal day 28. Double and triple labeling studies suggest that some of the Runx1/Runx3-double expressing neurons coexpressed TrkB, c-ret, and TrkC, which have been shown in the mechanoreceptive DRG neurons. These results suggest that Runx1/Runx3-double expressing neurons may represent mechanoreceptive properties in the DRG.
Hynds, Dianna L; Rangappa, Nagarathnamma; Ter Beest, Julia; Snow, Diane M; Rabchevsky, Alexander G
Transplantation of cellular populations to facilitate regrowth of damaged axons is a common experimental therapy for spinal cord injury. Schwann cells (SC) or microglia grafted into injury sites can promote axonal regrowth of central projections of dorsal root ganglion (DRG) sensory neurons. We sought to determine whether the addition of microglia or microglia-derived secretory products alters DRG axon regrowth upon cultures of SC. Rat DRG explants were grown on monolayers consisting of either SC, microglia, SC exposed to microglia-conditioned medium (MCM), or co-cultures with different relative concentrations of microglia. Image analysis revealed that, compared to SC alone, the extent of neurite outgrowth was significantly greater on SC-microglia co-cultures. Immunocytochemistry for extracellular matrix molecules showed that microglial cells stained positively for growth-promoting thrombospondin, whereas laminin and the inhibitory chondroitin sulfate proteoglycans (CSPGs) were localized primarily to SC. Notably, immunoreactivity for CSPGs appeared reduced in areas associated with DRG outgrowth in co-cultures and SC exposed to MCM. These results show that microglia or their secreted products can augment SC-mediated DRG regrowth in vitro, indicating that co-grafting SC with microglia provides a novel approach to augment sensory fiber regeneration after spinal cord injury.
Valeyev, A Y; Hackman, J C; Wood, P M; Davidoff, R A
1. Whole cell voltage-clamp studies of gamma-aminobutyric acid (GABA) receptors were performed on large (> 80 microns) cultured human dorsal root ganglion (DRG) neurons. 2. GABA and pentobarbital sodium when applied in micromolar concentrations evoked inward Cl- currents in DRG neurons voltage clamped at negative membrane potentials. 3. Diazepam (10 microM) and pentobarbital (10 microM) upmodulated the GABA current by approximately 149 and 168%, respectively. 4. The GABA currents in human DRG cells were unaffected by the classical GABA antagonists picrotoxin and bicuclline (100 microM). In contrast, the GABA responses evoked in adult rat DRG cells cultured in an identical manner were inhibited by both antagonists. The glycine receptor antagonist strychnine (100 microM) did not alter GABA currents in human DRG cells. 5. Human DRG cells did not respond to glycine (10-100 microM) or taurine (10-100 microM). The GABAB agonist baclofen had no effect on the holding current when patch pipettes were filled with 130 mM KCl. The GABAB antagonists saclofen applied either alone or with GABA was without effect. 6. The differences between the GABA receptors described here and GABA receptors in other species may reflect the presence of receptor subunits unique to human DRG cells.
Liu, Song; Bréjot, Thomas; Cressant, Arnaud; Bacci, Josette; Saïd, Gérard; Tadié, Marc; Heard, Jean Michel
Loss of dorsal root ganglion neuron, or injury to dorsal roots, induces permanent somatosensory defect without therapeutic option. We explored an approach to restoring hind limb somatosensory innervation after elimination of L4, L5 and L6 dorsal root ganglion neurons in rats. Somatosensory pathways were reconstructed by connecting L4, L5 and L6 lumbar dorsal roots to T10, T11 and T12 intercostal nerves, respectively, thus allowing elongation of thoracic ganglion neuron peripheral axons into the sciatic nerve. Connection of thoracic dorsal root ganglion neurons to peripheral tissues was documented 4 and 7 months after injury. Myelinated and unmyelinated fibers regrew in the sciatic nerve. Nerve terminations expressing calcitonin-gene-related-peptide colonized the footpad skin. Retrograde tracing showed that T10, T11 and T12 dorsal root ganglion neurons expressing calcitonin-gene-related-peptide or the neurofilament RT97 projected axons to the sciatic nerve and the footpad skin. Recording of somatosensory evoked potentials in the upper spinal cord indicated connection between the sciatic nerve and the central nervous system. Hind limb retraction in response to nociceptive stimulation of the reinnervated footpads and reversion of skin lesions suggested partial recovery of sensory function. Proprioceptive defects persisted. Delayed somatosensory reinnervation of the hind limb after destruction of lumbar dorsal root neurons in rats indicates potential approaches to reduce chronic disability after severe injury to somatosensory pathways.
Ogihara, Yuuki; Masuda, Tomoyuki; Ozaki, Shigeru; Yoshikawa, Masaaki; Shiga, Takashi
Somatosensation is divided into proprioception and cutaneous sensation. Dorsal root ganglion (DRG) neurons project their fibers toward peripheral targets including muscles and skin, and centrally to the spinal cord. Proprioceptive DRG neurons transmit information from muscle spindles and Golgi tendon organs to the spinal cord. We previously showed that Runt-related transcription factor 3 (Runx3) is expressed in these neurons and their projections to the ventral spinal cord and muscle spindles are lost in Runx3-deficient (Runx3(-/-) ) mouse embryos. Although Runx3 is likely to contribute to the fate decision and projection of proprioceptive DRG neurons, the precise roles for Runx3 in these phenomena are unknown. To identify genes regulated by Runx3 in embryonic DRGs, we performed microarray analyses using cDNAs isolated from wild-type and Runx3(-/-) DRGs of embryonic day (E) 12.5 and selected two transcript variants of the tyrosine kinase receptor C (TrkC) gene. These variants, Ntrk3 variant 1 (Ntrk3-v1) and variant 2 (Ntrk3-v2), encode full-length and truncated receptors of neurotrophin-3, respectively. Using double in situ hybridization, we found that most of Ntrk3-v1 mRNA expression in E14.5 DRGs depended on Runx3 but that more than half of Ntrk3-v2 mRNA one were expressed in a Runx3-independent manner. Furthermore, our data revealed that the rate of Ntrk3-v1 and Ntrk3-v2 colocalization in DRGs changed from E14.5 to E18.5. Together, our data suggest that Runx3 may play a crucial role in the development of DRGs by regulating the expression of Ntrk3 variants and that DRG neurons expressing Ntrk3-v1 but not Ntrk3-v2 may differentiate into proprioceptive ones.
Maino, Paolo; Koetsier, Eva; Kaelin-Lang, Alain; Gobbi, Claudio; Perez, Roberto
Small fiber neuropathy is a disorder of the peripheral nerves with typical symptoms of burning, sharp, and shooting pain and sensory disturbances in the feet. Pain treatment depends principally on the underlying etiology with concurrent administration of antidepressants, anticonvulsants, opioids, and topical treatments like capsaicin and local anesthetics. However, treatments for pain relief in these patients frequently fail. We describe the first case of intractable painful small fiber neuropathy of the foot successfully treated with spinal cord stimulation of the left L5 dorsal root ganglion.A 74-year-old man presented at our clinic with severe intractable pain, dysesthesia, and allodynia of the left foot caused by idiopathic small fiber neuropathy, confirmed by skin biopsy. His pain score was 8 on a standard 0 - 10 numeric rating scale. As the pain was not satisfactorily controlled by conventional therapy, dorsal root ganglion stimulation was proposed to the patient and, after informed consent, a specifically designed percutaneous stimulation lead was placed over the left L5 dorsal root ganglion and connected to an external neurostimulator. After a positive trial of 10 days, a permanent neurostimulator was implanted. Twenty months post-implantation the patient continued to experience stimulation-induced paresthesia covering the entire pain area and reported a pain rating of 4.Results from the case report demonstrate that the dorsal root ganglion is a promising neural stimulation target to treat neuropathic pain due to intractable small fiber neuropathy. Prospective controlled studies are warranted to confirm the efficacy of this treatment as an option for the aforementioned condition.Key words: Dorsal root ganglion stimulation, small fiber neuropathy, neuropathic pain.
Han, Chongyang; Hoeijmakers, Janneke G J; Liu, Shujun; Gerrits, Monique M; te Morsche, Rene H M; Lauria, Giuseppe; Dib-Hajj, Sulayman D; Drenth, Joost P H; Faber, Catharina G; Merkies, Ingemar S J; Waxman, Stephen G
Patients with small fibre neuropathy typically manifest pain in distal extremities and severe autonomic dysfunction. However, occasionally patients present with minimal autonomic symptoms. The basis for this phenotypic difference is not understood. Sodium channel Na(v)1.7, encoded by the SCN9A gene, is preferentially expressed in the peripheral nervous system within sensory dorsal root ganglion and sympathetic ganglion neurons and their small diameter peripheral axons. We recently reported missense substitutions in SCN9A that encode functional Na(v)1.7 variants in 28% of patients with biopsy-confirmed small fibre neuropathy. Two patients with biopsy-confirmed small fibre neuropathy manifested minimal autonomic dysfunction unlike the other six patients in this series, and both of these patients carry the Na(v)1.7/R185H variant, presenting the opportunity to compare variants associated with extreme ends of a spectrum from minimal to severe autonomic dysfunction. Herein, we show by voltage-clamp that R185H variant channels enhance resurgent currents within dorsal root ganglion neurons and show by current-clamp that R185H renders dorsal root ganglion neurons hyperexcitable. We also show that in contrast, R185H variant channels do not produce detectable changes when studied by voltage-clamp within sympathetic neurons of the superior cervical ganglion, and have no effect on the excitability of these cells. As a comparator, we studied the Na(v)1.7 variant I739V, identified in three patients with small fibre neuropathy characterized by severe autonomic dysfunction as well as neuropathic pain, and show that this variant impairs channel slow inactivation within both dorsal root ganglion and superior cervical ganglion neurons, and renders dorsal root ganglion neurons hyperexcitable and superior cervical ganglion neurons hypoexcitable. Thus, we show that R185H, from patients with minimal autonomic dysfunction, does not produce detectable changes in the properties of
Godel, Tim; Bäumer, Philipp; Pham, Mirko; Köhn, Anja; Muschol, Nicole; Kronlage, Moritz; Kollmer, Jennifer; Heiland, Sabine; Bendszus, Martin; Mautner, Victor-Felix
To evaluate functional and morphometric magnetic resonance neurography of the dorsal root ganglion and peripheral nerve segments in patients with Fabry painful neuropathy. In this prospective study, the lumbosacral dorsal root ganglia and proximal peripheral nerve segments of the lower extremity were examined in 11 male patients with Fabry disease by a standardized 3T magnetic resonance neurography protocol. Volumes of L3 to S2 dorsal root ganglia, perfusion parameters of L5-S1 dorsal root ganglia and the spinal nerve L5, and the cross-sectional area of the proximal sciatic nerve were compared to healthy controls. Dorsal root ganglia of patients with Fabry disease were symmetrically enlarged by 78% (L3), 94% (L4), 122% (L5), 115% (S1), and 119% (S2) (p < 0.001). In addition, permeability of the blood-tissue interface was decreased by 53% (p < 0.001). This finding was most pronounced in the peripheral zone of the dorsal root ganglion containing the cell bodies of the primary sensory neurons (p < 0.001). Spinal nerve permeability showed no difference between patients with Fabry disease and controls (p = 0.7). The sciatic nerve of patients with Fabry disease at the thigh level showed an increase in cross-sectional area by 48% (p < 0.001). Patients with Fabry disease have severely enlarged dorsal root ganglia with dysfunctional perfusion. This may be due to glycolipid accumulation in the dorsal root ganglia mediating direct neurotoxic effects and decreased neuronal blood supply. These alterations were less pronounced in peripheral nerve segments. Thus, the dorsal root ganglion might play a key pathophysiologic role in the development of neuropathy and pain in Fabry disease. © 2017 American Academy of Neurology.
Sharp, A A; Fedorovich, Y
Our understanding of the role of somatosensory feedback in regulating motility during chicken embryogenesis and fetal development in general has been hampered by the lack of an approach to selectively alter specific sensory modalities. In adult mammals, pyridoxine overdose has been shown to cause a peripheral sensory neuropathy characterized by a loss of both muscle and cutaneous afferents, but predominated by a loss of proprioception. We have begun to explore the sensitivity of the nervous system in chicken embryos to the application of pyridoxine on embryonic days 7 and 8, after sensory neurons in the lumbosacral region become post-mitotic. Upon examination of the spinal cord, dorsal root ganglion and peripheral nerves, we find that pyridoxine causes a loss of neurotrophic tyrosine kinase receptor type 3-positive neurons, a decrease in the diameter of the muscle innervating nerve tibialis, and a reduction in the number of large diameter axons in this nerve. However, we found no change in the number of Substance P or calcitonin gene-related peptide-positive neurons, the number of motor neurons or the diameter or axonal composition of the femoral cutaneous nerve. Therefore, pyridoxine causes a peripheral sensory neuropathy in embryonic chickens largely consistent with its effects in adult mammals. However, the lesion may be more restricted to proprioception in the chicken embryo. Therefore, pyridoxine lesion induced during embryogenesis in the chicken embryo can be used to assess how the loss of sensation, largely proprioception, alters spontaneous embryonic motility and subsequent motor development.
Hervey-Jumper, Shawn L; McKeever, Paul E; Gebarski, Stephen S; Muraszko, Karin M; Maher, Cormac O
Infantile hemangiomas are tumors commonly seen in children. Few authors have reported infantile hemangiomas affecting the CNS, and there are no prior reports detailing spontaneous resolution of a histologically proven juvenile hemangioma within a dorsal root ganglion. The authors report the case of a newborn boy with a large cutaneous hemangioma in the midline of his back. Spinal MR images were obtained to rule out associated spinal cord tethering, and an intradural spinal lesion was unexpectedly discovered. Biopsy revealed an intradural infantile hemangioma within the dorsal root ganglion, and, based on this diagnosis, no resection was performed. Sixteen months following the biopsy, the cutaneous hemangioma had become involuted and the intradural hemangioma had completely resolved. The behavior of the intradural component in this case follows the natural history of many cutaneous infantile hemangiomas.
Hoshino, Natalia; Vatterott, Phillip; Egwiekhor, Amina; Rochlin, M. William
Geniculate axons are initially guided to discrete epithelial placodes in the lingual and palatal epithelium that subsequently differentiate into taste buds. In vivo approaches show that brain-derived neurotrophic factor (BDNF) mRNA is concentrated in these placodes, that BDNF is necessary for targeting taste afferents to these placodes, and that BDNF misexpression disrupts guidance. We used an in vitro approach to determine whether BDNF may act directly on geniculate axons as a trophic factor and as an attractant, and whether there is a critical period for responsiveness to BDNF. We show that BDNF promotes neurite outgrowth from geniculate ganglion explants dissected from embryonic day (E) 15, E18, infant, and adult rats cultured in collagen gels, and that there is a concentration optimum for neurite extension. Gradients of BDNF derived from slow-release beads caused the greatest bias in neurite outgrowth at E15, when axons approach the immature gustatory papillae. Further, neurites advanced faster toward the BDNF bead than away from it, even if the average amount of neurotrophic factor encountered was the same. We also found that neurites that contact BDNF beads did not advance beyond them. At E18, when axons would be penetrating pregustatory epithelium in vivo, BDNF continued to exert a tropic effect on geniculate neurites. However, at postnatal and adult stages, the influence of BDNF was predominantly trophic. Our data support a role for BDNF acting as an attractant for geniculate axons during a critical period that encompasses initial targeting but not at later stages. PMID:20639634
Yu, Hongwei; Fischer, Gregory; Hogan, Quinn H
Transferring genetic molecules into the peripheral sensory nervous system to manipulate nociceptive pathophysiology is a powerful approach for experimental modulation of sensory signaling and potentially for translation into therapy for chronic pain. This can be efficiently achieved by the use of recombinant adeno-associated virus (rAAV) in conjunction with nociceptor-specific regulatory transgene cassettes. Among different routes of delivery, direct injection into the dorsal root ganglia (DRGs) offers the most efficient AAV-mediated gene transfer selectively into the peripheral sensory nervous system. Here, we briefly discuss the advantages and applications of intraganglionic microinjection, and then provide a detailed approach for DRG injection, including a list of the necessary materials and description of a method for performing DRG microinjection experiments. We also discuss our experience with several adeno-associated virus (AAV) options for in vivo transgene expression in DRG neurons.
Fadda, A.; Bärtschi, M.; Hemphill, A.; Widmer, H. R.; Zurbriggen, A.; Perona, P.; Vidondo, B.; Oevermann, A.
Neurological disorders in ruminants have an important impact on veterinary health, but very few host-specific in vitro models have been established to study diseases affecting the nervous system. Here we describe a primary neuronal dorsal root ganglia (DRG) culture derived from calves after being conventionally slaughtered for food consumption. The study focuses on the in vitro characterization of bovine DRG cell populations by immunofluorescence analysis. The effects of various growth factors on neuron viability, neurite outgrowth and arborisation were evaluated by morphological analysis. Bovine DRG neurons are able to survive for more than 4 weeks in culture. GF supplementation is not required for neuronal survival and neurite outgrowth. However, exogenously added growth factors promote neurite outgrowth. DRG cultures from regularly slaughtered calves represent a promising and sustainable host specific model for the investigation of pain and neurological diseases in bovines. PMID:27936156
Parke, Wesley Wilkin; Whalen, Joseph Leo
A descriptive anatomic investigation of the vasculature of the dorsal root ganglions. To determine whether the blood supply of the various spinal ganglions is sufficiently consistent to derive a "generic" description and illustration that would be applicable to all spinal levels, and to ascertain whether this vascular pattern is inherently predisposed to the development of a closed compartment syndrome. The few previous descriptions of spinal ganglionic vasculature do not include photographic evidence showing uniformity in the arterial distribution plan at all ganglionic levels. The venous drainage, although verbally reconstructed from microscopic sections, lacks any indication of its probable role in the etiology of a compartment syndrome. Three perinatal cadavers received latex/India ink injections, and their removed radiculomedullary systems were cleared, transilluminated, and macroscopically photographed. Paravertebral sections were grossly removed from the spines of two adult anatomic cadavers and received retrograde venous injections of a fine suspension of barium sulfate. The intervertebral foraminal tissues were then dissected from the bone, and radiographs of them were made. For comparative reference, a nerve root/ganglion complex of a rabbit was arterially injected with a more dilute preparation of the latex/India ink suspension. Macroscopic photographs of perinatal dorsal root ganglions showed that the pattern of the intraganglionic arterial distribution was sufficiently consistent to allow a graphic rendering and labeling of a "generic" ganglion. The series of incomplete retrograde venous injections adequately indicated the pressure labile location of a periganglionic venous plexus. The common development, structure, and function of the human dorsal root ganglions have resulted in the evolution of a uniform nutritional vascular pattern that can be conceptualized in a single visual image. Its plan of a primarily internal arterialization with a
Isstas, Marcel; Teichert, Manuel; Bolz, Jürgen; Lehmann, Konrad
The maturation of cortical inhibition provided by parvalbumin-containing basket cells derived from the medial ganglionic eminence (MGE) is a key event in starting the enhanced visual cortical plasticity during the critical period. Although it is generally assumed that a further increase in inhibition closes the critical period again, it was recently shown that embryonic interneurons derived from the MGE can induce an additional, artificial critical period when injected into the visual cortex of young mice. It has, however, remained open whether this effect was indeed specific for MGE-derived cells, and whether critical period-like plasticity could also be induced in fully adult animals. To clarify these issues, we injected explants from either the MGE or the caudal ganglionic eminence (CGE) into the visual cortices of fully adult mice, and performed monocular deprivation 33 days later for 4 days. Animals implanted with MGE cells, but not with CGE cells, showed marked ocular dominance plasticity. Immunohistochemistry confirmed that the injected cells from both sources migrated far in the host cortex, that most developed into neurons producing GABA, and that only cells from the MGE expressed parvalbumin. Thus, our results confirm that the plasticity-inducing effect of embryonic interneurons is specific for cells from the MGE, and is independent of the host animal's age.
Huang, M H; Horackova, M; Negoescu, R M; Wolf, S; Armour, J A
To determine the response characteristics of dorsal root ganglion neurones that may serve sensory functions during myocardial ischaemia. Extracellular recordings were made from 54 spontaneously active and 5 normally quiescent dorsal root ganglion neurones (T2-T5) in 22 anaesthetized open-chest dogs under control conditions and during epicardial mechanical or chemical stimulation and myocardial ischaemia. The activity of 78% of spontaneously active and all quiescent neurones with left ventricular sensory fields was modified by left ventricular ischaemia. Forty-six spontaneously active neurones (85%) were polysensory with respect to mechanical and chemical stimuli. The 5 quiescent neurones responded only to chemical stimuli. Spontaneously active neurones associated with left ventricular mechanosensory endings (37 neurones) generated four different activity patterns in response to similar mechanical stimuli (high or low pressure active, high-low pressure active, high-low pressure inactive). A fifth group generated activity which was not related to chamber dynamics. Adenosine, adenosine 5'-triphosphate, substance P and bradykinin modified 72, 61, 65 and 63% of the spontaneously active neurones, respectively. Maximum local mechanical or chemical stimuli enhanced activity to similar degrees, as did ischaemia. Each ischaemia-sensitive neurone displayed unique activity patterns in response to similar mechanical or chemical stimuli. Most myocardial ischemia-sensitive dorsal root ganglion neurones associated with epicardial neurites sense mechanical and multiple chemical stimuli, a small population sensing only mechanical or chemical stimuli. Activity patterns generated by these neurones depend on their primary sensory characteristics or those of other neurones that may converge on them, as well as the type and magnitude of the stimuli that impinge upon their sensory fields, both normally and during ischaemia.
Kolesár, Dalibor; Kolesárová, Mária; Kyselovič, Ján
The main aim of the present review is to provide at first a short survey of the basic anatomical description of sensory ganglion neurons in relation to cell size, conduction velocity, thickness of myelin sheath, and functional classification of their processes. In addition, we have focused on discussing current knowledge about the distribution pattern of neuronal nitric oxide synthase containing sensory neurons especially in the dorsal root ganglia in different animal species; hence, there is a large controversy in relation to interpretation of the results dealing with this interesting field of research.
Dolphin, A. C.; Scott, R. H.
Voltage-dependent inward calcium currents (ICa) activated in cultured rat dorsal root ganglion neurones were reversibly reduced in a dose-dependent manner by (-)-baclofen (10 microM to 100 microM). Baclofen (100 microM) reduced the calcium-dependent slow outward potassium current (IK(Ca)). This current was abolished in calcium-free medium and by 300 microM cadmium chloride. The action of baclofen on IK(Ca) was reduced when the calcium concentration in the medium was increased from 5 mM to 30 mM. The calcium independent fast transient voltage-dependent outward current (IK(Vt] was also reduced by baclofen; this effect remained present when Ca2+-free medium was used to prevent contamination by IK(Ca). 4-Aminopyridine (500 microM) reduced IK(Vt) and induced a small increase in ICa. The action of baclofen on ICa was partially antagonized by 4-aminopyridine. GABAB receptor-mediated inhibition of ICa in cultured rat dorsal root ganglion neurones involves a direct mechanism rather than resulting indirectly from an increase in the residual outward potassium currents activated by depolarization. The reduction in ICa by baclofen was variable and dependent on the amplitude of control ICa, larger currents being more resistant to the baclofen-induced inhibition. PMID:2423173
Yang, Xiangdong; Liu, Zhen; Li, Zhenzhong
Background: Norepinephrine (NE) is a key neurotransmitter that functionally activates adrenoreceptors expressed in sympathetic neurons. Functional α1-adrenoreceptors are also expressed in dorsal root ganglion (DRG) primary sensory neurons and regulate neurogenic inflammation and nociceptive responses. Galanin is involved in inflammation and nociception. It has been suggested that galanin receptor (GalR) 1 and GalR3 activation induces analgesia at the level of the spinal cord, while activation of GalR2 has a pronociceptive role in the periphery. Whether activation or inhibition of α-adrenoreceptors influences galanin expression remains unknown. Objective: The aim of the present study was to investigate whether the α-adrenoreceptor agonist NE, the α1-adrenoreceptor antagonist prazosin, and the α2-adrenoreceptor antagonist yohimbine affect galanin expression in primary cultured DRG neurons. Methods: DRG was dissected from 240 embryonic 15-day-old Wistar rats, cultured as dissociated cells for 2 days, and then exposed to NE (10-4 mol/L) for another 4 days. In the NE + prazosin group and the NE + yohimbine group, DRG neurons were pretreated with prazosin (10-6 mol/L) and yohimbine (10-5 mol/L), respectively, 10 minutes prior to the NE challenge. The neurons cultured continuously in media served as the controls. All of the cultured samples were processed to detect galanin mRNA and galanin peptide expression by reverse transcriptase-polymerase chain reaction and Western blot, respectively. Five samples were tested for each procedure. Results: Forty samples were prepared for this study and included in the analysis. After 4 days of incubation, mean (SD) galanin mRNA/β-actin mRNA concentration ratio was significantly increased with NE compared with controls (0.3349 [0.0413] vs 0.2411 [0.0519]; P < 0.05). Pretreatment with prazosin seemed to block the effects of NE (0.2522 [0.0496]; P < 0.05 vs NE), while yohimbine did not appear to significantly alter the effects of NE
Ingole, I V; Ghosh, S K
With an ever increasing number of cell phone users since late twentieth [corrected] century, magnitude of the problem of exposure to radiation emitted by cell phone is self evident. Extensive research had been devoted to incriminate or absolve it as a health hazard. Radiofrequency radiation emitted by cell phone had been stated to be a potent carcinogen, cytotoxic, genotoxic, mutagenic and neurobehavioral teratogen. Its effect on the brain had been a subject of extensive research evidently due to its proximity to the user's brain. While considering the biological effects of radiofrequency radiation, its intensity, frequency and the duration of exposure are important determinants. Nevertheless the results of these different studies have not been unequivocal. Considering the contradictory reports, the present work was undertaken to study the effect of such an exposure on the developing neural tissue of chick embryo. The processes of cell division and differentiation are fundamental to the development of any living being and are a sensitive index of any insult sustained at this stage. Neurons of dorsal root ganglion were selected for the present study as these ganglia were fully differentiated as early as fourth day of embryonic life. By varying duration of exposure, the embryos were exposed to different doses of radiation, sacrificed at different periods of incubation and subjected to histological processing. On light microscopic study it was observed that developing neurons of dorsal root ganglion suffered a damage which was dose dependent and persisted in spite of giving the exposure-free period between two exposures.
Sokolowski, Bernd H A
The acquisition of ion channels is critical to the formation of neuronal pathways in the peripheral and central nervous systems. This study describes the different types of inward currents (Ii) recorded from the soma of isolated cochleovestibular ganglion (CVG) cells of the embryonic chicken, Gallus gallus. Cells were isolated for whole-cell tight-seal recording from embryonic day (ED) 3, an age when the CVG is a cell cluster, to ED 9, an age when the cochlear and vestibular ganglia (CG, VG) are distinct structures. Results show Na+ and Ca2+ currents (INa and ICa) are acquired by ED 3, although INa dominates with greater density levels that peak by ED 6-7 in VG neurons. In the CG, INa acquisition is slower, reaching peak values by ED 8-9. Isolation of ICa, using Ba2+ as the charge carrier, showed both transient (IBaT)- and sustained (IBaL)-type currents on ED 3. Unlike INa, IBa density varied with age and ganglion. Total IBa increased steadily, showing a decline only in CG cells on ED 8-9 as a result of a decrease in IBaT. IBaL density increased over time, reaching a maximum on ED 6-7 in VG cells, followed by a decline on ED 8-9. In comparison, IBaL in CG neurons, did not increase significantly beyond mean values measured on ED 5. The early onset of these currents and the variations in Ca2+ channel expression between the ganglia suggests that intracellular signals relevant to phenotypic differentiation begin within these early time frames. Copyright 2006 Wiley-Liss, Inc.
Bernd, P.; Represa, J. )
We have investigated the possibility that nerve growth factor (NGF) may play a role in the development of the inner ear. Primordia of the inner ear, the otic vesicle (OV) and cochleovestibular ganglion (CVG), were isolated from 72-hr (stage 19-20) quail embryos and examined for the presence of NGF receptors. Quantitative binding studies revealed that both OV and CVG exhibited specific 125I-NGF binding; levels of nonspecific binding were 6 to 26% of total binding. Scatchard analysis yielded a linear plot, indicating the presence of a single class of NGF receptor. The average binding constant (Kd) was 8.0 nM for OV and 8.6 nM for CVG, corresponding to the low affinity (site II) NGF receptor. Examination of light microscopic radioautographs indicated that most of the specific 125I-NGF binding was located in the ventromedial wall of the OV, with little or no binding in the lateral wall and endolymphatic primordia. These studies were corroborated by microdissection of OV, in which 70% of the radioactivity was found to be localized in the medial half of the OV. In CVG, specific 125I-NGF binding was more concentrated in the cochlear portion of the ganglion, with silver grains primarily over areas containing support cells and immature neurons. Quantitative binding studies with isolated cochlear and vestibular ganglia obtained from 144-hr (stage 29-30) quail embryos revealed that the cochlear ganglion exhibited three times more specific 125I-NGF binding than the vestibular ganglion. The presence of NGF receptors on OV and CVG suggests that these structures are responsive to and/or dependent upon NGF. The following paper examines the question of whether NGF serves either as a mitogen, a survival factor, or a differentiation factor in this system.
Siemionow, Kris; Weinstein, James N; McLain, Robert F
The membrane, nucleus, and cytoplasmic contents of satellite cells were evaluated using a transmission electron microscope. To delineate satellite cell morphometries. The role of the satellite support cells associated with the neuronal cell bodies remains poorly understood. Previous research has identified one type of satellite support cells. Dorsal root ganglions were excised from 10 adult New Zealand White rabbits. Sections from L2-L5 ganglions were prepared, cut, and analyzed under a transmission electron microscope. A total of 190 neurons and their associated satellite cells were selected for analysis. Three subgroups of satellite cells were identified. The two predominant subgroups consisted of previously described satellite cells. The third subgroup consisted of highly complex and unusual cells. Nineteen satellite cells (4%) did not conform to any previous description of glial cells. Cells were characterized by larger nuclei, with numerous inclusions, and by extensively convoluted reflections of the cellular membrane. These cells were "perched" or "piggy-backed" on top of a convoluted and multilayered cytoplasmic sheet. A new type of support cell representing a different cell line or a highly adapted cell with specific functional capacities was identified.
Milikan, J M; Bolsover, S R
We have used fluorescently labelled calmodulins to probe the activity of calmodulin in living dorsal root ganglion cells. Calmodulin labelled with the fluorophore 5-([4,6 dichlorotriazin-2yl]amino)-fluorescein (FL-CaM) does not change its fluorescence when it binds calcium, while calmodulin labelled at lysine 75 with 2-chloro-(6-(4-N,N-diethylamino-phenyl)-1,4,5-triazin-4-yl (TA-CaM), an environment-sensitive probe, increases its fluorescence when it binds calcium. We micro-injected FL-CaM or TA-CaM into rat dorsal root ganglion cells and found that both probes localise to the cell nucleus. In contrast, endogenous cellular calmodulin, in dorsal root ganglion cells as in hippocampal neurones, is predominantly cytosolic unless the neurones are depolarised, then it moves to the nucleus. FL-CaM and TA-CaM, introduced into dorsal root ganglion cells via a patch pipette, also immediately move to the nucleus, indicating that the nuclear localisation is a property of the labelled calmodulins. Although the subcellular distribution of FL-CaM and TA-CaM does not necessarily match that of endogenous calmodulin, we show that FL-CaM can be used as a control for TA-CaM when studying calmodulin activation in different cellular compartments.
Redox modulation of fast inactivation has been described in certain cloned A-type voltage-gated K(+) (Kv) channels in expressing systems, but the effects remain to be demonstrated in native neurons. In this study, we examined the effects of cysteine-specific redox agents on the A-type K(+) currents in acutely dissociated small diameter dorsal root ganglion (DRG) neurons from rats. The fast inactivation of most A-type currents was markedly removed or slowed by the oxidizing agents 2,2'-dithio-bis(5-nitropyridine) (DTBNP) and chloramine-T. Dithiothreitol, a reducing agent for the disulfide bond, restored the inactivation. These results demonstrated that native A-type K(+) channels, probably Kv1.4, could switch the roles between inactivating and non-inactivating K(+) channels via redox regulation in pain-sensing DRG neurons. The A-type channels may play a role in adjusting pain sensitivity in response to peripheral redox conditions.
Bei, Wei-Jian; Xu, An-Long; Li, Chu-Yuan; Cabot, Peter J; Hermanussen, Siobhan
To investigate the effects of flavonoids from the leaves of Diospyros kaki L (FLDK) on the adhesion between the lymphocyte and the neurone. Centrifugal assay for fluorescence-bsaed cell adhesion was used to assay the adhesion between the lymphocyte and the dorsal root ganglion (DRG). The adhesion was significantly suppressed in the presence of FLDK dose-dependently at 5, 25 microg/mL concentration. FLDK was also effective to inhibit the adhesion under the challenge of ICAM-1 by 28.5% and 50.1%, respectively. Furthermore, FLDK enforced the inhibition of anti-NCAM antibody on the lymphocyte adhesion to DRG cells. FLDK might contribute to the prevention and treatment of the inflammation injury under neuron insult such as ischemia/reperfusion, neurotrauma and other neurodegenerative disease by inhibiting the adhesion between lymphocytes and neurons.
Liem, Liong; van Dongen, Eric; Huygen, Frank J; Staats, Peter; Kramer, Jeff
Chronic neuropathic pain is a widespread problem with negative personal and societal consequences. Despite considerable clinical neuroscience research, the goal of developing effective, reliable, and durable treatments has remained elusive. The critical role played by the dorsal root ganglion (DRG) in the induction and maintenance of chronic pain has been largely overlooked in these efforts, however. It may be that, by targeting this site, robust new options for pain management will be revealed. This review summarizes recent advances in the knowledge base for DRG-targeted treatments for neuropathic pain:• Pharmacological options including the chemical targeting of voltage-dependent calcium channels, transient receptor potential channels, neurotrophin production, potentiation of opioid transduction pathways, and excitatory glutamate receptors.• Ablation or modulation of the DRG via continuous thermal radiofrequency and pulsed radiofrequency treatments.• Implanted electrical neurostimulator technologies.• Interventions involving the modification of DRG cellular function at the genetic level by using viral vectors and gene silencing methods.
Bischoff, Ulrike; Bräu, Michael E; Vogel, Werner; Hempelmann, Gunter; Olschewski, Andrea
Hyperpolarizing voltage steps evoke slowly activating inward currents in a variety of neurones and in cardiac cells. This hyperpolarization-activated inward current (Ih) is thought to play a significant role in cell excitability, firing frequency, or in setting of the resting membrane potential in these cells. We studied the effects of lidocaine, mepivacaine, QX-314 and bupivacaine as well as its enantiomers on Ih in the membrane of dorsal root ganglion neurones (DRG). The patch-clamp technique was applied to small dorsal root ganglion neurones identified in 200 μM thin slices of young rat DRGs. Under voltage-clamp conditions, the whole-cell Ih current was recorded in the presence of different concentrations of the local anaesthetics. In current-clamp mode the resting membrane potential and the voltage response of DRG neurones to injected current pulses were investigated. Ih was reversibly blocked by bupivacaine, lidocaine and mepivacaine applied externally in clinically relevant concentrations. Concentration–response curves gave half-maximum inhibiting concentrations of 55, 99 and 190 μM, respectively. Bupivacaine block of the Ih current was not stereoselective. No significant effect was observed when QX-314 was applied to the external surface of the membrane. In current-clamp experiments 60 μM bupivacaine slightly hyperpolarized the membrane. The membrane stimulation by low-amplitude current pulses in the presence of bupivacaine showed an increase of the hyperpolarizing responses. Our findings suggest an important role of the Ih-block by local anaesthetics in the complex mechanism of drug action during epidural and spinal anaesthesia. PMID:12890706
Medici, Tom; Shortland, Peter J
Parvalbumin (PV) is a calcium binding protein that identifies a subpopulation of proprioceptive dorsal root ganglion (DRG) neurons. Calcitonin gene-related peptide (CGRP) is also expressed in a high proportion of muscle afferents but its relationship to PV is unclear. Little is known of the phenotypic responses of muscle afferents to nerve injury. Sciatic nerve axotomy or L5 spinal nerve ligation and section (SNL) lesions were used to explore these issues in adult rats using immunocytochemistry. In naive animals, the mean PV expression was 25 % of L4 or L5 dorsal root ganglion (DRG) neurons, and this was unchanged 2 weeks after sciatic nerve axotomy. Colocalization studies with the injury marker activating transcription factor 3 (ATF3) showed that approximately 24 % of PV neurons expressed ATF3 after sciatic nerve axotomy suggesting that PV may show a phenotypic switch from injured to uninjured neurons. This possibility was further assessed using the spinal nerve ligation (SNL) injury model where injured and uninjured neurons are located in different DRGs. Two weeks after L5 SNL there was no change in total PV staining and essentially all L5 PV neurons expressed ATF3. Additionally, there was no increase in PV-ir in the adjacent uninjured L4 DRG cells. Co-labelling of DRG neurons revealed that less than 2 % of PV neurons normally expressed CGRP and no colocalization was seen after injury. These experiments clearly show that axotomy does not produce down regulation of PV protein in the DRG. Moreover, this lack of change is not due to a phenotypic switch in PV immunoreactive (ir) neurons, or de novo expression of PV-ir in uninjured neurons after nerve injury. These results further illustrate differences that occur when muscle afferents are injured as compared to cutaneous afferents.
Du, Zhengqing; Meng, Ziqiang
This study addressed the effect of sulfur dioxide (SO(2)) derivatives on high-voltage-activated calcium currents (HVA-I(Ca)) in somatic membrane of freshly isolated rat dorsal root ganglion (DRG) neurons by using the whole-cell configuration of patch-clamp technique. High-threshold Ca(2+) channels are highly expressed in small dorsal root ganglion neurons. SO(2) derivatives increased the amplitudes of calcium currents in a concentration-dependent and voltage-dependent manner. The 50% enhancement concentrations (EC(50)) of SO(2) derivatives on HVA-I(Ca) was about 0.4 microM. In addition, SO(2) derivatives significantly shifted the activation and inactivation curve in the depolarizing direction. Parameters for the fit of a Boltzmann equation to mean values for the activation were V(1/2)=-17.9+/-1.3 mV before and -12.5+/-1.1 mV after application 0.5 microM SO(2) derivatives 2 min (P<0.05). The half inactivation of HVA-I(Ca) was shifted 9.7 mV to positive direction (P<0.05). Furthermore, SO(2) derivatives significantly prolonged the slow constant of inactivation, slowed the fast recovery but markedly accelerated the slow recovery of HVA-I(Ca) from inactivation. From HP of -60 mV 0.5 microM SO(2) derivatives increased the amplitude of HVA-I(Ca) with a depolarizing voltage step to -10 mV about 54.0% in small DRG neurons but 33.3% in large DRG neurons. These results indicated a possible correlation between the change of calcium channels and SO(2) inhalation toxicity, which might cause periphery neurons abnormal regulation of nociceptive transmission via calcium channels.
Kim, Koohyun; Jo, Daehyun; Kim, EungDon
Latent varicella zoster virus reactivates mainly in sensory ganglia such as the dorsal root ganglion (DRG) or trigeminal ganglion. The DRG contains many receptor channels and is an important region for pain signal transduction. Sustained abnormal electrical activity to the spinal cord via the DRG in acute herpes zoster can result in neuropathic conditions such as postherpetic neuralgia (PHN). Although the efficacy of pulsed radiofrequency (PRF) application to the DRG in various pain conditions has been previously reported, the application of PRF to the DRG in patients with herpes zoster has not yet been studied. The aim of the present study was to compare the clinical effects of PRF to the DRG in patients with herpes zoster to those of PRF to the DRG in patients with PHN. Retrospective comparative study. University hospital pain center in Korea. The medical records of 58 patients who underwent PRF to the DRG due to zoster related pain (herpes zoster or PHN) were retrospectively analyzed. Patients were divided into 2 groups according to the timing of PRF after zoster onset: an early PRF group (within 90 days) and a PHN PRF group (more than 90 days). The efficacy of PRF was assessed by a numeric rating scale (NRS) and by recording patient medication doses before PRF and at one week, 4 weeks, 8 weeks, and 12 weeks after PRF. Pain intensity was decreased after PRF in all participants. However, the degree of pain reduction was significantly higher in the early PRF group. Moreover, more patients discontinued their medication in the early PRF group, and the PRF success rate was also higher in the early PRF group. The relatively small sample size from a single center, short duration of review of medical records, and the retrospective nature of the study. PRF to the DRG is a useful treatment for treatment-resistant cases of herpes zoster and PHN. Particularly in herpes zoster patients with intractable pain, application of PRF to the DRG should be considered for pain control
Hayakawa, Tetsu; Kuwahara, Sachi; Maeda, Seishi; Tanaka, Koichi; Seki, Makoto
We have examined whether calcitonin gene-related peptide immunoreactive (CGRP-ir) neurons in the glossopharyngeal ganglia innervate the soft palate, the root of tongue, and the pharynx of the rat. Immunohistochemical observations revealed that numerous CGRP-ir neurons are located in the superior glossopharyngeal ganglion located ventrolateral to the medulla oblongata in the cranial cavity, and that CGRP-ir neurons are also located in the inferior glossopharyngeal ganglion at the jugular foramen. When Fluorogold was injected into the soft palate, the root of tongue, or the pharyngeal constrictor muscles, many retrogradely Fluorogold-labeled neurons were found in the superior glossopharyngeal ganglion and the nodose ganglion, and several Fluorogold-labeled neurons were found in the inferior glossopharyngeal ganglion. Double labeling with immunohistochemistry for CGRP and Fluorogold showed that in every case of injections of Fluorogold into the soft palate, the root of tongue, or the pharynx, about 30% of the Fluorogold-labeled neurons in the superior glossopharyngeal ganglion expressed CGRP-like immunoreactivity, while no double-labeled neurons were found in the inferior glossopharyngeal ganglion or the nodose ganglion. These results indicate that nociceptive sensory information from the soft palate, the root of tongue, and the pharynx might be conveyed by the neurons in the superior glossopharyngeal ganglion to the nucleus tractus solitarii.
Garza, Aníbal; López-Ramírez, Omar; Vega, Rosario; Soto, Enrique
Acid-sensing ionic channels (ASICs) have been shown to have a significant role in a growing number of physiological and pathological processes, such as nociception, synaptic transmission and plasticity, mechanosensation, and acidosis-induced neuronal injury. The discovery of pharmacological agents targeting ASICs has significant therapeutic potential and use as a research tool. In our work, we studied the action of transient perfusion (5-15 s) of aminoglycosides (AGs) (streptomycin and neomycin) on the proton-gated ionic currents in dorsal root ganglion (DRG) neurons of the rat and in human embryonic kidney (HEK)-293 cells. In DRG neurons, streptomycin and neomycin (30 microM) produced a significant, concentration-dependent, and reversible reduction in the amplitude of the proton-gated current, and a slowing of the desensitization rate of the ASIC current. Gentamycin (30 microM) also showed a significant reversible action on the ASIC currents. The curves of the pH effect for streptomycin and neomycin indicated that their effect was not significantly affected by pH. In HEK-293 cells, streptomycin (30 microM) produced a significant reduction in the amplitude of the proton-gated current. Neomycin and gentamycin had no significant action. Reduction of extracellular Ca(2+) concentration produced a significant increase in the action of streptomycin and neomycin on the desensitization time course of ASIC currents. These results indicate that ASICs are molecular targets for AGs, which may contribute to the understanding of their actions on excitable cells. Moreover, AGs may constitute a source to develop novel molecules with a greater affinity, specificity, and selectivity for the different ASIC subunits.
Zhang, Weiwei; Li, Zhenzhong
Targets of neuronal innervations play a vital role in regulating the survival and differentiation of innervating neurotrophin-responsive neurons. During development, neurons extend axons to their targets, and then their survival become dependent on the trophic substances secreted by their target cells. Sensory endings were present on myoblasts, myotubes, and myofibers in all intrafusal bundles regardless of age. The interdependence of sensory neurons and skeletal muscle (SKM) cells during both embryonic development and the maintenance of the mature functional state has not been fully understood. In the present study, neuromuscular cocultures of organotypic dorsal root ganglion (DRG) explants and dissociate SKM cells were established. Using this culture system, the morphological relationship between DRG neurons and SKM cells, neurites growth and neuronal migration were investigated. The migrating neurons were determined by fluorescent labeling of microtubule-associated protein-2 (MAP-2) and neurofilament 200 (NF-200) or growth-associated protein 43 (GAP-43). The expression of NF-200 and GAP-43 and their mRNAs was evaluated by Western blot assay and real time-PCR analysis. The results reveal that DRG explants showed more dense neurites outgrowth in neuromuscular cocultures as compared with that in the culture of DRG explants alone. The number of total migrating neurons (the MAP-2-expressing neurons) and the percentage NF-200-immunoreactive (IR) and GAP-43-IR neurons increased significantly in the presence of SKM cells. The levels of NF-200 and GAP-43 and their mRNAs increased significantly in neuromuscular cocultures as compared with that in the culture of DRG explants alone. These results suggested that target SKM cells play an important role in regulating neuronal protein synthesis, promoting neuritis outgrowth and neuronal migration of DRG explants in vitro. These results not only provide new clues for a better understanding of the association of SKM cells with
Palmgren, Björn; Jin, Zhe; Jiao, Yu; Kostyszyn, Beata; Olivius, Petri
At present severe damage to hair cells and sensory neurons in the inner ear results in non-treatable auditory disorders. Cell implantation is a potential treatment for various neurological disorders and has already been used in clinical practice. In the inner ear, delivery of therapeutic substances including neurotrophic factors and stem cells provide strategies that in the future may ameliorate or restore hearing impairment. In order to describe a surgical auditory nerve trunk approach, in the present paper we injected the neuronal tracer horseradish peroxidase (HRP) into the central part of the nerve by an intra cranial approach. We further evaluated the applicability of the present approach by implanting statoacoustic ganglion (SAG) cells into the same location of the auditory nerve in normal hearing rats or animals deafened by application of β-bungarotoxin to the round window niche. The HRP results illustrate labeling in the cochlear nucleus in the brain stem as well as peripherally in the spiral ganglion neurons in the cochlea. The transplanted SAGs were observed within the auditory nerve trunk but no more peripheral than the CNS-PNS transitional zone. Interestingly, the auditory nerve injection did not impair auditory function, as evidenced by the auditory brainstem response. The present findings illustrate that an auditory nerve trunk approach may well access the entire auditory nerve and does not compromise auditory function. We suggest that such an approach might compose a suitable route for cell transplantation into this sensory cranial nerve. Copyright © 2011 Elsevier B.V. All rights reserved.
Pan, Bin; Yu, Hongwei; Fischer, Gregory J; Kramer, Jeffery M; Hogan, Quinn H
Dorsal root ganglion (DRG) electrical stimulation (ganglionic field stimulation [GFS]) is effective in relieving clinical pain, but its mechanism is unknown. We therefore developed a rat model for GFS to test analgesic effects in the context of neuropathic pain. GFS was applied with a bipolar electrode at L4, using parameters replicating clinical use (20 Hz, 150-μs pulse width, current at 80% of motor threshold). Neuropathic pain was generated by tibial nerve injury (TNI). Pain behavior was monitored by determining the threshold for withdrawal from punctate mechanical stimuli, by identifying hyperalgesic responses to noxious mechanical stimuli, and by hypersensitivity to cold. The affective dimension of pain was measured using conditioned place preference. We found that electrode insertion caused no behavioral evidence of pain and produced no histological evidence of DRG damage. GFS reversed TNI-induced hypersensitivity to cold and mechanical hyperalgesia and allodynia. Allodynia remained diminished 15 minutes after GFS. Conditioned place preference showed that GFS was not rewarding in uninjured control animals but was rewarding in animals subjected to TNI, which reveals analgesic efficacy of GFS for spontaneous pain. We conclude that GFS relieves neuropathic pain in rats. This model may provide a platform for identifying mechanisms and novel applications of GFS. We show that electrical stimulation of the DRG in rats reverses neuropathic pain behavior and provides a rewarding effect to animals with spontaneous neuropathic pain. This confirms analgesic efficacy of DRG stimulation in an animal model, and provides a platform for preclinical exploration. Copyright Â© 2016 American Pain Society. Published by Elsevier Inc. All rights reserved.
Ayar, Ahmet; Kalkan, Omer-Faruk; Canpolat, Sinan; Tasdemir, Tamer; Ozan, Ulku
Background The aim of this study was to investigate the possible neurotoxic effects of 3 root canal sealers (RCSs) (AH Plus, GuttaFlow, iRoot SP) on cultured rat trigeminal ganglion (TG) neurons. Material and Methods Primary cultures of TG neurons were obtained from 1 to 2-day old rats. Freshly mixed RCSs were incubated in sterile phosphate buffered saline and cells were incubated with supernatants of the RCSs for different time intervals (1-, 3-, 6- and 24-h; 1 or 1/10 diluted) and viability/cytotoxicity was tested by counting the number of live cells. Pair of dishes with cells from the same culture incubated with only culture medium was considered as negative controls. Cell images were captured and acquired at x200 magnification using a microscope equipped with a camera using special image program. The viable cells were manually counted assigned from the images for each dose and incubation duration. Data was analysed by using 1-way analysis of variance with Tukey post hoc tests. Results There was no significant change in cell viability after short duration of incubation (1- and 3-h) with the supernatant of any of RCSs, except for undiluted-AH Plus at 3-h. When AH Plus was compared with other RCSs, for diluted supernatants, there was only significant difference between iRoot SP and AH Plus at 24-h (P<0.05). Whereas undiluted-AH Plus was significantly more cytotoxic for 3-, 6- and 24-h periods as compared to respective incubation periods of undiluted other groups (P<0.05). GuttaFlow groups had similar neurotoxic effect on cells for all test periods. Conclusions All tested RCSs exhibited a variable degree of neurotoxicity on these primary sensory neurons of orofacial tissues, depending on their chemical compositions. GuttaFlow and iRoot SP evoked a less toxic response to TG cells than AH Plus. Key words:Neurotoxicity, trigeminal ganglia, cell culture, root canal sealer, AH Plus, GuttaFlow, iRoot SP. PMID:28149460
Matsuoka, Akihiro J; Morrissey, Zachery D; Zhang, Chaoying; Homma, Kazuaki; Belmadani, Abdelhak; Miller, Charles A; Chadly, Duncan M; Kobayashi, Shun; Edelbrock, Alexandra N; Tanaka-Matakatsu, Miho; Whitlon, Donna S; Lyass, Ljuba; McGuire, Tammy L; Stupp, Samuel I; Kessler, John A
The ability to generate spiral ganglion neurons (SGNs) from stem cells is a necessary prerequisite for development of cell-replacement therapies for sensorineural hearing loss. We present a protocol that directs human embryonic stem cells (hESCs) toward a purified population of otic neuronal progenitors (ONPs) and SGN-like cells. Between 82% and 95% of these cells express SGN molecular markers, they preferentially extend neurites to the cochlear nucleus rather than nonauditory nuclei, and they generate action potentials. The protocol follows an in vitro stepwise recapitulation of developmental events inherent to normal differentiation of hESCs into SGNs, resulting in efficient sequential generation of nonneuronal ectoderm, preplacodal ectoderm, early prosensory ONPs, late ONPs, and cells with cellular and molecular characteristics of human SGNs. We thus describe the sequential signaling pathways that generate the early and later lineage species in the human SGN lineage, thereby better describing key developmental processes. The results indicate that our protocol generates cells that closely replicate the phenotypic characteristics of human SGNs, advancing the process of guiding hESCs to states serving inner-ear cell-replacement therapies and possible next-generation hybrid auditory prostheses. © Stem Cells Translational Medicine 2017;6:923-936.
Hoffman, E Matthew; Zhang, Zijia; Schechter, Ruben; Miller, Kenneth E
Glutamate is a neurotransmitter used at both the peripheral and central terminals of nociceptive primary sensory neurons, yet little is known concerning regulation of glutamate metabolism during peripheral inflammation. Glutaminase (GLS) is an enzyme of the glutamate-glutamine cycle that converts glutamine into glutamate for neurotransmission and is implicated in producing elevated levels of glutamate in central and peripheral terminals. A potential mechanism for increased levels of glutamate is an elevation in GLS expression. We assessed GLS expression after unilateral hind paw inflammation by measuring GLS immunoreactivity (ir) with quantitative image analysis of L4 dorsal root ganglion (DRG) neurons after one, two, four, and eight days of adjuvant-induced arthritis (AIA) compared to saline injected controls. No significant elevation in GLS-ir occurred in the DRG ipsilateral to the inflamed hind paw after one or two days of AIA. After four days AIA, GLS-ir was elevated significantly in all sizes of DRG neurons. After eight days AIA, GLS-ir remained elevated in small (<400 µm²), presumably nociceptive neurons. Western blot analysis of the L4 DRG at day four AIA confirmed the elevated GLS-ir. The present study indicates that GLS expression is increased in the chronic stage of inflammation and may be a target for chronic pain therapy.
Hoffman, E. Matthew; Zhang, Zijia; Schechter, Ruben; Miller, Kenneth E.
Glutamate is a neurotransmitter used at both the peripheral and central terminals of nociceptive primary sensory neurons, yet little is known concerning regulation of glutamate metabolism during peripheral inflammation. Glutaminase (GLS) is an enzyme of the glutamate-glutamine cycle that converts glutamine into glutamate for neurotransmission and is implicated in producing elevated levels of glutamate in central and peripheral terminals. A potential mechanism for increased levels of glutamate is an elevation in GLS expression. We assessed GLS expression after unilateral hind paw inflammation by measuring GLS immunoreactivity (ir) with quantitative image analysis of L4 dorsal root ganglion (DRG) neurons after one, two, four, and eight days of adjuvant-induced arthritis (AIA) compared to saline injected controls. No significant elevation in GLS-ir occurred in the DRG ipsilateral to the inflamed hind paw after one or two days of AIA. After four days AIA, GLS-ir was elevated significantly in all sizes of DRG neurons. After eight days AIA, GLS-ir remained elevated in small (<400 µm2), presumably nociceptive neurons. Western blot analysis of the L4 DRG at day four AIA confirmed the elevated GLS-ir. The present study indicates that GLS expression is increased in the chronic stage of inflammation and may be a target for chronic pain therapy. PMID:26771651
Raymon, H K; Thode, S; Zhou, J; Friedman, G C; Pardinas, J R; Barrere, C; Johnson, R M; Sah, D W
A renewable source of human sensory neurons would greatly facilitate basic research and drug development. We had established previously conditionally immortalized human CNS cell lines that can differentiate into functional neurons (). We report here the development of an immortalized human dorsal root ganglion (DRG) clonal cell line, HD10.6, with a tetracycline-regulatable v-myc oncogene. In the proliferative condition, HD10.6 cells have a doubling time of 1.2 d and exhibit a neuronal precursor morphology. After differentiation of clone HD10.6 for 7 d in the presence of tetracycline, v-myc expression was suppressed, and >50% of the cells exhibited typical neuronal morphology, stained positively for neuronal cytoskeletal markers, and fired action potentials in response to current injection. Furthermore, this cell line was fate-restricted to a neuronal phenotype; even in culture conditions that promote Schwann cell or smooth muscle differentiation of neural crest stem cells, HD10.6 differentiated exclusively into neurons. Moreover, differentiated HD10.6 cells expressed sensory neuron-associated transcription factors and exhibited capsaicin sensitivity. Taken together, these data indicate that we have established an immortalized human DRG cell line that can differentiate into sensory neurons with nociceptive properties. The cell line HD10.6 represents the first example of a human sensory neuronal line and will be valuable for basic research, as well as for the discovery of novel drug targets and clinical candidates.
Qu, Zu-Wei; Liu, Ting-Ting; Qiu, Chun-Yu; Li, Jia-Da; Hu, Wang-Ping
Chlorogenic acid (CGA) is one of the most abundant polyphenol compounds in the human diet. Recently, it is demonstrated to have potent antinociceptive effect. However, little is understood about the mechanism underlying CGA analgesia. Here, we have found that CGA can exert an inhibitory effect on the functional activity of native acid-sensing ion channels (ASICs) in rat dorsal root ganglion (DRG) neurons. First, CGA decreased the peak amplitude of proton-gated currents mediated by ASICs in a concentration-dependent manner. Second, CGA shifted the proton concentration-response curve downward, with a decrease of 41.76 ± 8.65% in the maximum current response to protons but with no significant change in the pH0.5 value. Third, CGA altered acidosis-evoked membrane excitability of rat DRG neurons and caused a significant decrease in the amplitude of the depolarization and the number of action potentials induced by acid stimuli. Finally, peripheral administered CGA attenuated nociceptive response to intraplantar injection of acetic acid in rats. ASICs are distributed in peripheral sensory neurons and participate in nociception. Our findings CGA inhibition of native ASICs indicated that CGA may exert analgesic action by modulating ASICs in the primary afferent neurons, which revealed a novel cellular and molecular mechanism underlying CGA analgesia.
Nie, Aifang; Wei, Cailing; Meng, Ziqiang
The effects of sodium metabisulfite (SMB), a general food preservative, on potassium currents in rat dorsal root ganglion (DRG) neurons were investigated using the whole-cell patch-clamp technique. SMB increased the amplitudes of both transient outward potassium currents and delayed rectifier potassium current in concentration- and voltage-dependent manner. The transient outward potassium currents (TOCs) include a fast inactivating (A-current or IA) current and a slow inactivating (D-current or ID) current. SMB majorly increased IA, and ID was little affected. SMB did not affect the activation process of transient outward currents (TOCs), but the inactivation curve of TOCs was shifted to more positive potentials. The inactivation time constants of TOCs were also increased by SMB. For delayed rectifier potassium current (IK), SMB shifted the activation curve to hyperpolarizing direction. SMB differently affected TOCs and IK, its effects major on A-type K+ channels, which play a role in adjusting pain sensitivity in response to peripheral redox conditions. SMB did not increase TOCs and IK when adding DTT in pipette solution. These results suggested that SMB might oxidize potassium channels, which relate to adjusting pain sensitivity in pain-sensing DRG neurons.
Lei, Zhen; Li, Xiaoyu; Wang, Guizhi; Fei, Jianchun; Meng, Tao; Zhang, Xinyu; Yu, Jingya; Yu, Jingui; Li, Jingxin
Acid-sensing ion channels (ASICs), part of the epithelial sodium channel/degenerin family, are activated by extracellular protons. The ASICs play a significant role in the acidosis-mediated perception of pain. The anaesthetic agent propofol also exerts antinociceptive effects, but the underlying mechanisms for this effect are not clear. We used whole-cell patch clamping to investigate the effect of propofol on proton-gated currents in: (i) rat dorsal root ganglion (DRG) neurons; and (ii) HEK293 cells transfected with either ASIC1a or ASIC3. Propofol inhibited the amplitude of proton-gated currents in DRG neurons, but did not change the sensitivity of ASICs to H(+). Notably, propofol altered acid-evoked excitability of rat DRG neurons and decreased the number of action potentials induced by acid stimuli. In addition, we demonstrated that propofol inhibited ASICs by directly binding with these channels in HEK293 cells. These results suggest that propofol inhibits proton-gated currents in DRG neurons and that inhibition of proton-gated currents explains, in part, the antinociceptive effects of propofol in primary afferent neurons.
Zhang, Hua; Verkman, A.S.
Dorsal root ganglion (DRG) neurons transduce peripheral pain signals through small-diameter, non-myelinated C-fibers, which, when injured, can regenerate to restore pain sensation. Water channel aquaporin-1 (AQP1) is expressed at the plasma membrane of cell bodies and axons of DRG neurons, where it modulates the sensing of certain types of pain. Here, we found that AQP1 is also involved in DRG axonal growth and regeneration by a mechanism that may involve water transport-facilitated extension of axonal outgrowths. Spontaneous and nerve growth factor-stimulated axonal extension was reduced in cultures of AQP1-deficient DRG neurons and DRG explants compared to the wildtype. Axonal growth in AQP1-deficient DRG cultures was rescued by transfection with AQP1 or a different water-transporting AQP (AQP4), but not by a non-water-transporting AQP1 mutant. Following sciatic nerve compression injury AQP1 expression was increased in DRG neurons in wildtype mice, and DRG axonal growth was impaired in AQP1-deficient mice. Our results indicate AQP1 as a novel determinant of DRG axonal regeneration and hence a potential therapeutic target to accelerate neuronal regeneration. PMID:25585012
Zhang, Hua; Verkman, A S
Dorsal root ganglion (DRG) neurons transduce peripheral pain signals through small-diameter, non-myelinated C-fibers, which, when injured, can regenerate to restore pain sensation. Water channel aquaporin-1 (AQP1) is expressed at the plasma membrane of cell bodies and axons of DRG neurons, where it modulates the sensing of certain types of pain. Here, we found that AQP1 is also involved in DRG axonal growth and regeneration by a mechanism that may involve water transport-facilitated extension of axonal outgrowths. Spontaneous and nerve growth factor-stimulated axonal extension was reduced in cultures of AQP1-deficient DRG neurons and DRG explants compared to the wildtype. Axonal growth in AQP1-deficient DRG cultures was rescued by transfection with AQP1 or a different water-transporting AQP (AQP4), but not by a non-water-transporting AQP1 mutant. Following sciatic nerve compression injury AQP1 expression was increased in DRG neurons in wildtype mice, and DRG axonal growth was impaired in AQP1-deficient mice. Our results indicate AQP1 as a novel determinant of DRG axonal regeneration and hence a potential therapeutic target to accelerate neuronal regeneration. Copyright © 2015 Elsevier Inc. All rights reserved.
Lopez-Santiago, Luis F.; Brackenbury, William J.; Chen, Chunling; Isom, Lori L.
Nociceptive dorsal root ganglion (DRG) neurons express tetrodotoxin-sensitive (TTX-S) and -resistant (TTX-R) Na+ current (INa) mediated by voltage-gated Na+ channels (VGSCs). In nociceptive DRG neurons, VGSC β2 subunits, encoded by Scn2b, selectively regulate TTX-S α subunit mRNA and protein expression, ultimately resulting in changes in pain sensitivity. We hypothesized that VGSCs in nociceptive DRG neurons may also be regulated by β1 subunits, encoded by Scn1b. Scn1b null mice are models of Dravet Syndrome, a severe pediatric encephalopathy. Many physiological effects of Scn1b deletion on CNS neurons have been described. In contrast, little is known about the role of Scn1b in peripheral neurons in vivo. Here we demonstrate that Scn1b null DRG neurons exhibit a depolarizing shift in the voltage dependence of TTX-S INa inactivation, reduced persistent TTX-R INa, a prolonged rate of recovery of TTX-R INa from inactivation, and reduced cell surface expression of Nav1.9 compared with their WT littermates. Investigation of action potential firing shows that Scn1b null DRG neurons are hyperexcitable compared with WT. Consistent with this, transient outward K+ current (Ito) is significantly reduced in null DRG neurons. We conclude that Scn1b regulates the electrical excitability of nociceptive DRG neurons in vivo by modulating both INa and IK. PMID:21555511
Tata, Ada Maria; De Stefano, M Egle; Tomassy, Giulio Srubek; Vilaró, M Teresa; Levey, Allan I; Biagioni, Stefano
The vesicular acetylcholine transporter (VAChT) is a transmembrane protein required, in cholinergic neurons, for selective storage of acetylcholine into synaptic vesicles. Although dorsal root ganglion (DRG) neurons utilize neuropeptides and amino acids for neurotransmission, we have previously demonstrated the presence of a cholinergic system. To investigate whether, in sensory neurons, the vesicular accumulation of acetylcholine relies on the same mechanisms active in classical cholinergic neurons, we investigated VAChT presence, subcellular distribution, and activity. RT-PCR and Western blot analysis demonstrated the presence of VAChT mRNA and protein product in DRG neurons and in the striatum and cortex, used as positive controls. Moreover, in situ hybridization and immunocytochemistry showed VAChT staining located mainly in the medium/large-sized subpopulation of the sensory neurons. A few small neurons were also faintly labeled by immunocytochemistry. In the electron microscope, immunolabeling was associated with vesicle-like elements distributed in the neuronal cytoplasm and in both myelinated and unmyelinated intraganglionic nerve fibers. Finally, [(3)H]acetylcholine active transport, evaluated either in the presence or in the absence of ATP, also demonstrated that, as previously reported, the uptake of acetylcholine by VAChT is ATP dependent. This study suggests that DRG neurons not only are able to synthesize and degrade ACh and to convey cholinergic stimuli but also are capable of accumulating and, possibly, releasing acetylcholine by the same mechanism used by the better known cholinergic neurons.
Yuan, Huijun; Lin, Jiarui; Lan, Tonghan
Nano red elemental selenium (Nano-Se), was demonstrated to be useful in medical and scientific researches. Here, we investigated the effects of Nano-Se on sodium currents on rat dorsal root ganglion neurons (DRG), using the whole-cell patch clamp method. Nano-Se reversibly decrease the I(Na)(TTX-S) in a concentration-dependent, time-dependent and open-channel block manners without affecting I(Na)(TTX-R). It shifted the steady-state activation and inactivation curves for I(Na) to more negative potentials. In the research of recovery from inactivation, the recovery time constant is longer in the present of Nano-Se. Nano-Se had a weaker inhibitory effect on I(Na), compared with marked decrease caused by selenite which indicated that Nano-Se is less neurotoxic than selenite in short-term/large dose treatments and had similar bio availability to sodium selenite. The results of interaction between the effects of Nano-Se and selenite on sodium currents indicated a negative allosteric interaction between the selenite binding site and the Nano-Se binding site or that they have the same competitive binding site.
Yu, Shan-Shan; Yu, Kuai; Gu, Yan; Ruan, Di-Yun
The physiological role of taurine, an abundant free amino acid in the neural system, is still poorly understood. The aim of this study was to investigate its effect on TTX-sensitive (TTX-S) and TTX-resistant (TTX-R) Na+ currents in enzymatically dissociated neurons from rat dorsal root ganglion (DRG) with conventional whole-cell recording manner under voltage-clamp conditions. A TTX-S Na+ current was recorded preferentially from large DRG neurons and a TTX-R Na+ current preferentially from small ones. For TTX-S Na+ channel, taurine of the concentration > or = 10 mM shifted the activation curve in the depolarizing direction and the inactivation curve in the hyperpolarizing direction. There was no change in the activation curve for TTX-R Na+ channel and the inactivation curve was shifted in the hyperpolarizing direction slightly in the presence of taurine > or = 20 mM. When the recovery kinetics was examined, the presence of taurine resulted in a slower recovery from inactivation of TTX-S currents and no change of TTX-R ones. All the effects of taurine were weakly concentration-dependent and partly recovered quite slowly after washout. Our data indicate that taurine alters the properties of Na+ currents in intact DRG neurons. These may contribute to the understanding of taurine as a natural neuroprotectant and the potential of taurine as a useful medicine for the treatment of sensory neuropathies.
Jia, Zhanfeng; Ling, Jennifer; Gu, Jianguo G
Rapidly adapting mechanically activated channels (RA) are expressed on somatosensory neurons and thought to play a role in mechanical transduction. Because mechanical sensations can be significantly affected by temperatures, we examined thermal sensitivity of RA currents in cultured dorsal root ganglion (DRG) neurons to see if RA channel activity is highly temperature-dependent. RA currents were evoked from DRG neurons by membrane displacements and recorded by the whole-cell patch-clamp recording technique. We found that RA currents were significantly enhanced by warming temperatures from 22 to 32 °C and reduced by cooling temperatures from 24 to 14 °C. RA channel activation exhibited steep temperature-dependence with a large temperature coefficient (Q10>5) and a high activation energy (Ea>30 kcal/mol). We further showed that RA channel activation by mechanical stimulation led to membrane depolarization, which could result in action potential firing at 22 °C or 32 °C but not at 14 °C. Taken together, our results provide the measurements of thermal dynamics and activation energy of RA channels, and suggest that a high energy barrier is present for RA channels to open. These findings are in agreement with temperature sensitivity of mechanical sensations in mammals.
Shang, Shujiang; Zhu, Feipeng; Liu, Bin; Chai, Zuying; Wu, Qihui; Hu, Meiqin; Wang, Yuan; Huang, Rong; Zhang, Xiaoyu; Wu, Xi; Sun, Lei; Wang, Yeshi; Wang, Li; Xu, Huadong; Teng, Sasa; Liu, Bing; Zheng, Lianghong; Zhang, Chen; Zhang, Fukang; Feng, Xinghua; Zhu, Desheng
Transient receptor potential A1 (TRPA1) is a nonselective cation channel implicated in thermosensation and inflammatory pain. In this study, we show that TRPA1 (activated by allyl isothiocyanate, acrolein, and 4-hydroxynonenal) elevates the intracellular Ca2+ concentration ([Ca2+]i) in dorsal root ganglion (DRG) neurons in the presence and absence of extracellular Ca2+. Pharmacological and immunocytochemical analyses revealed the presence of TRPA1 channels both on the plasma membrane and in endolysosomes. Confocal line-scan imaging demonstrated Ca2+ signals elicited from individual endolysosomes (“lysosome Ca2+ sparks”) by TRPA1 activation. In physiological solutions, the TRPA1-mediated endolysosomal Ca2+ release contributed to ∼40% of the overall [Ca2+]i rise and directly triggered vesicle exocytosis and calcitonin gene-related peptide release, which greatly enhanced the excitability of DRG neurons. Thus, in addition to working via Ca2+ influx, TRPA1 channels trigger vesicle release in sensory neurons by releasing Ca2+ from lysosome-like organelles. PMID:27799370
Shang, Shujiang; Zhu, Feipeng; Liu, Bin; Chai, Zuying; Wu, Qihui; Hu, Meiqin; Wang, Yuan; Huang, Rong; Zhang, Xiaoyu; Wu, Xi; Sun, Lei; Wang, Yeshi; Wang, Li; Xu, Huadong; Teng, Sasa; Liu, Bing; Zheng, Lianghong; Zhang, Chen; Zhang, Fukang; Feng, Xinghua; Zhu, Desheng; Wang, Changhe; Liu, Tao; Zhu, Michael X; Zhou, Zhuan
Transient receptor potential A1 (TRPA1) is a nonselective cation channel implicated in thermosensation and inflammatory pain. In this study, we show that TRPA1 (activated by allyl isothiocyanate, acrolein, and 4-hydroxynonenal) elevates the intracellular Ca(2+) concentration ([Ca(2+)]i) in dorsal root ganglion (DRG) neurons in the presence and absence of extracellular Ca(2+) Pharmacological and immunocytochemical analyses revealed the presence of TRPA1 channels both on the plasma membrane and in endolysosomes. Confocal line-scan imaging demonstrated Ca(2+) signals elicited from individual endolysosomes ("lysosome Ca(2+) sparks") by TRPA1 activation. In physiological solutions, the TRPA1-mediated endolysosomal Ca(2+) release contributed to ∼40% of the overall [Ca(2+)]i rise and directly triggered vesicle exocytosis and calcitonin gene-related peptide release, which greatly enhanced the excitability of DRG neurons. Thus, in addition to working via Ca(2+) influx, TRPA1 channels trigger vesicle release in sensory neurons by releasing Ca(2+) from lysosome-like organelles. © 2016 Shang et al.
I propose a model that replicates the kinetics of a rapidly-adapting mechanosensitive current generated by a dorsal root ganglion (DRG) neuron. When the DRG neuron is mechanically stimulated, an ionic current called a mechanosensitive current flows across its membrane. The kinetics of mechanosensitive currents are broadly classified into three types; rapidly adapting (RA), intermediately adapting, and slowly adapting. The kinetics of RA mechanosensitive currents are particularly intriguing. An RA mechanosensitive current is initially evoked by and rapidly adapts to a mechanical stimulus, but can also respond to an additional stimulus. Furthermore, an antecedent stimulus immediately followed by an additional stimulus suppresses reactivation of the current. The features of the kinetics depend on the characteristics of the mechanotransducer channels. Physiologists have proposed three factors associated with mechanotransducer channels, invoking activation, adaptation, and inactivation. In the present study, these factors are incorporated into an RA mechanosensitive current model. Computer simulations verified that the proposed model replicates the kinetics of real RA DRG mechanosensitive currents. The mechanosensitive current elicited by successive pulse-form stimuli was predominantly desensitized by the inactivating factor. Both the inactivating and adapting factors were involved in desensitization of a double-decker stimulus. The reduction of the sensitivity with decreasing velocity of the stimulus was mainly controlled by the adapting factor.
Repić, Tihana; Madirazza, Katarina; Bektur, Ezgi; Sapunar, Damir
Our study focuses on characterization of dorsal root ganglion (DRG) neurons cultured on silicon micro-pillar substrates (MPS) with the ultimate goal of designing micro-electrode arrays (MEAs) for successful electrophysiological recordings of DRG neurons. Adult and neonatal DRG neurons were cultured on MPS and glass coverslips for 7 days in vitro. DRG neuronal distribution and morphometric analysis, including neurite alignment and length, was performed on MPS areas with different pillar width and spacing. We showed that MPS provide an environment for growth of adult and neonatal DRG neurons as permissive as control glass surfaces. Neonatal DRG neurons were present on MPS areas with narrow pillar spacing, while adult neurons preferred wider pillar spacing. Compared to the control glass surfaces the neonatal and adult DRG neurons in regions with narrow pillar spacing range developed a smaller number of longer neurites. In the same area, neurites were preferentially oriented along three directional axes at 30°, 90° and 150°. MPS architecture influenced growth directionality of all main DRG neuronal subtypes. We can conclude that specific micro-pillar substrate topography affects the morphology of DRG neurons. This knowledge can enable development of MEAs with precisely defined physical features for various neuroscience applications. PMID:28008963
Han, Sungmin; Chu, Jun-Uk; Kim, Hyungmin; Park, Jong Woong; Youn, Inchan
Proprioceptive afferent activities could be useful for providing sensory feedback signals for closed-loop control during functional electrical stimulation (FES). However, most previous studies have used the single-unit activity of individual neurons to extract sensory information from proprioceptive afferents. This study proposes a new decoding method to estimate ankle and knee joint angles using multiunit activity data. Proprioceptive afferent signals were recorded from a dorsal root ganglion with a single-shank microelectrode during passive movements of the ankle and knee joints, and joint angles were measured as kinematic data. The mean absolute value (MAV) was extracted from the multiunit activity data, and a dynamically driven recurrent neural network (DDRNN) was used to estimate ankle and knee joint angles. The multiunit activity-based MAV feature was sufficiently informative to estimate limb states, and the DDRNN showed a better decoding performance than conventional linear estimators. In addition, processing time delay satisfied real-time constraints. These results demonstrated that the proposed method could be applicable for providing real-time sensory feedback signals in closed-loop FES systems. PMID:28276474
Sun, Wenjing; Smith, Daniel; Bryn, Steven; Borgens, Richard; Shi, Riyi
Axonal demyelination is a critical pathological phenomenon associated with spinal cord injury and multiple sclerosis (MS). Previous studies demonstrated that 4-Aminopyridine, a fast potassium channel blocker, enhances impulse conduction on damaged and/or demyelinated axons, allowing for functional recovery in spinal cord injuries and MS, but with severe therapeutic limitations. To continue to explore the therapeutic value of blocking fast potassium channels while circumventing the side effects of 4-AP, we have developed three novel 4-AP derivatives that enhance impulse conduction in spinal cord trauma. In the current study, we have shown that one of these three derivatives, N-(4-pyridyl) methyl carbamates (MC), significantly inhibits a fast, I(A) like potassium current in guinea pig dorsal root ganglion cells in a whole cell patch clamp configuration. This inhibition of I(A) likely plays a critical role in MC's ability to restore conduction in mechanically injured spinal cord axons and may present a viable alternative to 4-AP for individuals with spinal cord injury or MS. From this, compounds with greater efficacy and perhaps less side effects will likely emerge in the near future, which will greatly enhance the functional restoration and lessen the suffering of SCI and MS patients.
Robas, Nicola; Mead, Emma; Fidock, Mark
MrgX2 is a recently identified orphan G-protein-coupled receptor whose ligand and physiological function were unknown. Here we describe cortistatin, a neuropeptide for which no specific receptor has been identified previously, as a high potency ligand at MrgX2. Cortistatin has several biological functions including roles in sleep regulation, locomotor activity, and cortical function. Using a "reverse pharmacology" approach, we have identified a number of additional cyclic peptide agonists for MrgX2, determined their rank order of potency, and demonstrated that this receptor has a pharmacological profile distinct from the other characterized members of the Mrg (Mas-related genes) family. In MrgX2-expressing cells, cortistatin-stimulated increases in intracellular Ca2+ but had no effect on basal or forskolin-stimulated cAMP levels, suggesting that this receptor is Gq-coupled. Immunohistochemical and quantitative PCR studies show MrgX2 to have a limited expression profile, both peripheral and within the central nervous system, with highest levels in dorsal root ganglion.
Hernández, M A; Avila, J; Moya, F; Alberto, C
In primary cultures of dorsal root ganglion cells from rat embryos, neurons undergo a morphological transformation from a bipolar to a differentiated pseudo-unipolar shape, resembling their developmental stages in vivo. Cells present in these cultures are characterized here by immunological criteria using monoclonal and polyclonal antibodies against microtubule associated proteins MAP1 and MAP2 and against tubulin. After development for seven days in culture, antibodies against microtubule associated proteins MAP1 brightly labeled cells with neuronal morphology and lightly stained cells with the shape of Schwann cells. In addition, an extended network of neuronal processes was labeled with this antibody. Anti-microtubule associated protein MAP2 stained only neurons and a more restricted network of neuronal processes. The compartmentalization of microtubule associated protein MAP2 during the maturation process was followed by double-labeling with antibodies to microtubule associated proteins MAP1 and MAP2. Initially, microtubule associated protein MAP2 was present in the cell body and the two processes of bipolar neurons. Subsequently, the labeling of both processes changed, depending on neuronal morphology. In neurons in which both processes were approaching one another, one of these neurites was stained predominantly with anti-microtubule associated protein MAP2. Finally, in pseudo, unipolar neurons, anti-microtubule associated protein MAP2 labeling was found in the cell body and excluded from the more distal processes.
Han, Sungmin; Chu, Jun-Uk; Kim, Hyungmin; Park, Jong Woong; Youn, Inchan
Proprioceptive afferent activities could be useful for providing sensory feedback signals for closed-loop control during functional electrical stimulation (FES). However, most previous studies have used the single-unit activity of individual neurons to extract sensory information from proprioceptive afferents. This study proposes a new decoding method to estimate ankle and knee joint angles using multiunit activity data. Proprioceptive afferent signals were recorded from a dorsal root ganglion with a single-shank microelectrode during passive movements of the ankle and knee joints, and joint angles were measured as kinematic data. The mean absolute value (MAV) was extracted from the multiunit activity data, and a dynamically driven recurrent neural network (DDRNN) was used to estimate ankle and knee joint angles. The multiunit activity-based MAV feature was sufficiently informative to estimate limb states, and the DDRNN showed a better decoding performance than conventional linear estimators. In addition, processing time delay satisfied real-time constraints. These results demonstrated that the proposed method could be applicable for providing real-time sensory feedback signals in closed-loop FES systems.
Viatchenko-Karpinski, Viacheslav; Gu, Jianguo G
Primary afferent fibers use mechanically activated (MA) currents to transduce innocuous and noxious mechanical stimuli. However, it is largely unknown about the differences in MA currents between the afferents for sensing innocuous and noxious stimuli. In the present study, we used dorsal root ganglion (DRG) neurons acutely dissociated from rats and studied their MA currents and also their intrinsic membrane properties. Recorded from small-sized DRG neurons, we found that most of these neurons were mechanically sensitive (MS) showing MA currents. The MS neurons could be classified into nociceptive-like mechanically sensitive (Noci-MS) and non-nociceptive-like mechanically sensitive (nonNoci-MS) neurons based on their action potential shapes. Noci-MS neurons responded to mechanical stimulation with three types of MA currents, rapidly adapting (RA), intermediately adapting (IA), and slowly adapting (SA) currents. In contrast, almost all nonNoci-MS neurons showed RA current type in response to mechanical stimulation. Mechanical thresholds had a broad range for both nonNoci-MS and Noci-MS neurons, and the thresholds were not significantly different between them. However, MA current densities were significantly smaller in Noci-MS than in nonNoci-MS neurons. Noci-MS and nonNoci-MS neurons also showed significant differences in their electrophysiological properties including action potential (AP) thresholds and AP firing patterns. These differences may contribute to the differential sensory encoding for innocuous and noxious mechanical stimuli. Copyright © 2016. Published by Elsevier Ireland Ltd.
Hara, Tomomi; Chiba, Terumasa; Abe, Kenji; Makabe, Akiko; Ikeno, Souichi; Kawakami, Kazuyoshi; Utsunomiya, Iku; Hama, Toshihiro; Taguchi, Kyoji
Peripheral neuropathy is a common adverse effect of paclitaxel treatment. To analyze the contribution of transient receptor potential vanilloid 1 (TRPV1) in the development of paclitaxel-induced thermal hyperalgesia, TRPV1 expression in the rat dorsal root ganglion (DRG) was analyzed after paclitaxel treatment. Behavioral assessment using the tail-flick test showed that intraperitoneal administration of 2 and 4 mg/kg paclitaxel induced thermal hyperalgesia after days 7, 14, and 21. Paclitaxel-induced thermal hyperalgesia after day 14 was significantly inhibited by the TRP antagonist ruthenium red (3 mg/kg, s.c.) and the TRPV1 antagonist capsazepine (30 mg/kg, s.c.). Paclitaxel (2 and 4 mg/kg) treatment increased the expression of TRPV1 mRNA and protein in DRG neurons. Immunohistochemistry showed that paclitaxel (4 mg/kg) treatment increased TRPV1 protein expression in small and medium DRG neurons 14 days after treatment. Antibody double labeling revealed that isolectin B4-positive small DRG neurons co-expressed TRPV1. TRPV1 immunostaining was up-regulated in paw skin day 14 after paclitaxel treatment. Moreover, in situ hybridization histochemistry revealed that most of the TRPV1 mRNA-labeled neurons in the DRG were small or medium in size. These results suggest that paclitaxel treatment increases TRPV1 expression in DRG neurons and may contribute to functional peripheral neuropathic pain.
Kayano, Tomohiko; Kitamura, Naoki; Moriya, Taiki; Kuwahara, Takeshi; Komagiri, You; Toescu, Emil C; Shibuya, Izumi
Adult rat dorsal root ganglion (DRG) neurons cultured in the presence of 100-ng/mL NGF were reported to show spontaneous action potentials in the cell-attached recording. In this study, underlying mechanisms were examined in the whole-cell and outside-out voltage clamp recording. In 75% neurons with on-cell firing, transient inward current spikes were repetitively recorded in the voltage clamp mode at -50 mV in the whole-cell configuration (named "Isp"). Isp with stable amplitudes occurred in an all-or-none fashion, and was abolished by TTX (< 100 nM), lidocaine (< 1 mM) and a reduction of extracellular Na(+) (154 to 100 mM) in an all-or-none fashion, suggesting that Isp reflects spontaneous dicharges occurring at the loosely voltage-clamped regions. Isp was also observed in the excised outside-out patches and the kinetics and the sensitivity to TTX and lidocaine resembled those in the whole-cell. Spontaneous action potentials were also recorded in the current clamp mode. Small subthreshold spikes often preceded the action potentials. When the localized discharge affected a whole-somatic membrane potential to overcome a threshold, the action potential generated. These results indicate that the triggering sources of the action potential exist in the somatic membrane itself in NGF-treated DRG neurons.
Xiao, Yun; Wu, Yang; Zhao, Bo; Xia, Zhongyuan
Voltage-gated potassium channels (KV) regulate pain transmission by controlling neuronal excitability. Changes in KV expression patterns may thus contribute toward hyperalgesia following nerve injury. The aim of this study was to characterize KV current density in dorsal root ganglion (DRG) neurons following chronic constriction injury (CCI) of the right sciatic nerve, a robust model of post-traumatic neuropathic pain. The study examined changes in small-diameter potassium ion currents (<30 µm) in neurons in the L4-L6 DRG following CCI by whole-cell patch-clamping and the association with post-CCI mechanical and thermal nociceptive thresholds. Compared with the control group, 7 days after CCI, the mechanical force and temperature required to elicit ipsilateral foot withdrawal decreased significantly, indicating tactile allodynia and thermal hyperalgesia. Post-CCI neurons had a significantly lower rheobase current and depolarized resting membrane potential than controls, suggesting KV current downregulation. Some ipsilateral DRG neurons also had spontaneous action potentials and repetitive firing. There was a 55% reduction in the total KV current density caused by a 55% decrease in the sustained delayed rectifier potassium ion current (IK) density and a 17% decrease in the transient A-type potassium ion current (IA) density. These results indicated that changes in DRG neuron IK and IA current density and concomitant afferent hyperexcitability may contribute toward neuropathic pain following injury. The rat CCI model may prove valuable for examining pathogenic mechanisms and potential therapies, such as KV channel modulators.
Pachuau, Judith; Martin-Caraballo, Miguel
In this study we have characterized the functional expression of T-type Ca(2+) channels in developing chick nodose neurons, a population of placode-derived sensory neurons innervating the heart and various visceral organs. Voltage-gated Ca(2+) currents were measured using whole cell patch clamp recordings in neurons acutely isolated between embryonic day (E) 7 and E20, prior to hatching. E7 nodose neurons express relatively large high voltage-activated (HVA) Ca(2+) currents. HVA current density progressively increases between E7 and E17. T-type Ca(2+) currents were restricted to a few nodose neurons between E7 and E10 but were present in approximately 60% of nodose neurons by E17. T-type Ca(2+) channels regulate the response of nodose neurons to injection of hyperpolarizing currents, but do not have any effect on the action potential waveform. Nickel ions blocked T-type Ca(2+) currents in a concentration-dependent manner with an IC(50) of 17 microM. The high sensitivity of T-type Ca(2+) channels to nickel blockade combined with sequencing of a partial cDNA suggests that T-type Ca(2+) currents are generated by alpha1H subunits in chick nodose neurons. Steady-state activation and inactivation kinetics were similar to those previously reported for other alpha1H channels in mammalian neurons. Semi-quantitative PCR analysis indicates that alpha1H mRNA was present in chick nodose neurons by E7, suggesting that the functional expression of T-type Ca(2+) channels involves a posttranscriptional mechanism. These findings demonstrate a distinct pattern of T-type Ca(2+) channel functional expression in placode-derived neurons when compared with CNS neurons.
Thayer, S A; Miller, R J
1. Simultaneous whole-cell patch-clamp and Fura-2 microfluorimetric recordings of calcium currents (ICa) and the intracellular free Ca2+ concentration ([Ca2+]i) were made from neurones grown in primary culture from the dorsal root ganglion of the rat. 2. Cells held at -80 mV and depolarized to 0 mV elicited a ICa that resulted in an [Ca2+]i transient which was not significantly buffered during the voltage step and lasted long after the cell had repolarized and the current ceased. The process by which the cell buffered [Ca2+]i back to basal levels could best be described with a single-exponential equation. 3. The membrane potential versus ICa and [Ca2+]i relationship revealed that the peak of the [Ca2+]i transient evoked at a given test potential closely paralleled the magnitude of the ICa suggesting that neither voltage-dependent nor Ca2(+)-induced Ca2+ release from intracellular stores made a significant contribution to the [Ca2+]i transient. 4. When the cell was challenged with Ca2+ loads of different magnitude by varying the duration or potential of the test pulse, [Ca2+]i buffering was more effective for larger Ca2+ loads. The relationship between the integrated ICa and the peak of the [Ca2+]i transient reached an asymptote at large Ca2+ loads indicating that Ca2(+)-dependent processes became more efficient or that low-affinity processes had been recruited. 5. Inhibition of Ca2+ influx with neuropeptide Y demonstrated that inhibition of a large ICa produced minor alterations in the peak of the [Ca2+]i transient, while inhibition of smaller currents produced corresponding decreases in the [Ca2+]i transient. Thus, inhibition of the ICa was reflected by a change in the peak [Ca2+]i only when submaximal Ca2+ loads were applied to the cell, implying that modulation of [Ca2+]i is dependent on the activation state of the cells. 6. Intracellular dialysis with the mitochondrial Ca2+ uptake blocker Ruthenium Red in whole-cell patch-clamp experiments removed the buffering
Li, Li; Zhao, Lei; Wang, Yang; Ma, Ke-tao; Shi, Wen-yan; Wang, Ying-zi; Si, Jun-qiang
The mechanism underlying the modulatory effect of substance P (SP) on GABA-activated response in rat dorsal root ganglion (DRG) neurons was investigated. In freshly dissociated rat DRG neurons, whole-cell patch-clamp technique was used to record GABA-activated current and sharp electrode intracellular recording technique was used to record GABA-induced membrane depolarization. Application of GABA (1-1000 μmol/L) induced an inward current in a concentration-dependent manner in 114 out of 127 DRG neurons (89.8 %) examined with whole-cell patch-clamp recordings. Bath application of GABA (1-1000 μmol/L) evoked a depolarizing response in 236 out of 257 (91.8%) DRG neurons examined with intracellular recordings. Application of SP (0.001-1 μmol/L) suppressed the GABA-activated inward current and membrane depolarization. The inhibitory effects were concentration-dependent and could be blocked by the selective neurokinin 1 (NK1) receptors antagonist spantide but not by L659187 and SR142801 (1 μmol/L, n=7), selective antagonists of NK2 and NK3. The inhibitory effect of SP was significantly reduced by the calcium chelator BAPTA-AM, phospholipase C (PLC) inhibitor U73122, and PKC inhibitor chelerythrine, respectively. The PKA inhibitor H-89 did not affect the SP effect. Remarkably, the inhibitory effect of SP on GABA-activated current was nearly completely removed by a selective PKCε inhibitor epilon-V1-2 but not by safingol and LY333531, selective inhibitors of PKCα and PKCβ. Our results suggest that NK1 receptor mediates SP-induced inhibition of GABA-activated current and membrane depolarization by activating intracellular PLC-Ca²⁺-PKCε cascade. SP might regulate the excitability of peripheral nociceptors through inhibition of the "pre-synaptic inhibition" evoked by GABA, which may explain its role in pain and neurogenic inflammation.
Cai, Qi; Qiu, Chun-Yu; Qiu, Fang; Liu, Ting-Ting; Qu, Zu-Wei; Liu, Yu-Min; Hu, Wang-Ping
Extracellular acidosis is a common feature in pain-generating pathological conditions. Acid-sensing ion channels (ASICs), pH sensors, are distributed in peripheral sensory neurons and participate in nociception. Morphine exerts potent analgesic effects through the activation of opioid receptors for various pain conditions. A cross-talk between ASICs and opioid receptors in peripheral sensory neurons has not been shown so far. Here, we have found that morphine inhibits the activity of native ASICs in rat dorsal root ganglion (DRG) neurons. Morphine dose-dependently inhibited proton-gated currents mediated by ASICs in the presence of the TRPV1 inhibitor capsazepine. Morphine shifted the proton concentration-response curve downwards, with a decrease of 51.4±3.8% in the maximum current response but with no significant change in the pH0.5 value. Another μ-opioid receptor agonist DAMGO induced a similar decrease in ASIC currents compared with morphine. The morphine inhibition of ASIC currents was blocked by naloxone, a specific opioid receptor antagonist. Pretreatment of forskolin, an adenylyl cyclase activator, or the addition of cAMP reversed the inhibitory effect of morphine. Moreover, morphine altered acid-evoked excitability of rat DRG neurons and decreased the number of action potentials induced by acid stimuli. Finally, peripheral applied morphine relieved pain evoked by intraplantar of acetic acid in rats. Our results indicate that morphine can inhibit the activity of ASICs via μ-opioid receptor and cAMP dependent signal pathway. These observations demonstrate a cross-talk between ASICs and opioid receptors in peripheral sensory neurons, which was a novel analgesic mechanism of morphine.
Ng, KY; Yeung, BHS; Wong, YH; Wise, H
Background and Purpose Hyper-nociceptive PGE2 EP4 receptors and prostacyclin (IP) receptors are present in adult rat dorsal root ganglion (DRG) neurones and glial cells in culture. The present study has investigated the cell-specific expression of two other Gs-protein coupled hyper-nociceptive receptor systems: β-adrenoceptors and calcitonin gene-related peptide (CGRP) receptors in isolated DRG cells and has examined the influence of neurone–glial cell interactions in regulating adenylyl cyclase (AC) activity. Experimental Approach Agonist-stimulated AC activity was determined in mixed DRG cell cultures from adult rats and compared with activity in DRG neurone-enriched cell cultures and pure DRG glial cell cultures. Key Results Pharmacological analysis showed the presence of Gs-coupled β2-adrenoceptors and CGRP receptors, but not β1-adrenoceptors, in all three DRG cell preparations. Agonist-stimulated AC activity was weakest in DRG neurone-enriched cell cultures. DRG neurones inhibited IP receptor-stimulated glial cell AC activity by a process dependent on both cell–cell contact and neurone-derived soluble factors, but this is unlikely to involve purine or glutamine receptor activation. Conclusions and Implications Gs-coupled hyper-nociceptive receptors are readily expressed on DRG glial cells in isolated cell cultures and the activity of CGRP, EP4 and IP receptors, but not β2-adrenoceptors, in glial cells is inhibited by DRG neurones. Studies using isolated DRG cells should be aware that hyper-nociceptive ligands may stimulate receptors on glial cells in addition to neurones, and that variable numbers of neurones and glial cells will influence absolute measures of AC activity and affect downstream functional responses. PMID:22924655
Takeda, M; Ikeda, M; Tanimoto, T; Lipski, J; Matsumoto, S
The aim of the study was to examine the effects of alpha(2)-adrenoreceptor agonists on the excitability of trigeminal root ganglion (TRG) neurons using the perforated patch-clamp technique, and to determine whether these neurons express mRNA for alpha(2)-adrenoreceptors. In current-clamp mode, the resting membrane potential was -57.4+/-1.2 mV (n=26). Most neurons (71%) were hyperpolarized by clonidine (5-50 microM) in a concentration-dependent manner. The response was associated with an increase of cell input resistance. In addition, clonidine reduced the repetitive firing evoked by depolarizing current pulses. An alpha(2)-adrenergic agonist, UK14,304, (10-20 microM) also hyperpolarized TRG neurons. The clonidine- and UK14,304-induced hyperpolarization was blocked by idazoxan (alpha(2)-adrenoreceptor antagonist). In voltage-clamp, clonidine (1-50 microM) reversibly reduced the hyperpolarization- and time-dependent cationic current. The effect was mimicked by UK14,304 (10-20 microM), and antagonized by idazoxan. Hyperpolarization-activated cationic current was blocked by extracellular Cs(+) (2 mM) or a specific blocker, ZD7288 (20 microM). Analysis of tail currents revealed that a reversal potential of the clonidine-sensitive component of hyperpolarization-activated cationic current was -46 mV. Single-cell reverse transcription-polymerase chain reaction analysis demonstrated the expression of mRNA for alpha(2A)- and alpha(2C)-adrenoreceptors. These results demonstrate that activation of alpha(2)-adrenoreceptors can hyperpolarize TRG neurons, and that the inhibitory effect is associated with inhibition of hyperpolarization-activated cationic current. Our results suggest that activation of alpha(2)-adrenoreceptors in the absence of nerve injury may have an inhibitory effect on nociceptive transmission in the trigeminal system at the level of both TRG neuronal cell bodies and primary afferent terminals.
Zhao, Yuanting; Qin, Yue; Liu, Tuanjiang; Hao, Dingjun
Neuropathic pain, which is characterized by hyperalgesia, allodynia and spontaneous pain, is one of the most painful symptoms that can be experienced in the clinic. It often occurs as a result of injury to the peripheral nerves, dorsal root ganglion (DRG), spinal cord or brain. The renin-angiotensin system (RAS) plays an important role in nociception. As an essential component of the RAS, the angiotensin (Ang)-(1-7)/Mas axis may be involved in antinociception. The aim of the present study was to explore the expression pattern of Mas in DRG neurons following chronic nerve injury and examine the effects of Mas inhibition and activation on neuropathic pain in a chronic constriction injury (CCI) rat model. The results showed, that compared with the sham group, CCI caused a time-dependent induction of Mas expression at both the mRNA and the protein levels in DRG neurons. Consistent with the results, isolated DRG neurons showed a time-dependent increase in Ang-(1-7) binding on the cell membrane following the CCI surgery, but not the sham surgery. Compared with the sham control groups, CCI significantly decreased the paw withdrawal latency and threshold, and this was markedly improved and aggravated by intrathecal injection of the selective Mas agonist Ang-(1-7) and the selective Mas inhibitor D-Pro7-Ang-(1-7), respectively. In conclusion, this study has provided the first evidence, to the best of our knowledge, that the Mas expression in DRG neurons is time-dependently induced by chronic nerve injury and that the intrathecal activation and inhibition of Mas can improve and aggravate CCI-induced neuropathic pain, respectively. This study has provided novel insights into the pathophysiological process of neuropathic pain and suggests that the Ang-(1-7)/Mas axis could be an effective therapeutic target for neuropathic pain, warranting further study.
Bevan, S; Yeats, J
1. The responses of adult and neonatal rat dorsal root ganglion (DRG) neurones to buffered acidic solutions were studied with both voltage clamp and radioactive ion flux techniques. Electrophysiological experiments were made on acutely isolated neurones and ion flux experiments were made on cells that had been in culture for 3-6 days. 2. Acid solutions of pH < 6.2 evoked a sustained, slowly inactivating inward current in neurones voltage clamped at negative holding potentials. The size of the current increased with increasing proton concentrations. This response was restricted to a sub-population (approximately 45%) of adult and neonatal rat DRG neurones and was distinct from a rapidly activating and inactivating proton-induced inward sodium current that was also found in DRG neurones. 3. The proton-activated sustained current was due to an increase in cation conductance that allowed K+, Cs+ and Na+ to pass with PK/PNa = 1.32 and PCs/PNa = 1.12. 4. Radioactive ion efflux experiments made on neonatal rat cultured DRG neurones showed that protons also increased the permeability to both [14C]guanidinium and 86Rb+ ions. The half-maximal increase in efflux rate for 86Rb+ occurred at pH 5.8. Acid solution also stimulated the efflux of 86Rb+ in cultures of adult rat neurones. 5. Cells that showed a late, sustained proton-activated current also responded to capsaicin. In addition, no proton-activated fluxes of either [14C]guanidinium or 86Rb+ ions were observed in cultures of DRG neurones that had been treated with high concentrations of capsaicin (10 microM) to kill the capsaicin-sensitive neurones. Thus this proton-activated current is restricted largely, if not exclusively, to capsaicin-sensitive peripheral sensory neurones. PMID:1726795
1. The properties of gamma-aminobutyric acid (GABA)-activated chloride channels in dorsal root ganglion (DRG) neurones obtained from rats and cats were examined using the single-electrode voltage clamp in conjunction with suction-electrode techniques. 2. GABA-evoked currents showed voltage-sensitive kinetics. Time constants (tau D) were measured from voltage-jump relaxations and tau D became briefer with membrane hyperpolarization. tau D was 33 ms at -120 mV with 60 microM-GABA and changed e-fold for 188 mV. tau D decreased as GABA concentration was increased - the extrapolated tau D at 'zero' GABA concentration was approximately equal to 50 ms at -120 mV. 3. The steady-state current in GABA was curvilinear, rectifying at negative potentials. The instantaneous current was linear with symmetrical chloride concentrations (140 mM) on both sides of the cell membrane. 4. Muscimol was a more effective agonist than GABA, while piperidine-4-sulphonic acid and ethylenediamine monocarbamate were only weakly effective agonists. Taurine and glycine had no detectable agonist activity. 5. Ion substitution experiments revealed the permeability sequence I- greater than Br- greater than Cl- greater than F- greater than propionate (1.88 greater than 1.21 greater than 1.0 approximately equal to 0.1 approximately equal to 0.1). 6. The presence of iodide and bromide ions externally caused an increase in chloride efflux at membrane potentials more negative than -40 mV, and caused a prolongation of voltage-jump relaxations. Relaxations in fluoride and propionate solutions were faster than those seen in chloride.
Mizuta, Kotaro; Fujita, Tsugumi; Kumamoto, Eiichi
Although opioids inhibit action potential (AP) conduction in primary-afferent fibers, this has not yet been fully examined. We investigated by using the sharp glass microelectrode technique how opioids (morphine, codeine, and ethylmorphine) affect APs recorded from adult rat dorsal root ganglion (DRG) neurons in response to sciatic nerve stimulation. The DRG neurons were classified into three types, Aα/β, Aδ, and C, according to AP characteristics, including the fiber conduction velocity (CV) of the neuron. AP of the Aα/β neuron was reduced in peak amplitude by each of the opioids in a reversible and concentration-dependent manner. The potency sequence was ethylmorphine > codeine = morphine (IC(50) = 0.70, 2.5, and 2.9 mM, respectively), indicating that this AP inhibition is related to the chemical structure of the opioid. Each of the Aδ and C neuron APs was also inhibited by the opioids; ethylmorphine had a tendency to inhibit APs more effectively than codeine and morphine. This inhibition was variable in extent among neurons and was either comparable to or greater than that of the Aα/β neuron AP. The opioid-induced AP inhibitions were unaffected by nonspecific opioid-receptor antagonist naloxone; opioid-receptor agonists did not affect APs. In conclusion, the opioids inhibited APs in DRG neurons without opioid-receptor activation; this inhibition was different among neurons having different primary-afferent fiber CVs and also among the three kinds of opioid. The inhibition by opioid of primary-afferent fiber AP conduction is suggested to be distinct in extent among fibers conveying distinct types of nociceptive information. Copyright © 2012 Wiley Periodicals, Inc.
Li, Zhisong; Gu, Xiyao; Sun, Linlin; Wu, Shaogen; Liang, Lingli; Cao, Jing; Lutz, Brianna Marie; Bekker, Alex; Zhang, Wei; Tao, Yuan-Xiang
Peripheral nerve injury-induced changes in gene transcription and translation in primary sensory neurons of the dorsal root ganglion (DRG) are considered to contribute to neuropathic pain genesis. Transcription factors control gene expression. Peripheral nerve injury increases the expression of myeloid zinc finger protein 1 (MZF1), a transcription factor, and promotes its binding to the voltage-gated potassium 1.2 (Kv1.2) antisense (AS) RNA gene in the injured DRG. However, whether DRG MZF1 participates in neuropathic pain is still unknown. Here, we report that blocking the nerve injury-induced increase of DRG MZF1 through microinjection of MZF1 siRNA into the injured DRG attenuated the initiation and maintenance of mechanical, cold, and thermal pain hypersensitivities in rats with chronic constriction injury (CCI) of the sciatic nerve, without affecting locomotor functions and basal responses to acute mechanical, heat, and cold stimuli. Mimicking the nerve injury-induced increase of DRG MZF1 through microinjection of recombinant adeno-associated virus 5 expressing full-length MZF1 into the DRG produced significant mechanical, cold, and thermal pain hypersensitivities in naive rats. Mechanistically, MZF1 participated in CCI-induced reductions in Kv1.2 mRNA and protein and total Kv current and the CCI-induced increase in neuronal excitability through MZF1-triggered Kv1.2 AS RNA expression in the injured DRG neurons. MZF1 is likely an endogenous trigger of neuropathic pain and might serve as a potential target for preventing and treating this disorder.
Lu, Shao-Gang; Zhang, Xiulin; Gold, Michael S
Primary afferent neurons are functionally heterogeneous. To determine whether this functional heterogeneity reflects, in part, heterogeneity in the regulation of the concentration of intracellular Ca2+ ([Ca2+]i), the magnitude and decay of evoked Ca2+ transients were assessed in subpopulations of dorsal root ganglion (DRG) neurons with voltage clamp and fura-2 ratiometric imaging. To determine whether differences in evoked Ca2+ transients among subpopulations of DRG neurons reflected differences in the contribution of Ca2+ regulatory mechanisms, pharmacological techniques were employed to assess the contribution of influx, efflux, release and uptake pathways. Subpopulations of DRG neurons were defined by cell body size, binding of the plant lectin IB4 and responsiveness to the algogenic compound capsaicin (CAP). Ca2+ transients were evoked with 30 mm K+ or voltage steps to 0 mV. There were marked differences between subpopulations of neurons with respect to both the magnitude and decay of the Ca2+ transient, with the largest and most slowly decaying Ca2+ transients in small-diameter, IB4-positive, CAP-responsive neurons. The smallest and most rapidly decaying transients were in large-diameter, IB4-negative and CAP-unresponsive DRG neurons. These differences were not due to a differential distribution of voltage-gated Ca2+ currents. However, these differences did appear to reflect a differential contribution of other influx, efflux, release and uptake mechanisms between subpopulations of neurons. These results suggest that electrical activity in subpopulations of DRG neurons will have a differential influence on Ca2+-regulated phenomena such as spike adaptation, transmitter release and gene transcription. Significantly more activity should be required in large-diameter non-nociceptive afferents than in small-diameter nociceptive afferents to have a comparable influence on these processes. PMID:16945973
Lynch, Paul J; McJunkin, Tory; Eross, Eric; Gooch, Stacie; Maloney, Jillian
Postherpetic neuralgia (PHN) is the most common complication following an acute varicella zoster virus infection. PHN often results in a chronic severe pain condition refractory to conservative pain management treatments. Peripheral nerve stimulation over the affected spinal nerve root may be an effective treatment option for patients with intractable PHN. To describe a successful case of peripheral nerve stimulation of the second cervical dorsal root ganglion for the treatment of intractable PHN. An 80-year-old man with a 15-month history of severe PHN was referred to our clinic for pain management. His pain was localized to the left side in the distribution of the C2 dermatome. The patient's pain was unresponsive to comprehensive conventional treatments for PHN including physical therapy, membrane stabilizing medications, opioids, anti-inflammatories, cervical epidural steroid injections, cervical facet joint injections, and dorsal root ganglion blockade with pulsed radiofrequency. After failing to respond to conservative and interventional therapies, a peripheral nerve stimulator trial was conducted for a period of seven days. The lead was placed within the epidural space over the atlanto-axial joint under fluoroscopy to stimulate the left C2 nerve root. This trial resulted in a significant decrease of the patient's pain, and discontinuation of all pain medications. We describe a case of successful electrode placement at the C2 spinal level for the treatment of refractory PHN. © 2010 International Neuromodulation Society.
Tang, Zhanying; Cui, Xuejun; Hu, Zhijun; Xiao, Jing; Li, Weiwei; Yang, Qiangling; Liu, Dan; Lin, Jie; Wang, Yongjun; Shi, Qi
The aim of this study was to observe the effects of Yiqi Huayu recipe on TRPV4 expression in radicular neuralgia model induced by chronic constriction to the rat lumber nerve root. Healthy male SD rats were divided into 3 groups for radicular neuralgia (RN) model construction: the sham operation group, model groups (day 3, 7, 14 and 28), and medication groups (day 3, 7, 14 and 28). Von-Frey hairs test was performed to detect the 50% with drawal threshold (50% TPW) for rats of each group. The expression of TRPV4 in dorsal root ganglion was detected at both mRNA and protein level. Rats from all model groups displayed hyperalgesia with significantly reduced 50% TPW values compared with sham-operation group (P<0.01); Yiqi Huayu recipe medication groups showed higher 50% TPW than model group since 7 days post medication (P<0.01); the medication groups showed decreased TRPV4 expression than that of model groups (P<0.01). In conclusion, Yiqi Huayu recipe alleviates nerve root constriction induced radicular neuralgia by repressing TRPV4 expression in dorsal root ganglion. PMID:26770465
Saijilafu; Zhou, Feng-Quan
It is well known that mature neurons in the central nervous system (CNS) cannot regenerate their axons after injuries due to diminished intrinsic ability to support axon growth and a hostile environment in the mature CNS(1,2). In contrast, mature neurons in the peripheral nervous system (PNS) regenerate readily after injuries(3). Adult dorsal root ganglion (DRG) neurons are well known to regenerate robustly after peripheral nerve injuries. Each DRG neuron grows one axon from the cell soma, which branches into two axonal branches: a peripheral branch innervating peripheral targets and a central branch extending into the spinal cord. Injury of the DRG peripheral axons results in substantial axon regeneration, whereas central axons in the spinal cord regenerate poorly after the injury. However, if the peripheral axonal injury occurs prior to the spinal cord injury (a process called the conditioning lesion), regeneration of central axons is greatly improved(4). Moreover, the central axons of DRG neurons share the same hostile environment as descending corticospinal axons in the spinal cord. Together, it is hypothesized that the molecular mechanisms controlling axon regeneration of adult DRG neurons can be harnessed to enhance CNS axon regeneration. As a result, adult DRG neurons are now widely used as a model system to study regenerative axon growth(5-7). Here we describe a method of adult DRG neuron culture that can be used for genetic study of axon regeneration in vitro. In this model adult DRG neurons are genetically manipulated via electroporation-mediated gene transfection(6,8). By transfecting neurons with DNA plasmid or si/shRNA, this approach enables both gain- and loss-of-function experiments to investigate the role of any gene-of-interest in axon growth from adult DRG neurons. When neurons are transfected with si/shRNA, the targeted endogenous protein is usually depleted after 3-4 days in culture, during which time robust axon growth has already occurred
Alvarez-Leefmans, F J; Gamiño, S M; Giraldez, F; Noguerón, I
1. Intracellular Cl- activity (aiCl) and membrane potential (Em) were measured in frog dorsal root ganglion neurones (DRG neurones) using double-barrelled Cl- -selective microelectrodes. In standard Ringer solution buffered with HEPES (5 mM), equilibrated with air or 100% O2, the resting membrane potential was -57.7 +/- 1.0 mV and aiCl was 23.6 +/- 1.0 mM (n = 53). The value of aiCl was 2.6 times the activity expected for an equilibrium distribution and the difference between Em and ECl was 25 mV. 2. Removal of external Cl- led to a reversible fall in aiCl. Initial rates of decay and recovery of aiCl were 4.1 and 3.3 mM min-1, respectively. During the recovery of aiCl following return to standard Ringer solution, most of the movement of Cl- occurred against the driving force for a passive distribution. Changes in aiCl were not associated with changes in Em. Chloride fluxes estimated from initial rates of change in aiCl when external Cl- was removed were too high to be accounted for by electrodiffusion. 3. The intracellular accumulation of Cl- was dependent on the extracellular Cl- activity (aoCl). The relationship between aiCl and aoCl had a sigmoidal shape with a half-maximal activation of about 50 mM-external Cl-. 4. The steady-state aiCl depended on the simultaneous presence of extracellular Na+ and K+. Similarly, the active reaccumulation of Cl- after intracellular Cl- depletion was abolished in the absence of either Na+ or K+ in the bathing solution. 5. The reaccumulation of Cl- was inhibited by furosemide (0.5-1 x 10(-3) M) or bumetanide (10(-5) M). The decrease in aiCl observed in Cl- -free solutions was also inhibited by bumetanide. 6. Cell volume changes were calculated from the observed changes in aiCl. Cells were estimated to shrink in Cl- -free solutions to about 75% their initial volume, at an initial rate of 6% min-1. 7. The present results provide direct evidence for the active accumulation of Cl- in DRG neurones. The mechanism of Cl- transport is
Mattila, R K; Harila, K; Kangas, S M; Paavilainen, H; Heape, A M; Mohr, I J; Hukkanen, V
After a primary lytic infection at the epithelia, herpes simplex virus type 1 (HSV-1) enters the innervating sensory neurons and translocates to the nucleus, where it establishes a quiescent latent infection. Periodically, the virus can reactivate and the progeny viruses spread back to the epithelium. Here, we introduce an embryonic mouse dorsal root ganglion (DRG) culture system, which can be used to study the mechanisms that control the establishment, maintenance and reactivation from latency. Use of acyclovir is not necessary in our model. We examined different phases of the HSV-1 life cycle in DRG neurons, and showed that WT HSV-1 could establish both lytic and latent form of infection in the cells. After reactivating stimulus, the WT viruses showed all markers of true reactivation. In addition, we showed that deletion of the γ(1)34.5 gene rendered the virus incapable of reactivation, even though the virus was clearly able to replicate and persist in a quiescent form in the DRG neurons.
Purkiss, J; Welch, M; Doward, S; Foster, K
Capsaicin, the pungent component of "hot" chili peppers, selectively activates a distinct population of primary sensory neurons responsive to noxious stimuli. Many of these fibres express neuropeptides including the tachykinin, substance P. Using cultured dorsal root ganglion neurons, we found that capsaicin (10 microM) stimulated a 2-fold increase in release of substance P in the absence of extracellular Ca(2+). Elevated potassium (75 mM) was unable to induce release under these conditions. The introduction of Ca(2+) enhanced capsaicin-induced release and brought about a robust response to potassium. Preincubation of cells with botulinum neurotoxin A (100 nM) completely blocked potassium-induced release but the capsaicin response, in the absence of Ca(2+), was unaffected. However, toxin treatment dramatically reduced capsaicin-stimulated release in the presence of Ca(2+). It is concluded that capsaicin induces release of substance P from dorsal root ganglion neurons via two mechanisms, one requiring extracellular Ca(2+) and the intact synaptosomal-associated protein 25 kDa (SNAP-25), and the other independent of extracellular Ca(2+) and not involving SNAP-25.
Huang, Yimei; Yang, Hongqin; Chen, Jiangxu; Shen, Xiuqiu; Zheng, Liqin; Wang, Yuhua; Xie, Shusen
As one of the most important second messengers, calcium in nerve cells plays a critical role in neuronal processes, including excitability, neurotransmitter release, synaptic plasticity. Modulation of the calcium concentration is an important means of regulating diverse neuronal functions. To evaluate the role of calcium, quantitative measurement of cytosolic free calcium concentrations is necessary. There are several optical techniques that are available for measurement of calcium in live cells. Laser scanning confocal microscopy and two-photon microscopy are two prevalent techniques for their advantage in spatial resolution. In this paper, calcium in dorsal root ganglion neurons was imaged by laser scanning confocal microscopy and two-photon microscopy with Fluo-3, a calcium specific fluorescence probe. Both of spatial resolution and photobleaching, two common limitations of optical image modality, were compared between laser scanning confocal microscopy and two-photon microscopy, respectively. Three dimension images showed that laser scanning confocal microscopy and two-photon microscopy had not only similar lateral resolution but also parallel vertical resolution. However, Laser scanning confocal microscopy had an advantage over the two-photon microcopy in photobleaching. These results indicated that laser scanning confocal microscopy was more suitable than two-photon microscopy to be applied in imaging calcium in dorsal root ganglion neurons with Fluo-3.
Ma, Zhong; Hasenstein, Karl H
Vertical orientation of emerging roots typically is the first response of plants to gravity. Although root gravitropism has been studied extensively, no conclusive data on the onset of gravisensing exist. We determined the inception of gravisensitivity in flax (Linum usitatissimum) roots by clinorotating germinating seeds after various periods of static orientation (gravistimulation) of imbibed seeds. Gravitropic competency was established about 8 h after imbibition, 11 h prior to germination. The time was determined based on 50% of the newly emerged roots curving in the direction of the gravity vector during static imbibition, despite subsequent clinorotation. The threshold value was affected by the orientation of the seeds. Upward orientation of the micropyle/radicle reduced the number of graviresponding roots to about one-half. Prolonged clinorotation weakened the graviresponse. Gravisensing was accompanied by the development of amyloplasts, but the actin cytoskeleton was not involved because imbibition in Latrunculin B did not affect the onset of gravisensitivity or germination, and the development of F-actin in untreated controls was observed only after the onset of gravisensitivity.
Qu, Yu-Juan; Jia, Lei; Zhang, Xiao; Wei, Hui; Yue, Shou-Wei
The aim of the present study was to investigate whether the MAPK pathways were involved in the mechanism of neuropathic pain in rats with chronic compression of the dorsal root ganglion. We determined the paw withdrawal mechanical threshold (PWMT) of rats before and after CCD surgery and then after p38, JNK, or ERK inhibitors administration. Western blotting, RT-PCR, and immunofluorescence of dorsal root ganglia were performed to investigate the protein and mRNA level of MAPKs and also the alternation in distributions of positive neurons in dorsal root ganglia. Intrathecal administration of MAPKs inhibitors, SB203580 (p38 inhibitor), SP600125 (JNK inhibitor), and U0126 (ERK inhibitor), resulted in a partial reduction in CCD-induced mechanical allodynia. The reduction of allodynia was associated with significant depression in the level of both MAPKs mRNA and protein expression in CCD rats and also associated with the decreased ratios of large size MAPKs positive neurons in dorsal root ganglia. In conclusion, the specific inhibitors of MAPKs contributed to the attenuation of mechanical allodynia in CCD rats and the large size MAPKs positive neurons in dorsal root ganglia were crucial.
Zuidema, X.; Breel, J.; Wille, F.
Chronic perineal pain limits patients in physical and sexual activities, leading to social and psychological distress. In most cases, this pain develops after surgery in the urogenital area or as a consequence of trauma. Neuromodulation is one of the options in chronic postsurgical perineal pain treatment. We present a case of refractory perineal pain after right sided surgical resection of a Bartholin's cyst which was treated with third sacral nerve root/dorsal root ganglion stimulation using the transforaminal approach. We describe a new anchorless lead placement technique using a unique curved lead delivery sheath. We postulate that this new posterior foraminal technique of lead placement is simple, safe, and reversible and may lower the occurrence of lead related complications. PMID:27123351
Zhou, Yu; Zhao, Zhi-Qi
High-threshold Ca(2+) channels and tetrodotoxin-resistant Na(+) channels are highly expressed in small dorsal root ganglion neurons. In acutely isolated rat dorsal root ganglion neurons, the effects of neomycin, one of the aminoglycoside antibiotics, on high-threshold Ca(2+) currents and tetrodotoxin-resistant Na(+) currents were examined using whole-cell patch recording. We showed for the first time that neomycin dose-dependently inhibited peak high-threshold Ca(2+) currents and peak tetrodotoxin-resistant Na(+) currents with half-maximal inhibitory concentrations at 3.69 microM (n=20) and 1213.44 microM (n=25), respectively. Inactivation properties of high-threshold Ca(2+) currents and activation properties of tetrodotoxin-resistant Na(+) currents were also affected by neomycin with reduction of excitability of small dorsal root ganglion neurons. Half-maximal inactivation voltage of high-threshold Ca(2+) currents was -45.56 mV before and -50.46 mV after application of neomycin (n=10). Half-maximal activation voltage of tetrodotoxin-resistant Na(+) currents was -19.93 mV before and -11.19 mV after administration of neomycin (n=15). These results suggest that neomycin can inhibit high-threshold Ca(2+) currents and tetrodotoxin-resistant Na(+) currents in small dorsal root ganglion neurons, which may contribute to neomycin-induced peripheral and central analgesia.
Zheng, Li-qin; Wang, Yu-hua; He, Yi-peng; Zhou, Jie; Yang, Hong-qin; Zhang, Yan-ding; Xie, Shu-sen
Nitric oxide (NO) and nitric oxide synthase (NOS) have an important role in pain signaling transmission in animal models. Low-level laser therapy (LLLT) is known to have an analgesic effect, but the mechanism is unclear. The aim of the study is to investigate the influence of LLLT on NO release and NOS synthesis in dorsal root ganglion (DRG) neurons, in order to find whether LLLI can ameliorate pain through modulating NO production at the cellular level. The results show that in stress conditions, the laser irradiation at 658 nm can modulate NO production in DRG neurons with soma diameter of about 20 μm in a short time after illumination, and affect NOS synthesis in a dose-dependent manner. It is demonstrated that LLLT might treat pain by altering NO release directly and indirectly in DRG neurons.
Kikuchi, Shin; Ninomiya, Takafumi; Kohno, Takayuki; Kojima, Takashi; Tatsumi, Haruyuki
Cobalt is a trace element that localizes in the human body as cobalamin, also known as vitamin B12. Excessive cobalt exposure induces a peripheral neuropathy, the mechanisms of which are yet to be elucidated. We investigated how cobalt may affect mitochondrial motility in primary cultures of rat dorsal root ganglion (DRG). We observed mitochondrial motility by time-lapse imaging after DsRed2 tagging via lentivirus, mitochondrial structure using transmission electron microscopy (TEM), and axonal swelling using immunocytochemical staining. The concentration of cobaltous ion (Co(2+)) required to significantly suppress mitochondrial motility is lower than that required to induce axonal swelling following a 24-h treatment. Exposure to relatively low concentrations of Co(2+) for 48 h suppressed mitochondrial motility without leading to axonal swelling. TEM images indicated that Co(2+) induces mitochondrial destruction. Our results show that destruction of the axonal mitochondria precedes the axonal degeneration induced by Co(2+) exposure.
Murata, Ryo; Ohtori, Seiji; Ochiai, Nobuyasu; Takahashi, Norimasa; Saisu, Takashi; Moriya, Hideshige; Takahashi, Kazuhisa; Wada, Yuichi
Although extracorporeal shockwave has been applied in the treatment of various diseases, the biological basis for its analgesic effect remains unclear. Therefore, we investigated the dorsal root ganglion neurons of rats following shockwave exposure to the footpad to elucidate its effect on the peripheral nervous system. We used activating transcription factor 3 (ATF3) and growth-associated phosphoprotein (GAP-43) as markers for nerve injury and axonal regeneration, respectively. The average number of neurons immunoreactive for ATF3 increased significantly in the treated rats at all experimental time points, with 78.3% of those neurons also exhibiting immunoreactivity for GAP-43. Shockwave exposure induced injury of the sensory nerve fibers within the exposed area. This phenomenon may be linked to the desensitization of the exposure area, not the cause of pain, considering clinical research with a particular absence of painful adverse effect. Subsequent active axonal regeneration may account for the reinnervation of exposed area and the amelioration of the desensitization.
Madelaine, Romain; Busolin, Giorgia; Nikaido, Masataka; Colanesi, Sarah; Camargo-Sosa, Karen; Toppo, Stefano; Blader, Patrick; Tiso, Natascia; Kelsh, Robert N.
The development of functional peripheral ganglia requires a balance of specification of both neuronal and glial components. In the developing dorsal root ganglia (DRGs), these components form from partially-restricted bipotent neuroglial precursors derived from the neural crest. Work in mouse and chick has identified several factors, including Delta/Notch signaling, required for specification of a balance of these components. We have previously shown in zebrafish that the Sry-related HMG domain transcription factor, Sox10, plays an unexpected, but crucial, role in sensory neuron fate specification in vivo. In the same study we described a novel Sox10 mutant allele, sox10baz1, in which sensory neuron numbers are elevated above those of wild-types. Here we investigate the origin of this neurogenic phenotype. We demonstrate that the supernumerary neurons are sensory neurons, and that enteric and sympathetic neurons are almost absent just as in classical sox10 null alleles; peripheral glial development is also severely abrogated in a manner similar to other sox10 mutant alleles. Examination of proliferation and apoptosis in the developing DRG reveals very low levels of both processes in wild-type and sox10baz1, excluding changes in the balance of these as an explanation for the overproduction of sensory neurons. Using chemical inhibition of Delta-Notch-Notch signaling we demonstrate that in embryonic zebrafish, as in mouse and chick, lateral inhibition during the phase of trunk DRG development is required to achieve a balance between glial and neuronal numbers. Importantly, however, we show that this mechanism is insufficient to explain quantitative aspects of the baz1 phenotype. The Sox10(baz1) protein shows a single amino acid substitution in the DNA binding HMG domain; structural analysis indicates that this change is likely to result in reduced flexibility in the HMG domain, consistent with sequence-specific modification of Sox10 binding to DNA. Unlike other Sox10
Jiao, Ling; Zhang, Tengda; Wang, Huixing; Zhang, Wenyi; Fan, Saijun; Huo, Xiaodong; Zheng, Baosen; Ma, Wenting
The use of iodine-125 ((125)I) in cancer treatment has been shown to relieve patients' pain. Considering dorsal root ganglia are critical for neural transmission between the peripheral and central nervous systems, we assumed that (125)I could be implanted into rat dorsal root ganglia to provide relief for neuropathic pain. (125)I seeds with different radioactivity (0, 14.8, 29.6 MBq) were implanted separately through L4-5 and L5-6 intervertebral foramen into the vicinity of the L5 dorsal root ganglion. von Frey hair results demonstrated the mechanical pain threshold was elevated after implanting (125)I seeds from the high radioactivity group. Transmission electron microscopy revealed that nuclear membrane shrinkage, nucleolar margination, widespread mitochondrial swelling, partial vacuolization, lysosome increase, and partial endoplasmic reticulum dilation were visible at 1,440 hours in the low radioactivity group and at 336 hours in the high radioactivity group. Abundant nuclear membrane shrinkage, partial fuzzy nuclear membrane and endoplasmic reticulum necrosis were observed at 1,440 hours in the high radioactivity group. No significant difference in combined behavioral scores was detected between preoperation and postoperation in the low and high radioactivity groups. These results suggested that the mechanical pain threshold was elevated after implanting (125)I seeds without influencing motor functions of the hind limb, although cell injury was present.
Wen, Xianjie; Lai, Xiaohong; Li, Xiaohong; Zhang, Tao; Liang, Hua
In this study, we identified the subtype of Calcium/calmodulin-dependent protein kinase II (CaMK II) mRNA in dorsal root ganglion neurons and observed the effects of ropivacaine hydrochloride in different concentration and different exposure time on the mRNA expression. Dorsal root ganglion neurons were isolated from the SD rats and cultured in vitro. The mRNA of the CaMK II subtype in dorsal root ganglion neurons were detected by real-time PCR. As well as, the dorsal root ganglion neurons were treated with ropivacaine hydrochloride in different concentration (1mM,2mM, 3mM and 4mM) for the same exposure time of 4h, or different exposure time (0h,2h,3h,4h and 6h) at the same concentration(3mM). The changes of the mRNA expression of the CaMK II subtype were observed with real-time PCR. All subtype mRNA of the CaMK II, CaMK IIα, CaMK IIβ, CaMK II δ, CaMK IIγ, can be detected in dorsal root ganglion neurons. With the increased of the concentration and exposure time of the ropivacaine hydrochloride, all the subtype mRNA expression increased. Ropivacaine hydrochloride up-regulate the CaMK IIβ, CaMK IIδ, CaMK IIg mRNA expression with the concentration and exposure time increasing. The nerve blocking or the neurotoxicity of the ropivacaine hydrochloride maybe involved with CaMK II.
Leffler, Andreas; Linte, Ramona Madalina; Nau, Carla; Reeh, Peter; Babes, Alexandru
Heat-activated ion channels from the vanilloid-type TRP group (TRPV1-4) seem to be central for heat-sensitivity of nociceptive sensory neurons. Displaying a high-threshold (> 52 degrees C) for activation, TRPV2 was proposed to act as a sensor for intense noxious heat in mammalian sensory neurons. However, although TRPV2 is expressed in a distinct population of thinly myelinated primary afferents, a widespread expression in a variety of neuronal and non-neuronal tissues suggests a more diverse physiological role of TRPV2. In its role as a heat-sensor, TRPV2 has not been thoroughly characterized in terms of biophysical and pharmacological properties. In the present study, we demonstrate that the features of heterologously expressed rat TRPV2 closely resemble those of high-threshold heat-evoked currents in medium- and large-sized capsaicin-insensitive rat dorsal root ganglion (DRG) neurons. Both in TRPV2-expressing human embryonic kidney (HEK)293t cells and in DRGs, high-threshold heat-currents were sensitized by repeated activation and by the TRPV1-3 agonist, 2-aminoethoxydiphenyl borate (2-APB). In addition to a previously described block by ruthenium red, we identified the trivalent cations, lanthanum (La(3+)) and gadolinium (Gd(3+)) as potent blockers of TRPV2. Thus, we present a new pharmacological tool to distinguish between heat responses of TRPV2 and the closely related capsaicin-receptor, TRPV1, which is strongly sensitized by trivalent cations. We demonstrate that self-sensitization of heat-evoked currents through TRPV2 does not require extracellular calcium and that TRPV2 can be activated in cell-free membrane patches in the outside-out configuration. Taken together our results provide new evidence for a role of TRPV2 in mediating high-threshold heat responses in a subpopulation of mammalian sensory neurons.
Niimura, Yuri; Aminaka, Yuichi; Hayashi, Kensuke
Migrating neurons have leading processes that direct cell movement in response to guidance cues. We investigated the involvement of glycogen synthase kinase 3 (GSK3) in the formation of leading processes and migration of neurons in vitro. We used embryonic rat medial ganglionic eminence (MGE) neurons, which are precursors of inhibitory neurons that migrate into the cerebral cortex. When MGE neurons were placed on an astrocyte layer, they migrated freely with the highest speed among neurons from other parts of the embryonic forebrain. When they were cultured alone, they showed bipolar morphology and extended leading processes within 20 h. Their leading processes had large growth cones, but did not elongate during 3 days in culture, indicating that leading processes are distinct from short axons. Next, we examined the effect of GSK3 inhibitors on leading processes and the migratory behavior of MGE neurons. MGE neurons treated with GSK3 inhibitors showed multipolar morphology and altered process shapes. Moreover, migration of MGE neurons on the astrocyte layer was significantly decreased in the presence of GSK3 inhibitors. These data suggest that GSK3 is involved in the formation of leading processes and in the migration of MGE neurons.
Imani, Farnad; Gharaei, Helen; Rezvani, Mehran
Chronic pain following lower-limb amputation is now a well-known neuropathic, chronic-pain syndrome that usually presents as a combination of phantom and stump pain. Controlling these types of neuropathic pain is always complicated and challenging. If pharmacotherapy does not control the patient's pain, interventional procedures have to be taken. The aim of this study was to evaluate the efficacy of using pulsed radiofrequency (PRF) on the dorsal root ganglia at the L4 and L5 nerve roots to improve phantom pain. Two patients with phantom pain were selected for the study. After a positive response to segmental nerve blockade at the L4 and L5 nerve roots, PRF was performed on the L4 and L5 dorsal root ganglia. Global clinical improvement was good in one patient, with a 40% decrease in pain on the visual analogue scale (VAS) in 6 months, and moderate in the second patient, with a 30% decrease in pain scores in 4 months. PRF of the dorsal root ganglia at the L4 and L5 nerve roots may be an effective therapeutic option for patients with refractory phantom pain.
Cuellar, J M; Montesano, P X; Antognini, J F; Carstens, E
Herniation of the nucleus pulposus (NP) from lumbar intervertebral discs commonly results in radiculopathic pain possibly through a neuroinflammatory response. NP sensitizes dorsal horn neuronal responses, but it is unknown whether this reflects a central or peripheral sensitization. To study central sensitization, we tested if NP enhances windup--the progressive increase in the response of a nociceptive spinal neuron to repeated electrical C-fiber stimulation--a phenomenon that may partly account for temporal summation of pain. Single-unit recordings were made from wide dynamic range (WDR; n = 36) or nociceptive-specific (NS; n = 8) L5 dorsal horn neurons in 44 isoflurane-anesthetized rats. Subcutaneous electrodes delivered electrical stimuli (20 pulses, 3 times the C-fiber threshold, 0.5 ms) to the receptive field on the hindpaw. Autologous NP was harvested from a tail disc and placed onto the L5 dorsal root ganglion after recording of baseline responses (n = 22). Controls had saline applied similarly (n = 22). Electrical stimulus trains (0.1, 0.3, and 1 Hz; 5-min interstimulus interval) were repeated every 30 min for 3-6 h after each treatment. The total number of evoked spikes (summed across all 20 stimuli) to 0.1 Hz was enhanced 3 h after NP, mainly in the after-discharge (AD) period (latency > 400 ms). Total responses to 0.3 and 1.0 Hz were also enhanced at > or = 60 min after NP in both the C-fiber (100- to 400-ms latency) and AD periods, whereas the absolute windup (C-fiber + AD - 20 times the initial response) increased at > or = 90 min after treatment. In saline controls, windup was not enhanced at any time after treatment for any stimulus frequency, although there was a trend toward enhancement at 0.3 Hz. These results are consistent with NP-induced central sensitization. Mechanical responses were not significantly enhanced after saline or NP treatment. We speculate that inflammatory agents released from (or recruited by) NP affect the dorsal root
Kumamoto, Jun-Ichi; Nakatani, Masashi; Tsutsumi, Moe; Goto, Makiko; Denda, Sumiko; Takei, Kentaro; Denda, Mitsuhiro
The density of peripheral nerve fibres is increased in atopic dermatitis. Moreover, reduction in the fibres in a mouse model of atopic dermatitis reduces scratching behaviour. Thus, regulation of nerve fibre extension could be an effective strategy to reduce itching in pruritus dermatosis. In this study, we established a new coculture system of keratinocytes and dorsal-root-ganglion-derived cells using an apparatus, AXIS(™) , which consists of two different channels connected via a set of microgrooves, through which signalling molecules and axons, but not living cells, can pass. When we seeded keratinocytes in one chamber, extension of nerve fibres was observed from dorsal root ganglion cells seeded in the other chamber. Addition of anti-BDNF antibody in the keratinocyte-seeded chamber significantly reduced the extension. Application of Semaphorin 3A also reduced the extension by approximately 50%. We suggest that this coculture system may be useful for screening of anti-itching drugs.
Wen, Xianjie; Li, Xiaohong; Liang, Hua; Yang, Chenxiang; Zhong, Jiying; Wang, Hanbing; Liu, Hongzhen
CaMKIIγ in dorsal root ganglion neurons is closely related to the neuropathic pain, neuron injury induced by local anesthetics. To get great insight into the function of CaMKIIγ in dorsal root ganglion neurons, we need one cell model to specially inhibit the CaMKIIγ mRNA expression. The present study was aimed to establish one cell model to specially inhibit the CaMKIIγ mRNA expression. We designed the CaMKIIγ shRNA sequence and connected with pYr-1.1 plasmid. The ligation product of the CaMKIIγshRNA and pYr-1.1 plasmid was recombined with pAd/PL-DEST vector into pAD-CaMKIIγ-shRNA. adenovirus vector. pAD-CaMKIIγ-shRNA. adenovirus vector infected the dorsal root ganglion neuron to inhibit the CaMKIIγ mRNA expression in vitro. The pAD-CaMKIIγ-shRNA adenovirus vector was verified to be correct by the digestion, sequence. And pAD-CaMKIIγ-shRNA. adenovirus vector can infect the DRG cells to inhibit the CaMKIIγ mRNA or protein expression by the real-time polymerase chain reaction (PCR) or western blotting. Those results showed that we successfully constructed one adenovirus vector that can infect the dorsal root ganglion neuron to inhibit the CaMKIIγ mRNA and protein expression. That will supply with one cell model for the CaMKIIγ function study.
Wu, Shaogen; Marie Lutz, Brianna; Miao, Xuerong; Liang, Lingli; Mo, Kai; Chang, Yun-Juan; Du, Peicheng; Soteropoulos, Patricia; Tian, Bin; Kaufman, Andrew G.; Bekker, Alex; Hu, Yali
Background Peripheral nerve injury leads to changes in gene expression in primary sensory neurons of the injured dorsal root ganglia. These changes are believed to be involved in neuropathic pain genesis. Previously, these changes have been identified using gene microarrays or next generation RNA sequencing with poly-A tail selection, but these approaches cannot provide a more thorough analysis of gene expression alterations after nerve injury. Methods The present study chose to eliminate mRNA poly-A tail selection and perform strand-specific next generation RNA sequencing to analyze whole transcriptomes in the injured dorsal root ganglia following spinal nerve ligation. Quantitative real-time reverse transcriptase polymerase chain reaction assay was carried out to verify the changes of some differentially expressed RNAs in the injured dorsal root ganglia after spinal nerve ligation. Results Our results showed that more than 50 million (M) paired mapped sequences with strand information were yielded in each group (51.87 M–56.12 M in sham vs. 51.08 M–57.99 M in spinal nerve ligation). Six days after spinal nerve ligation, expression levels of 11,163 out of a total of 27,463 identified genes in the injured dorsal root ganglia significantly changed, of which 52.14% were upregulated and 47.86% downregulated. The largest transcriptional changes were observed in protein-coding genes (91.5%) followed by noncoding RNAs. Within 944 differentially expressed noncoding RNAs, the most significant changes were seen in long interspersed noncoding RNAs followed by antisense RNAs, processed transcripts, and pseudogenes. We observed a notable proportion of reads aligning to intronic regions in both groups (44.0% in sham vs. 49.6% in spinal nerve ligation). Using quantitative real-time polymerase chain reaction, we confirmed consistent differential expression of selected genes including Kcna2, Oprm1 as well as lncRNAs Gm21781 and 4732491K20Rik following spinal nerve
Benitez, Sergio; Seltzer, Alicia; Acosta, Cristian
AT2 receptor (AT2R) plays a functional role in foetal development. Its expression declines in most tissues soon after birth but stays high in sensory areas of the adult nervous system. In the dorsal root ganglia (DRG) the expression pattern of AT2R during development and the identity of the subpopulation expressing it remain unknown. Using a combination of semi-quantitative PCR, western blotting and immunohistochemistry we examined the expression of AT2R at mRNA and protein levels in rat DRGs from embryonic day 15 (E15) until postnatal day 30 (PN30). We found that both AT2R mRNA and protein levels exhibited only minor (statistically non-significant) fluctuations from E15 to PN30. Detailed quantitative analysis of ABC/DAB AT2R staining showed a) that the receptor was present in most neurons at E15 and E18 and b) that postnatally it was predominantly expressed by small DRG neurons. Given that small neurons are putative C-nociceptors and the proposed role of AT2R in neuropathic pain, we next examined whether these AT2R-positive neurons co-localized with Ret and trkA embryonically and with IB4-binding postnatally. Most AT2R-positive neurons expressed trkA embryonically and bound IB4 postnatally. We found strong positive statistically highly significant correlations between AT2R cytoplasmic%intensities and trkA at E15/E18 and with Ret only at E18. Cytoplasmic AT2R also strongly and positively correlated with IB4-binding at PN3, 15 and 30. Our demonstration that a subpopulation of C-nociceptor-like neurons expresses AT2R during development supports a role for this receptor in neuropathic pain.
Puopolo, Michelino; Martin-Eauclaire, Marie-France; Bean, Bruce P.
The voltage-activated sodium (Nav) channel Nav1.9 is expressed in dorsal root ganglion (DRG) neurons where it is believed to play an important role in nociception. Progress in revealing the functional properties and pharmacological sensitivities of this non-canonical Nav channel has been slow because attempts to express this channel in a heterologous expression system have been unsuccessful. Here, we use a protein engineering approach to dissect the contributions of the four Nav1.9 voltage sensors to channel function and pharmacology. We define individual S3b–S4 paddle motifs within each voltage sensor, and show that they can sense changes in membrane voltage and drive voltage sensor activation when transplanted into voltage-activated potassium channels. We also find that the paddle motifs in Nav1.9 are targeted by animal toxins, and that these toxins alter Nav1.9-mediated currents in DRG neurons. Our results demonstrate that slowly activating and inactivating Nav1.9 channels have functional and pharmacological properties in common with canonical Nav channels, but also show distinctive pharmacological sensitivities that can potentially be exploited for developing novel treatments for pain. PMID:21670206
Clark, SP; Bollag, WB; Westlund, KN; Ma, F; Falls, G; Xie, D; Johnson, M; Isales, CM; Bhattacharyya, MH
A commercial resin-based pine oil derived from Pinus palustris and Pinus elliottii was the major focus of this investigation. Extracts of pine resins, needles and bark are folk medicines commonly used to treat skin ailments, including burns. The American Burn Association estimates that 500,000 people with burn injuries receive medical treatment each year; one-half of US burn victims are children, most with scald burns. This systematic study was initiated as follow-up to personal anecdotal evidence acquired over more than 10 years by MH Bhattacharyya regarding pine oil’s efficacy for treating burns. The results demonstrate that pine oil counteracted dermal inflammation in both a mouse ear model of contact irritant-induced dermal inflammation and a 2nd degree scald burn to the mouse paw. Furthermore, pine oil significantly counteracted the tactile allodynia and soft tissue injury caused by the scald burn. In mouse dorsal root ganglion (DRG) neuronal cultures, pine oil added to the medium blocked ATP-activated, but not capsaicin-activated, pain pathways, demonstrating specificity. These results together support the hypothesis that a pine-oil-based treatment can be developed to provide effective in-home care for 2nd degree burns. PMID:23595692
Tang, He-Bin; Inoue, Atsuko; Iwasa, Mikiko; Hide, Izumi; Nakata, Yoshihiro
To clarify the molecular mechanism of substance P (SP) release from dorsal root ganglion (DRG) neurons, we investigated the involvement of several intracellular effectors in the regulation of SP release evoked by capsaicin, potassium or/and bradykinin. Bradykinin-evoked SP release from cultured adult rat DRG neurons was attenuated by either the mitogen-activated protein kinase kinase (MEK) inhibitor (U0126) or cycloheximide. As the long-term exposure of DRG neurons to bradykinin (3 h) resulted in extracellular signal-regulated kinase (ERK) phosphorylation at an early stage and thereafter induced cyclooxygenase-2 (COX-2) protein expression, which both contribute to the SP release triggered by bradykinin B2 receptor. The long-term exposure of DRG neurons to bradykinin enhanced the SP release by capsaicin, but attenuated that by potassium. Interestingly, the inositol 1,4,5-triphosphate (IP3)-induced calcium release blocker [2-aminoethyl diphenylborinate (2-APB)] not only inhibited the potassium-evoked SP release, but also completely abolished the enhancement of capsaicin-induced SP release by bradykinin from cultured DRG neurons. Together, these findings suggest that the molecular mechanisms of SP release by bradykinin involve the activation of MEK, and also require the de novo protein synthesis of COX-2 in DRG neurons. The IP3-dependent calcium release could be involved in the processes of the regulation by bradykinin of capsaicin-triggered SP release.
Naziroğlu, Mustafa; Uğuz, Abdülhadi Cihangir; Ismailoğlu, Özgür; Çiğ, Bilal; Özgül, Cemil; Borcak, Muhammed
We sought to determine the contribution of oxidative stress-dependent activation of TRPM2 and L-type voltage-gated Ca(2+) channels (VGCC) in dorsal root ganglion (DRG) neurons of rats after spinal cord injury (SCI). The rats were divided into 4 groups: control; sham control; SCI; and SCI+nimodipine groups. The neurons of the SCI groups were also incubated with non-specific TRPM2 channel blockers, 2-aminoethoxydiphenylborate (2-APB) and N-(p-amylcinnamoyl)anthranilic acid (ACA), before H2 O2 stimulation. The [Ca(2+) ]i concentrations were higher in the SCI group than in the control groups, although their concentrations were decreased by nimodipine and 2-APB. The H2 O2 -induced TRPM2 current densities in patch-clamp experiments were decreased by ACA and 2-APB incubation. In the nimodipine group, the TRPM2 channels of neurons were not activated by H2 O2 or cumene hydroperoxide. Increased Ca(2+) influx and currents in DRG neurons after spinal injury indicated TRPM2 and voltage-gated Ca(2+) channel activation. Copyright © 2013 Wiley Periodicals, Inc.
Fan, Chenglei; Chu, Xiao; Wang, Lin; Shi, Hao; Li, Tieshan
Arthritis pain affects people's long-term health, and recent studies have demonstrated that transient receptor potential vanilloid type 1 (TRPV1) plays a crucial role in arthritis pain. In addition, Pre-clinical evidence indicated that botulinum toxin type A (BoNT/A) has antinociceptive effect. The present study investigated the causality between the antinociceptive effects of BoNT/A and the expression of TRPV1 in dorsal root ganglion (DRG) in rats with adjuvant-arthritis pain. The results showed that BoNT/A significantly reduced adjuvant-arthritis nociceptive behaviors in a dose-dependent manner. Furthermore, the BoNT/A cleaved synaptosomal-associated protein of 25 kDa (cl-SNAP-25) was detected in the DRG using immunofluorescence after intra-articular administration. Although BoNT/A significantly reduced the protein levels of TRPV1, there were no significant changes in the mRNA levels of TRPV1 between CFA and BoNT/A (1U, 3U, 10U) group after BoNT/A retrograde axonal transport into the DRG with quantitative RT-PCR. This research provides evidence that the antinociceptive mechanism of BoNT/A might be mediated by reduction of TRPV1 expression through inhibition of its plasma membrane trafficking after intra-articular administration. Copyright © 2017 Elsevier Ltd. All rights reserved.
Zhang, Yongjie; Zheng, Yiyan; Zhang, Yi Ping; Shields, Lisa B E; Hu, Xiaoling; Yu, Panpan; Burke, Darlene A; Wang, Heming; Jun, Cai; Byers, Jonathan; Whittemore, Scott R; Shields, Christopher B
Injection of viral vectors into peripheral nerves may transfer specific genes into their dorsal root ganglion (DRG) neurons and motoneurons. However, myelin sheaths of peripheral axons block the entry of viral particles into nerves. We studied whether mild, transient peripheral nerve demyelination prior to intraneural viral vector injection would enhance gene transfer to target DRG neurons and motoneurons. The right sciatic nerve of C57BL/6 mice was focally demyelinated with 1% lysolecithin, and the left sciatic nerve was similarly injected with saline (control). Five days after demyelination, 0.5 microl of Ad5-GFP was injected into both sciatic nerves at the site of previous injection. The effectiveness of gene transfer was evaluated by counting GFP(+) neurons in the DRGs and ventral horns. After peripheral nerve demyelination, there was a fivefold increase in the number of infected DRG neurons and almost a 15-fold increase in the number of infected motoneurons compared with the control, nondemyelinated side. Focal demyelination reduced the myelin sheath barrier, allowing greater virus-axon contact. Increased CXADR expression on the demyelinated axons facilitated axoplasmic viral entry. No animals sustained any prolonged neurological deficits. Increased gene delivery into DRG neurons and motoneurons may provide effective treatment for amyotrophic lateral sclerosis, pain, and spinal cord injury.
Zhang, Yi; Yue, Jing; Ai, Midan; Ji, Zhigang; Liu, Zhiguo; Cao, Xuehong; Li, Li
We used optogenetic techniques in spinal cord and dorsal root ganglion (DRG) neuron studies. This study investigated changes in channelrhodopsin-2 (ChR2) expression in the spinal cord and DRG neurons using optogenetic techniques. The results show the possibility of using optogenetics to treat neuropathic pain. Previous studies have shown that activated ChR2 induces an increase in DRG neuron action potential. Western blot analysis was used to measure ChR2 protein levels in the spinal cord and DRG neurons or rats intrathecally injected with ChR2 lentivirus. Electrophysiology recording was used to detect differences in action potential levels in the spinal cord and calcium channel currents in the DRG neurons. Our studies showed that ChR2 expression increased the action potential in the spinal cord and increased calcium channel currents in DRG neurons. We successfully expressed the ChR2 protein in the spinal cord and DRG neurons. We also found that ChR2 increased the action potential in the spinal cord and activated the calcium channel in DRG neurons. These findings support the research possibilities of using optogenetic studies to improve treatment for neuropathic pain. N/A.
Wendland, Jens R; Schmidt, Klaus H; Koltzenburg, Martin; Petersen, Marlen
The neuropeptide galanin is known to have an antinociceptive effect under neuropathic conditions. After axotomy, galanin is upregulated in sensory neurons, presumably in the capsaicin-sensitive ones. Here, the sensitivity to capsaicin and the expression of galanin were simultaneously examined by double-staining in individual, dissociated rat dorsal root ganglion neurons (1) after axotomy of the sciatic nerve for up to 14 days and (2) in culture for up to 4 days without prior nerve injury. Ten days after axotomy, the proportion of capsaicin-sensitive neurons had decreased by 36 percentage points (from 63% to 27%), whereas the proportion of galaninergic neurons had increased by 33 percentage points (from 3% to 36%). These changes were also observed in neurons kept in culture, where the regulation was attenuated by the addition of nerve growth factor (NGF) or glial cell line-derived neurotrophic factor (GDNF) to the medium. After axotomy, galaninergic neurons had a soma size-distribution profile similar to the capsaicin-sensitive neurons, but there was no colocalization of capsaicin sensitivity and galanin expression in individual neurons. In culture, some neurons showed colocalization after 30 h and 48 h, but not after 6 h or 96 h. We conclude that the upregulation of galanin in an individual neuron is preceded by downregulation of its capsaicin sensitivity both in NGF-dependent peptidergic and in GDNF-dependent non-peptidergic neurons, indicating a change in phenotype.
Liu, Ting-Ting; Qu, Zu-Wei; Qiu, Chun-Yu; Qiu, Fang; Ren, Cuixia; Gan, Xiong; Peng, Fang; Hu, Wang-Ping
Levo-tetrahydropalmatine (l-THP), a main bioactive Chinese herbal constituent from the genera Stephania and Corydalis, has been in use in clinical practice for years in China as a traditional analgesic agent. However, the mechanism underlying the analgesic action of l-THP is poorly understood. This study shows that l-THP can exert an inhibitory effect on the functional activity of native acid-sensing ion channels (ASICs), which are believed to mediate pain caused by extracellular acidification. l-THP dose dependently decreased the amplitude of proton-gated currents mediated by ASICs in rat dorsal root ganglion (DRG) neurons. l-THP shifted the proton concentration-response curve downward, with a decrease of 40.93% ± 8.45% in the maximum current response to protons, with no significant change in the pH0.5 value. Moreover, l-THP can alter the membrane excitability of rat DRG neurons to acid stimuli. It significantly decreased the number of action potentials and the amplitude of the depolarization induced by an extracellular pH drop. Finally, peripherally administered l-THP inhibited the nociceptive response to intraplantar injection of acetic acid in rats. These results indicate that l-THP can inhibit the functional activity of ASICs in dissociated primary sensory neurons and relieve acidosis-evoked pain in vivo, which for the first time provides a novel peripheral mechanism underlying the analgesic action of l-THP.
Kolarcik, Christi L.; Catt, Kasey; Rost, Erika; Albrecht, Ingrid N.; Bourbeau, Dennis; Du, Zhanhong; Kozai, Takashi D.Y.; Luo, Xiliang; Weber, Douglas J.; Cui, X. Tracy
Objective The dorsal root ganglion (DRG) is an attractive target for implanting neural electrode arrays that restore sensory function or provide therapy via stimulation. However, penetrating microelectrodes designed for these applications are small and deliver low currents. For long-term performance of microstimulation devices, novel coating materials are needed in part to decrease impedance values at the electrode-tissue interface and to increase charge storage capacity. Approach Conductive polymer poly(3,4-ethylenedioxythiophene) (PEDOT) and multiwall carbon nanotubes (CNTs) were coated on the electrode surface and doped with the anti-inflammatory drug, dexamethasone. Electrode characteristics and the tissue reaction around neural electrodes as the result of stimulation, coating and drug release were characterized. Hematoxylin and eosin staining along with antibodies recognizing Iba1 (microglia/macrophages), NF200 (neuronal axons), NeuN (neurons), vimentin (fibroblasts), caspase-3 (cell death) and L1 (neural cell adhesion molecule) were used. Quantitative image analyses were performed using MATLAB. Main Results Our results indicate that coated microelectrodes have lower in vitro and in vivo impedance values. Significantly less neuronal death/damage was observed with coated electrodes as compared to non-coated controls. The inflammatory response with the PEDOT/CNT-coated electrodes was also reduced. Significance This study is the first to report on the utility of these coatings in stimulation applications. Our results indicate PEDOT/CNT coatings may be valuable additions to implantable electrodes used as therapeutic modalities. PMID:25485675
Wang, Siran; Ghezzi, Chiara E; White, James D; Kaplan, David L
Corneal tissue displays the highest peripheral nerve density in the human body. Engineering of biomaterials to promote interactions between neurons and corneal tissue could provide tissue models for nerve/cornea development, platforms for drug screening, as well as innovative opportunities to regenerate cornea tissue. The focus of this study was to develop a coculture system for differentiated human corneal stromal stem cells (dhCSSCs) and dorsal root ganglion neurons (DRG) to mimic the human cornea tissue interactions. Axon extension, connectivity, and neuron cell viability were studied. DRG neurons developed longer axons when cocultured with dhCSSCs in comparison to neuron cultures alone. To assess the mechanism involved in the coculture response, nerve growth factors (NGF) secreted by dhCSSCs including NGF, brain-derived neurotrophic factor (BDNF), glial cell-derived neurotrophic factor (GDNF), and neurotrophin-3 were characterized with greater focus on BDNF secretion. DhCSSCs also secreted collagen type I, an extracellular matrix molecule favorable for neuronal outgrowth. This coculture system provides a slowly degrading silk matrix to study neuronal responses in concert with hCSSCs related to innervation of corneal tissue with utility toward human corneal nerve regeneration and associated diseases. © 2015 Wiley Periodicals, Inc.
Wei, Jinrong; Li, Meng; Wang, Dieyu; Zhu, Hongyan; Kong, Xiangpeng; Wang, Shusheng; Zhou, You-Lang; Ju, Zhong; Jiang, Guo-Qin
Background Cancer-induced pain (CIP) is one of the most severe types of chronic pain with which clinical treatment remains challenging and the involved mechanisms are largely unknown. Suppressor of cytokine signaling 3 (SOCS3) is an important intracellular protein and provides a classical negative feedback loop, thus involving in a wide variety of processes including inflammation and nociception. However, the role of SOCS3 pathway in CIP is poorly understood. The present study was designed to investigate the role of SOCS3 in dorsal root ganglion (DRG) in the development of CIP. Method CIP was established by injection of Walker 256 mammary gland tumor cells into the rat tibia canal. Whole-cell patch clamping and Western blotting were performed. Results Following the development of bone cancer, SOCS3 expression was significantly downregulated in rat DRGs at L2–L5 segments. Overexpression of SOCS3, using lentiviral-mediated production of SOCS3 at spinal cord level, drastically attenuated mechanical allodynia and body weight-bearing difference, but not thermal hyperalgesia in bone cancer rats. In addition, overexpression of SOCS3 reversed the hyperexcitability of DRG neurons innervating the tibia, and reduced abnormal expression of toll-like receptors 4 in the DRGs. Conclusions These results suggest that SOCS3 might be a key molecular involved in the development of complicated cancer pain and that overexpression of SOCS3 might be an important strategy for treatment for mechanical allodynia associated with bone cancer. PMID:28326931
Li, Hao; Zhang, Weiwei; Liu, Guixiang; Li, Jianmin; Liu, Huaxiang; Li, Zhenzhong
The neurotrophic factor-like activity of monosialoganglioside (GM1) has been shown to activate tyrosine kinase receptors (Trk). Targets of neuronal innervation play a vital role in regulating the survival and differentiation of innervating neurotrophin-responsive neurons. Both GM1 and target skeletal muscle (SKM) cells are essential for the maintenance of the function of neurons. However, much less is known about the effects of GM1 or/and target SKM cells on the expression of Trk receptors in dorsal root ganglion (DRG) neurons. Here we have tested what extent to the expression of TrkA, TrkB, and TrkC receptors in primary cultured of DRG neurons in absence or presence of GM1 or/and SKM cells. In this experiment, we found that: (1) GM1 promoted expression of TrkA and TrkB but not TrkC in primary cultured DRG neurons; (2) target SKM cells promoted expression of TrkC but not TrkA and TrkB in neuromuscular cocultures without GM1 treatment; and (3) GM1 and target SKM cells had additional effects on expression of these three Trk receptors. The results of the present study offered new clues for a better understanding of the association of GM1 and target SKM on the expression of Trk receptors.
Wang, Jie; Wang, Xiao-Wei; Zhang, Yang; Yin, Cui-Ping; Yue, Shou-Wei
Chronic compression of the dorsal root ganglion (DRG) (CCD) in rats is a typical model of neuropathic pain. TRPV4 contributed to mechanical allodynia induced by the CCD model. Our previous study demonstrated that TRPV4 enhances neuropathic hyperalgesia through a NO-cGMP-PKG cascade. However, the underlying mechanism(s) is still largely unknown. Therefore, the aim of the present study was to test whether TRPV4-mediated Ca(2+) influx is involved in the TRPV4-NO pathway. Regulation of intracellular calcium concentration by intrathecal injection of TRPV4-targeted siRNA significantly decreased the behavioural hyperalgesia, NF-κB activity, and NO content in CCD rats. Intraperitoneal (i.p.) injection of mibefradil significantly induced dose-dependent increases in the paw withdrawal latency (PWL) and mechanical withdrawal thresholds (MWT), as well as decreases in NF-κB activity and NO content in DRG of CCD rats. Moreover, pre-treatment with 4α-PDD attenuated the suppressive effects of mibefradil on CCD-induced neuropathic hyperalgesia, NF-κB activity, and NO production. The data showed that TRPV4-mediated Ca(2+) influx might be engaged in the TRPV4-NO pathway in neuropathic hyperalgesia in the CCD model. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.
Zhang, Hua; Verkman, A. S.
Aquaporin-1 (AQP1) water channels are expressed in the plasma membrane of dorsal root ganglion (DRG) neurons. We found reduced osmotic water permeability in freshly isolated DRG neurons from AQP1−/− versus AQP1+/+ mice. Behavioral studies showed greatly reduced thermal inflammatory pain perception in AQP1−/− mice evoked by bradykinin, prostaglandin E2, and capsaicin as well as reduced cold pain perception. Patch clamp of freshly isolated DRG neurons showed reduced action potential firing in response to current injections. Single action potentials after pulse current injections showed reduced maximum inward current, suggesting impaired Nav1.8 Na+ function. Whole-cell Nav1.8 Na+ currents in Nav1.8-expressing ND7-23 cells showed slowed frequency-dependent inactivation after AQP1 transfection. Immunoprecipitation studies showed AQP1- Nav1.8 Na+ interaction, which was verified in live cells by single-particle tracking of quantum dot-labeled AQP1. Our results implicate the involvement of AQP1 in DRG neurons for the perception of inflammatory thermal pain and cold pain, whose molecular basis is accounted for, in part, by reduced Nav1.8-dependent membrane Na+ current. AQP1 is, thus, a novel target for pain management. PMID:20018876
Yuan, Huijun; Lan, Tonghan; Lin, Jiarui
Nano-Selenium, a novel Nano technology production, was demonstrated to be useful in medical and scientific researches. Here, we investigated the effects of Nano-Selenium on tetrodotoxin-sensitive (TTX-S) voltage-dependent Na+channels in isolated rat dorsal root ganglion neurons, using whole-cell patch-clamp method. Nano-Selenium irreversibly decreased TTX-S Na+current (I
Tonra, J R; Mendell, L M
Experiments were undertaken to examine anatomical correlates of physiological effects of rabbit sera raised against nerve growth factor (anti-NGF) on nociceptive afferents. This antiserum has been shown to deplete the population of A-delta high threshold mechanoreceptors and to reduce neurogenic vasodilatation. Because numerous studies implicate calcitonin gene related peptide (CGRP)-containing sensory neurons in these effects, immunocytochemical and anatomical techniques were used to examine the normal development of CGRP-immunoreactive (-IR) neurons in the dorsal root ganglion (DRG) of rats from 13 days to 19 weeks of age, and to compare this to the development in rats treated neonatally (postnatal days 2-14) with anti-NGF. In controls the rate of increase in the mean diameter of CGRP-IR cells was substantially greater between 13 days and 5 weeks of age than it was between 5 weeks and 19 weeks, in contrast to CGRP-negative neurons whose rate of growth remained relatively constant. Anti-NGF had no significant effect on growth rate, but rats treated with anti-NGF exhibited a reduced proportion of CGRP-IR neurons at 5 weeks. This deficit was reversed by 19 weeks unlike the physiological changes. These results indicate independent regulation of CGRP expression and nociceptor physiology by NGF.
Badea, Adina; McCracken, Joselle M; Tillmaand, Emily G; Kandel, Mikhail E; Oraham, Aaron W; Mevis, Molly B; Rubakhin, Stanislav S; Popescu, Gabriel; Sweedler, Jonathan V; Nuzzo, Ralph G
Understanding and controlling the interactions occurring between cells and engineered materials are central challenges toward progress in the development of biomedical devices. In this work, we describe materials for direct ink writing (DIW), an extrusion-based type of 3D printing, that embed a custom synthetic protein (RGD-PDL) within the microfilaments of 3D-hydrogel scaffolds to modify these interactions and differentially direct tissue-level organization of complex cell populations in vitro. The RGD-PDL is synthesized by modifying poly-d-lysine (PDL) to varying extents with peptides containing the integrin-binding motif Arg-Gly-Asp (RGD). Compositional gradients of the RGD-PDL presented by both patterned and thin-film poly(2-hydroxyethyl) methacrylate (pHEMA) substrates allow the patterning of cell-growth compliance in a grayscale form. The surface chemistry-dependent guidance of cell growth on the RGD-PDL-modified pHEMA materials is demonstrated using a model NIH-3T3 fibroblast cell line. The formation of a more complex cellular system-organotypic primary murine dorsal root ganglion (DRG)-in culture is also achieved on these scaffolds, where distinctive forms of cell growth and migration guidance are seen depending on their RGD-PDL content and topography. This experimental platform for the study of physicochemical factors on the formation and the reorganization of organotypic cultures offers useful capabilities for studies in tissue engineering, regenerative medicine, and diagnostics.
Lakritz, Jessica R.; Bodair, Ayman; Shah, Neal; O'Donnell, Ryan; Polydefkis, Michael J.; Miller, Andrew D.; Burdo, Tricia H.
HIV-associated sensory neuropathy remains the most common neurological complication of HIV infection and is characterized by dorsal root ganglion (DRG) inflammation and intraepidermal nerve fiber density (IENFD) loss. Chronic peripheral immune cell activation and accumulation may cause damage to the DRG, but has not been fully investigated yet. By using an SIV-infected, CD8-lymphocyte–depleted rhesus macaque model, we defined immune cells surrounding DRG neurons and their role in DRG pathology, measured cell traffic from the bone marrow to the DRGs using 5-bromo-2-deoxyuridine (BrdU) pulse, and serially measured IENFD. We found an increase in CD68+ and CD163+ macrophages in DRGs of SIV-infected animals. MAC387+ recently recruited monocytes/macrophages were increased, along with BrdU+ cells, in the DRGs of SIV-infected macaques. We demonstrated that 78.1% of all BrdU+ cells in DRGs were also MAC387+. The number of BrdU+ monocytes correlated with severe DRG histopathology, which included neuronophagia, neuronal loss, and Nageotte nodules. These data demonstrate that newly recruited MAC387+BrdU+ macrophages may play a significant role in DRG pathogenesis. IENFD decreased early (day 21), consistent with the development of sensory neuropathy in SIV-infected macaques. Decreased IENFD was associated with elevated BrdU+ cells in the DRG. These data suggest that increased recruitment of macrophages to DRG is associated with severe DRG histopathology and IENFD loss. PMID:25956030
Lee, Moon Chul; Nam, Taick Sang; Jung, Se Jung; Gwak, Young S.; Leem, Joong Woo
Chronic compression of dorsal root ganglion (CCD) results in neuropathic pain. We investigated the role of spinal GABA in CCD-induced pain using rats with unilateral CCD. A stereological analysis revealed that the proportion of GABA-immunoreactive neurons to total neurons at L4/5 laminae I–III on the injured side decreased in the early phase of CCD (post-CCD week 1) and then returned to the sham-control level in the late phase (post-CCD week 18). In the early phase, the rats showed an increase in both mechanical sensitivity of the hind paw and spinal WDR neuronal excitability on the injured side, and such increase was suppressed by spinally applied muscimol (GABA-A agonist, 5 nmol) and baclofen (GABA-B agonist, 25 nmol), indicating the reduced spinal GABAergic inhibition involved. In the late phase, the CCD-induced increase in mechanical sensitivity and neuronal excitability returned to pre-CCD levels, and such recovered responses were enhanced by spinally applied bicuculline (GABA-A antagonist, 15 nmol) and CGP52432 (GABA-B antagonist, 15 nmol), indicating the regained spinal GABAergic inhibition involved. In conclusion, the alteration of spinal GABAergic inhibition following CCD and leading to a gradual reduction over time of CCD-induced mechanical hypersensitivity is most likely due to changes in GABA content in spinal GABA neurons. PMID:26451259
Wu, Ying; Liu, Yongfeng; Hou, Panpan; Yan, Zonghe; Kong, Wenjuan; Liu, Beiying; Li, Xia; Yao, Jing; Zhang, Yuexuan; Qin, Feng; Ding, Jiuping
The transient receptor potential vanilloid receptor 1 (TRPV1) channel is a nonselective cation channel activated by a variety of exogenous and endogenous physical and chemical stimuli, such as temperature (≥42 °C), capsaicin, a pungent compound in hot chili peppers, and allyl isothiocyanate. Large-conductance calcium- and voltage-activated potassium (BK) channels regulate the electric activities and neurotransmitter releases in excitable cells, responding to changes in membrane potentials and elevation of cytosolic calcium ions (Ca(2+)). However, it is unknown whether the TRPV1 channels are coupled with the BK channels. Using patch-clamp recording combined with an infrared laser device, we found that BK channels could be activated at 0 mV by a Ca(2+) influx through TRPV1 channels not the intracellular calcium stores in submilliseconds. The local calcium concentration around BK is estimated over 10 μM. The crosstalk could be affected by 10 mM BAPTA, whereas 5 mM EGTA was ineffectual. Fluorescence and co-immunoprecipitation experiments also showed that BK and TRPV1 were able to form a TRPV1-BK complex. Furthermore, we demonstrated that the TRPV1-BK coupling also occurs in dosal root ganglion (DRG) cells, which plays a critical physiological role in regulating the "pain" signal transduction pathway in the peripheral nervous system.
Yan, Zonghe; Kong, Wenjuan; Liu, Beiying; Li, Xia; Yao, Jing; Zhang, Yuexuan; Qin, Feng; Ding, Jiuping
The transient receptor potential vanilloid receptor 1 (TRPV1) channel is a nonselective cation channel activated by a variety of exogenous and endogenous physical and chemical stimuli, such as temperature (≥42 °C), capsaicin, a pungent compound in hot chili peppers, and allyl isothiocyanate. Large-conductance calcium- and voltage-activated potassium (BK) channels regulate the electric activities and neurotransmitter releases in excitable cells, responding to changes in membrane potentials and elevation of cytosolic calcium ions (Ca2+). However, it is unknown whether the TRPV1 channels are coupled with the BK channels. Using patch-clamp recording combined with an infrared laser device, we found that BK channels could be activated at 0 mV by a Ca2+ influx through TRPV1 channels not the intracellular calcium stores in submilliseconds. The local calcium concentration around BK is estimated over 10 μM. The crosstalk could be affected by 10 mM BAPTA, whereas 5 mM EGTA was ineffectual. Fluorescence and co-immunoprecipitation experiments also showed that BK and TRPV1 were able to form a TRPV1-BK complex. Furthermore, we demonstrated that the TRPV1-BK coupling also occurs in dosal root ganglion (DRG) cells, which plays a critical physiological role in regulating the “pain” signal transduction pathway in the peripheral nervous system. PMID:24147119
Zhang, Yan; Jia, Yan-Yan; Guo, Jin-Lei; Liu, Pei-Qing; Jiang, Jian-Min
The (-)-gallocatechin-3-gallate (GCG) concentration in some tea beverages can account for as much as 50% of the total catechins. It has been shown that catechins have analgesic properties. Voltage-gated sodium channels (Nav) mediate neuronal action potentials. Tetrodotoxin inhibits all Nav isoforms, but Nav1.8 and Nav1.9 are relatively tetrodotoxin-resistant compared to other isoforms and functionally linked to nociception. In this study, the effects of GCG on tetrodotoxin-resistant Na+ currents were investigated in rat primary cultures of dorsal root ganglion neurons via the whole-cell patch-clamp technique. We found that 1 μM GCG reduced the amplitudes of peak current density of tetrodotoxin-resistant Na+ currents significantly. Furthermore, the inhibition was accompanied by a depolarizing shift of the activation voltage and a hyperpolarizing shift of steady-state inactivation voltage. The percentage block of GCG (1 μM) on tetrodotoxin-resistant Na+ current was 45.1% ± 1.1% in 10 min. In addition, GCG did not produce frequency-dependent block of tetrodotoxin-resistant Na+ currents at stimulation frequencies of 1 Hz, 2 Hz and 5 Hz. On the basis of these findings, we propose that GCG may be a potential analgesic agent.
Shi, Haohong; Luo, Xingjing
BACKGROUND 7, 8, 3'-trihydroxyflavone (THF) is a novel pro-neuronal small molecule that acts as a TrkB agonist. In this study, we examined the effect of THF on promoting neuronal growth and protecting anesthetics-induced neurotoxicity in dorsal root ganglion (DRG) neurons in vitro. MATERIAL AND METHODS Neonatal mouse DRG neurons were cultured in vitro and treated with various concentrations of THF. The effect of THF on neuronal growth was investigated by neurite outgrowth assay and Western blot. In addition, the protective effects of THF on bupivacaine-induced neurotoxicity were investigated by apoptosis TUNEL assay, neurite outgrowth assay, and Western blot, respectively. RESULTS THF promoted neurite outgrowth of DRG neurons in dose-dependent manner, with an EC50 concentration of 67.4 nM. Western blot analysis showed THF activated TrkB signaling pathway by inducing TrkB phosphorylation. THF also rescued bupivacaine-induced neurotoxicity by reducing apoptosis and protecting neurite retraction in DRG neurons. Furthermore, the protection of THF in bupivacaine-injured neurotoxicity was directly associated with TrkB phosphorylation in a concentration-dependent manner in DRG neurons. CONCLUSIONS THF has pro-neuronal effect on DRG neurons by promoting neurite growth and protecting against bupivacaine-induced neurotoxicity, likely through TrkB activation.
Jia, Zhanfeng; Ikeda, Ryo; Ling, Jennifer; Gu, Jianguo G
Rapidly adapting mechanically activated channels (RA) are expressed in primary afferent neurons and identified as Piezo2 ion channels. We made whole-cell voltage-clamp recordings from cultured dorsal root ganglion (DRG) neurons to study RA channel regulation. RA currents showed gradual increases in current amplitude (current "run-up") after establishing whole-cell mode when 0.33 mM GTP or 0.33 mM GTPγS was included in the patch pipette internal solution. RA current run-up was also observed in HEK293 cells that heterologously expressed Piezo2 ion channels. No significant RA current run-up was observed in DRG neurons when GTP was omitted from the patch pipette internal solution, when GTP was replaced with 0.33 mM GDP, or when recordings were made under the perforated patch-clamp recording configuration. Our findings revealed a GTP-dependent up-regulation of the function of piezo2 ion channels in DRG neurons.
Rapidly adapting mechanically activated channels (RA) are expressed in primary afferent neurons and identified as Piezo2 ion channels. We made whole-cell voltage-clamp recordings from cultured dorsal root ganglion (DRG) neurons to study RA channel regulation. RA currents showed gradual increases in current amplitude (current “run-up”) after establishing whole-cell mode when 0.33 mM GTP or 0.33 mM GTPγS was included in the patch pipette internal solution. RA current run-up was also observed in HEK293 cells that heterologously expressed Piezo2 ion channels. No significant RA current run-up was observed in DRG neurons when GTP was omitted from the patch pipette internal solution, when GTP was replaced with 0.33 mM GDP, or when recordings were made under the perforated patch-clamp recording configuration. Our findings revealed a GTP-dependent up-regulation of the function of piezo2 ion channels in DRG neurons. PMID:24344923
Mao, Qingxiang; Yuan, Jingjing; Xiong, Ming; Wu, Shaogen; Chen, Liyong; Bekker, Alex; Yang, Tiande
Peripheral nerve injury-caused hyperexcitability and abnormal ectopic discharges in the primary sensory neurons of dorsal root ganglion (DRG) play a key role in neuropathic pain development and maintenance. The two-pore domain background potassium (K2P) channels have been identified as key determinants of the resting membrane potential and neuronal excitability. However, whether K2P channels contribute to neuropathic pain is still elusive. We reported here that K2P1.1, the first identified mammalian K2P channel, was highly expressed in mouse DRG and distributed in small-, medium-, and large-sized DRG neurons. Unilateral lumbar (L) 4 spinal nerve ligation led to a significant and time-dependent reduction of K2P1.1 mRNA and protein in the ipsilateral L4 DRG, but not in the contralateral L4 or ipsilateral L3 DRG. Rescuing this reduction through microinjection of adeno-associated virus-DJ expressing full-length K2P1.1 mRNA into the ipsilateral L4 DRG blocked spinal nerve ligation-induced mechanical, thermal, and cold pain hypersensitivities during the development and maintenance periods. This DRG viral microinjection did not affect acute pain and locomotor function. Our findings suggest that K2P1.1 participates in neuropathic pain development and maintenance and may be a potential target in the management of this disorder. PMID:28326939
Kolarcik, Christi L.; Catt, Kasey; Rost, Erika; Albrecht, Ingrid N.; Bourbeau, Dennis; Du, Zhanhong; Kozai, Takashi D. Y.; Luo, Xiliang; Weber, Douglas J.; Cui, X. Tracy
Objective. The dorsal root ganglion is an attractive target for implanting neural electrode arrays that restore sensory function or provide therapy via stimulation. However, penetrating microelectrodes designed for these applications are small and deliver low currents. For long-term performance of microstimulation devices, novel coating materials are needed in part to decrease impedance values at the electrode-tissue interface and to increase charge storage capacity. Approach. Conductive polymer poly(3,4-ethylenedioxythiophene) (PEDOT) and multi-wall carbon nanotubes (CNTs) were coated on the electrode surface and doped with the anti-inflammatory drug, dexamethasone. Electrode characteristics and the tissue reaction around neural electrodes as a result of stimulation, coating and drug release were characterized. Hematoxylin and eosin staining along with antibodies recognizing Iba1 (microglia/macrophages), NF200 (neuronal axons), NeuN (neurons), vimentin (fibroblasts), caspase-3 (cell death) and L1 (neural cell adhesion molecule) were used. Quantitative image analyses were performed using MATLAB. Main results. Our results indicate that coated microelectrodes have lower in vitro and in vivo impedance values. Significantly less neuronal death/damage was observed with coated electrodes as compared to non-coated controls. The inflammatory response with the PEDOT/CNT-coated electrodes was also reduced. Significance. This study is the first to report on the utility of these coatings in stimulation applications. Our results indicate PEDOT/CNT coatings may be valuable additions to implantable electrodes used as therapeutic modalities.
Wang, Qun; Sun, Yanyuan; Ren, Yingna; Gao, Yandong; Tian, Li; Liu, Yang; Pu, Yanan; Gou, Xingchun; Chen, Yanke; Lu, Yan
Matrix metalloproteinases (MMPs) are widely implicated in inflammation and tissue remodeling associated with various neurodegenerative diseases and play an important role in nociception and allodynia. Extracellular Matrix Metalloproteinase Inducer (EMMPRIN) plays a key regulatory role for MMP activities. However, the role of EMMPRIN in the development of neuropathic pain is not clear. Western blotting, real-time quantitative RT-PCR (qRT-PCR), and immunofluorescence were performed to determine the changes of messenger RNA and protein of EMMPRIN/OX47 and their cellular localization in the rat dorsal root ganglion (DRG) after nerve injury. Paw withdrawal threshold test was examined to evaluate the pain behavior in spinal nerve ligation (SNL) model. The lentivirus containing OX47 shRNA was injected into the DRG one day before SNL. The expression level of both mRNA and protein of OX47 was markedly upregulated in ipsilateral DRG after SNL. OX47 was mainly expressed in the extracellular matrix of DRG. Administration of shRNA targeted against OX47 in vivo remarkably attenuated mechanical allodynia induced by SNL. In conclusion, peripheral nerve injury induced upregulation of OX47 in the extracellular matrix of DRG. RNA interference against OX47 significantly suppressed the expression of OX47 mRNA and the development of mechanical allodynia. The altered expression of OX47 may contribute to the development of neuropathic pain after nerve injury.
Hamer, John F; Purath, Traci A
This article investigates the degree and duration of pain relief from cervicogenic headaches or occipital neuralgia following treatment with radiofrequency ablation of the C2 dorsal root ganglion and/or third occipital nerves. It also addresses the procedure's complication rate and patient's willingness to repeat the procedure if severe symptoms recur. This is a single-center retrospective observational study of 40 patients with refractory cervicogenic headaches and or occipital neuralgia. Patients were all referred by a headache specialty clinic for evaluation for radiofrequency ablation of the C2 dorsal root ganglion and/or third occipital nerves. After treatment, patients were followed for a minimum of 6 months to a year. Patient demographics and the results of radiofrequency ablation were recorded on the same day, after 3-4 days, and at 6 months to 1 year following treatment. Thirty-five percent of patients reported 100% pain relief and 70% reported 80% or greater pain relief. The mean duration of improvement is 22.35 weeks. Complication rate was 12-13%. 92.5% of patients reported they would undergo the procedure again if severe symptoms returned. Radiofrequency ablation of the C2 dorsal root ganglion and/or third occipital nerve can provide many months of greater than 50% pain relief in the vast majority of recipients with an expected length of symptom improvement of 5-6 months. © 2014 American Headache Society.
Cai, Weihua; Cao, Jing; Ren, Xiuhua; Qiao, Liang; Chen, Xuemei; Li, Ming; Zang, Weidong
Abnormal acute pain after burn injury still torments patients severely. In this study, we investigated that one voltage gated sodium channel Nav1.7 plays a vital role in lowering heat pain threshold after burn injury, and the hypothesis that knockdown of Nav1.7 attenuates pain following burn injury. Sixty eight adult male Sprague-Dawley rats were divided into 4 treatment groups: (1) sham, which hind paw was put on the room temperature metal plate for 15 s (2) burn model, which hind paw was put on the 85 °C metal plate for 15 s. (3) Burn injury + lentiviral vector -SCN9AsiRNA-GFP (LV- SCN9AsiRNA-GFP group, n = 18), which receive the DRG microinjection of LV- SCN9AsiRNA-GFP on the zero day. (4) Burn injury + lentiviral vector negative control (LV-NC-GFP group, n = 18), which receive the DRG microinjection of empty lentiviral vector on the zero day. Both mechanical and heat threshold were measured from day 1 to 21. Meanwhile, expression of sodium channels Nav1.7 in injured dorsal root ganglia were measured on post-operative days 7(POD 7). Rats exhibited decreased thresholds on both mechanical allodynia and thermal withdrawl latency, accompanied by increased Nav1.7 and c-fos expression in dorsal root ganglion (DRG). And knockdown of Nav1.7 in L5DRG led to the attenuation of burn injury-induced mechanical allodynia and thermal hyperalgesia in the rats. We provide evidence that shRNA mediated knockdown of Nav1.7 attenuates burn induced pain in rats as well as decreased the activiation of c-fos protein.
Wang, Tao; Hurwitz, Olivia; Shimada, Steven G.; Qu, Lintao; Fu, Kai; Zhang, Pu; Ma, Chao; LaMotte, Robert H.
Radicular pain in humans is usually caused by intraforaminal stenosis and other diseases affecting the spinal nerve, root, or dorsal root ganglion (DRG). Previous studies discovered that a chronic compression of the DRG (CCD) induced mechanical allodynia in rats and mice, with enhanced excitability of DRG neurons. We investigated whether CCD altered the pain-like behavior and also the responses of cutaneous nociceptors with unmyelinated axons (C-fibers) to a normally aversive punctate mechanical stimulus delivered to the hairy skin of the hind limb of the mouse. The incidence of a foot shaking evoked by indentation of the dorsum of foot with an aversive von Frey filament (tip diameter 200 μm, bending force 20 mN) was significantly higher in the foot ipsilateral to the CCD surgery as compared to the contralateral side on post-operative days 2 to 8. Mechanically-evoked action potentials were electrophysiologically recorded from the L3 DRG, in vivo, from cell bodies visually identified as expressing a transgenically labeled fluorescent marker (neurons expressing either the receptor MrgprA3 or MrgprD). After CCD, 26.7% of MrgprA3+ and 32.1% MrgprD+ neurons exhibited spontaneous activity (SA), while none of the unoperated control neurons had SA. MrgprA3+ and MrgprD+ neurons in the compressed DRG exhibited, in comparison with neurons from unoperated control mice, an increased response to the punctate mechanical stimuli for each force applied (6, 20, 40, and 80 mN). We conclude that CCD produced both a behavioral hyperalgesia and an enhanced response of cutaneous C-nociceptors to aversive punctate mechanical stimuli. PMID:26356638
Wong, J.; Oblinger, M.M. )
The expression of major cytoskeletal protein mRNAs was studied in adult rat dorsal root ganglion (DRG) neurons after crushing either their central or peripheral branch axons. mRNA levels in DRG neurons were examined by quantitative in situ hybridization with radiolabeled cDNA probes specific for the low-molecular-weight neurofilament protein (NF-L) and beta-tubulin. The large-sized (greater than 1000 microns 2) neurons which give rise to myelinated axons in lumbar ganglia (L4 and L5) were studied 1 d through 8 weeks after either dorsal root or sciatic nerve crush. NF-L and beta-tubulin mRNA levels in axotomized DRG neurons were compared to those in contralateral control DRG neurons, as well as to those in normal (completely untreated) DRG cells. In the case of NF-L mRNA, changes were observed after central as well as peripheral branch axotomy and the time course and magnitude of changes were similar after both types of axotomy. NF-L mRNA levels initially decreased (first 2 weeks after crush) and then began to return towards control levels at longer survival times. Similar, but less pronounced, changes in NF-L mRNA levels also occurred in contralateral DRG neurons (which were uninjured); the changes in contralateral neurons were not simply a result of surgical stress since no changes in NF-L mRNA levels were observed in sham-operated DRG neurons. In the case of tubulin mRNA, changes were observed after central as well as peripheral branch axotomy by in situ hybridization, but the time course and magnitude of changes were different after each type of axotomy.
Fischer, Gregory; Kostic, Sandra; Nakai, Hiroyuki; Park, Frank; Sapunar, Damir; Yu, Hongwei; Hogan, Quinn
Direct injection of agents into the dorsal root ganglia (DRGs) offers the opportunity to manipulate sensory neuron function at a segmental level to explore pathophysiology of painful conditions. However, there is no described method that has been validated in detail for such injections in adult rats. We have found that 2 (µl of dye injected through a pulled glass pipette directly into the distal DRG, exposed by a minimal foraminotomy, produces complete filling of the DRG with limited extension into the spinal roots. Injection into the spinal nerve required 3 µl to achieve comparable DRG filling, produced preferential spread into the ventral root, and was accompanied by substantial leakage of injected solution from the injection site. Injections into the sciatic nerve of volumes up to 10 (µl did not reach the DRG. Transient hypersensitivity to mechanical stimulation at threshold (von Frey) and noxious levels (pin) developed after 2 µl saline injection directly into the DRG that was in part attributable to the surgical exposure procedure alone. Only minimal astrocyte activation in the spinal dorsal horn was evident after DRG saline injections. Injection of adeno-associated virus (AAV) vector conveying green fluorescent protein (GFP) transgene resulted in expression as soon as 1 day after injection into the DRG, including fibers in the spinal dorsal horn and columns. AAV injection into the DRG produced additional thermal hypersensitivity and withdrawal from the stroke of a brush and compromised motor performance. These findings demonstrate a method for selective injection of agents into single DRGs for anatomically restricted actions. PMID:21540055
Chen, Yi-Jen; Wang, Yan-Hsiung; Wang, Chau-Zen; Ho, Mei-Ling; Kuo, Po-Lin; Huang, Mao-Hsiung; Chen, Chia-Hsin
Dorsal root ganglia (DRG) are vulnerable to physical injury of the intervertebral foramen, and chronic compression of the DRG (CCD) an result in nerve root damage with persistent morbidity. The purpose of this study was to evaluate the effects of low level laser therapy (LLLT) on the DRG in a CCD model and to determine the mechanisms underlying these effects. CCD rats had L-shaped stainless-steel rods inserted into the fourth and fifth lumbar intervertebral foramen, and the rats were then subjected to 0 or 8 J/cm2 LLLT for 8 consecutive days following CCD surgery. Pain and heat stimuli were applied to test for hyperalgesia following CCD. The levels of TNF-α, IL-1β and growth-associated protein-43 (GAP-43) messenger RNA (mRNA) expression were measured via real-time PCR, and protein expression levels were analyzed through immunohistochemical analyses. Our data indicate that LLLT significantly decreased the tolerable sensitivity to pain and heat stimuli in the CCD groups. The expression levels of the pro-inflammatory cytokines TNF-α and IL-1β were increased following CCD, and we found that these increases could be reduced by the application of LLLT. Furthermore, the expression of GAP-43 was enhanced by LLLT. In conclusion, LLLT was able to enhance neural regeneration in rats following CCD and improve rat ambulatory behavior. The therapeutic effects of LLLT on the DRG during CCD may be exerted through suppression of the inflammatory response and induction of neuronal repair genes. These results suggest potential clinical applications for LLLT in the treatment of compression-induced neuronal disorders.
Wang, Dong; Pan, Hao; Zhu, Hang; Zhu, Li; He, Yong-Jiang; Wang, Jian; Jia, Gao-Yong
The nucleus pulposus (NP) is an avascular, hydrated tissue that permits the intervertebral disc to resist compressive loads to the spine. To determine the mechanisms by which intervertebral disc degeneration is caused by the nucleus pulposus, the expression and regulation of nuclear factor (NF)‑κB and acid sensing ion channel 3 (ASIC3) were examined. For the intervertebral disc degeneration model, NP was harvested from the tail of rats and applied to the L5 dorsal root ganglion (DRG). The mechanical pain withdrawal threshold (PWT) in NP model rats was assessed. Reverse transcription‑quantitative polymerase chain reaction and western blotting were used to examine NF‑κB and ASIC3 expression levels in DRG. Finally, the effect of the NF‑κB inhibitor pyrrolidine dithiocarbamate (PDTC) and the ASIC3 signaling pathway blocker amiloride were examined. Rats exposed to NP exhibited decreased PWT for 12 days, and NF‑κB and ASIC3 was upregulated in DRG induced by NP 14 days after surgery. After administration of amiloride and PDTC to DRG affected by NP, the levels of nitric oxide (NO), tumor necrosis factor‑α (TNF‑α), interleukin‑6 (IL‑6), NF‑κB and ASIC3 were downregulated, and the levels of aquaporin (AQP) 1 and AQP3 were significantly increased for 14 days. In conclusion, these results suggested that NF‑κB and ASIC3 may serve an important role in intervertebral disc degeneration caused by NP.
de Simone, Ambra; Hubbard, Rachel; Viñegra de la Torre, Natanael; Velappan, Yazhini; Wilson, Michael; Considine, Michael J; Soppe, Wim J J; Foyer, Christine H
The aim of this study was to characterize redox changes in the nuclei and cytosol occurring during the mitotic cell cycle in the embryonic roots of germinating Arabidopsis seedlings, and to determine how redox cycling was modified in mutants with a decreased capacity for ascorbate synthesis. Using an in vivo reduction-oxidation (redox) reporter (roGFP2), we show that transient oxidation of the cytosol and the nuclei occurred at G1 in the synchronized dividing cells of the Arabidopsis root apical meristem, with reduction at G2 and mitosis. This redox cycle was absent from low ascorbate mutants in which nuclei were significantly more oxidized than controls. The cell cycle-dependent increase in nuclear size was impaired in the ascorbate-deficient mutants, which had fewer cells per unit area in the root proliferation zone. The transcript profile of the dry seeds and size of the imbibed seeds was strongly influenced by low ascorbate but germination, dormancy release and seed aging characteristics were unaffected. These data demonstrate the presence of a redox cycle within the plant cell cycle and that the redox state of the nuclei is an important factor in cell cycle progression. Controlled oxidation is a key feature of the early stages of the plant cell cycle. However, sustained mild oxidation restricts nuclear functions and impairs progression through the cell cycle leading to fewer cells in the root apical meristem. Antioxid. Redox Signal. 00, 000-000.
Moreno-López, Y; Martínez-Lorenzana, G; Condés-Lara, M; Rojas-Piloni, G
Oxytocin (OT) secreted by the hypothalamo-spinal projection exerts antinociceptive effects in the dorsal horn. Electrophysiological evidence indicates that OT could exert these effects by activating OT receptors (OTR) directly on dorsal horn neurons and/or primary nociceptive afferents in the dorsal root ganglia (DRG). However, little is known about the identity of the dorsal horn and DRG neurons that express the OTR. In the dorsal horn, we found that the OTR is expressed principally in neurons cell bodies. However, neither spino-thalamic dorsal horn neurons projecting to the contralateral thalamic ventral posterolateral nucleus (VPL) and posterior nuclear group (Po) nor GABaergic dorsal horn neurons express the OTR. The OTR is not expressed in skin nociceptive terminals or in dorsal horn nociceptive fibers. In the DRG, however, the OTR is expressed predominantly in non-peptidergic C-fiber cell bodies, but not in peptidergic or mechanoreceptor afferents or in skin nociceptive terminals. Our results suggest that the antinociceptive effects of OT are mediated by direct activation of dorsal horn neurons and peripheral actions on nociceptive, non-peptidergic C-afferents in the DRG.
Salman, Maria; Naseem, Imrana; Hassan, Iftekhar; Khan, Aijaz A; Alhazza, Ibrahim M
Cis-Diamminedichloroplatinum II- (CP-) induced neurotoxicity is one of the least explored aspects of this drug. Dorsal root ganglia (DRG) cells are considered as the primary target, and their damage plays a vital role in pathogenesis and etiology of CP-induced neurotoxicity. The present study is aimed at confirming if riboflavin (RF) has any protective role in shielding the DRG from CP-induced toxicity. After conducting the established treatment strategy on mice under photoillumination, it was observed that, despite the fact that RF alone is partially toxic, its combination with CP significantly ameliorated the drug-induced damage in DRG cells as evidenced by histological analysis. In addition, it was interesting to observe that the combination group (RF + CP) was able to induce apoptosis in the target cells up to a significant extent which is considered as the most preferred way of countering cancer cells. Therefore, RF can act as an effective adjuvant compound in CP-based chemoradiotherapy to improve clinical outcomes in the contemporary anticancer treatment regimes.
Briggs, C A; Chandraraj, S
Sixty-five lumbosacral regions from adult cadavers were dissected and the position and relations of the lumbosacral ligament noted. The lumbosacral ligament was present in all specimens; in 22 (34%) it extended medially across the ventral ramus of the fifth lumbar nerve, and in six (9%) of these the underlying nerve was compressed and visibly flattened. On two of these specimens the nerve, together with its dorsal root ganglion, was removed, processed, and stained with Masson's trichrome. The compressed nerve showed increased thickness of endoneurial and perineurial connective tissue, and the cells of the dorsal root ganglion were smaller and surrounded by increased connective tissue, particularly at the periphery of the ganglion. Observation of the lumbosacral ligament and surrounding anatomical structures suggests that anatomical variation in this region may be attributed to the health of the lumbosacral articular elements. In those specimens showing compression of the fifth lumbar spinal nerve there was also narrowing of the lumbosacral interspace. In these the disc itself was compressed and showed degenerative changes. The articular processes at the lumbosacral joint were irregular, with thinning and fissuring of the artiuclar cartilage. It is suggested that the processes which lead to the further development of the ligament, by the formation of additional fibrous bands, are mechanical in nature and result from instability at the lumbosacral region itself. Instability subsequently leads to the initiation of a chain of degenerative changes, involving pathology at the lumbosacral disc and zygapophyseal joints.(ABSTRACT TRUNCATED AT 250 WORDS)
Cao, Lijun; Xun, Junmei; Jiang, Xinghua; Tan, Rong
Mas is a functional binding site for angiotensin (Ang)-(1-7), a critical component of the renin-angiotensin system that is involved in processing nociceptive information. A recent study reported the localization of Mas in rat dorsal root ganglia (DRG) and demonstrated that Ang-(1-7) produced a dose-dependent peripheral antinociceptive effect in rats through the Mas receptor by an opioid-independent mechanism. In the present study, we for the first time examined the effect of propofol on Mas expression in cultured DRG neurons. We treated rat DRG neurons with propofol at different concentrations (0.1, 0.5, 1, 5 or 10 microM) for different length of time (0.5, 1, 2, 4 or 6 h) with or without transcription inhibitor actinomycin D or different kinase inhibitors. Propofol increased the Mas receptormRNA level in a statistically significant dose- and time-dependent manner within 4 h, which led to dose-dependent up-regulation of the Mas receptor protein level as well as Ang-(1-7) binding on the cell membrane. Actinomycin D (1 mg/ml) and p38 mitogen-activated protein kinase inhibitor PD169316 (25 microM) completely abolished the effect of propofol on Mas receptor expression in DRG neurons. In conclusion, we demonstrate that propofol markedly up-regulates Mas receptor expression at the transcription level in DRG neurons by a p38 MAPK-dependent mechanism. This study provides new insights into the mechanisms of action of propofol in peripheral antinociception, and suggests a new regulatory mechanism on the Ang-(1-7)/Mas axis in the peripheral nervous system.
Haskins, William; Benitez, Sergio; Mercado, Juan M; Acosta, Cristian G
Tandem pore-domain Halothane Inhibited K(+) channel (THIK1) is a two-pore-domain potassium channel (K2P) present in dorsal root ganglia (DRG). We previously demonstrated that THIK1 mRNA levels in the DRG dropped ipsilaterally 1day after CFA-induced cutaneous inflammation (CFA1). In this study we aimed to identify the currently unknown DRG subpopulations expressing THIK1, and to investigate the relationship between the channel and both inflammatory and spontaneous pain in normal rats. Using a combination of immunohistochemistry, western blotting and behavioural tests, we found that all small neurons and large groups of medium and large DRG neurons express THIK1. Myelinated and unmyelinated fibers, nerve endings in the skin and lamina I and II of the spinal cord also express the channel. THIK1 staining co-localizes with IB4-binding and trkA suggesting that the channel is expressed by nociceptors. At CFA1, both cytoplasmic and edge (membrane-associated) THIK1 staining were significantly reduced only in small neurons ipsilaterally compared to normal. At 4days after inflammation (CFA4), edge THIK1 staining levels in small neurons decreased bilaterally compared to normal. Medium and large size DRG neurons showed no change in THIK1 expression either at CFA1 or CFA4. Ipsilateral (but not contralateral) mean %intensities of THIK1 in small neurons at CFA1 correlated strongly negatively with spontaneous foot lifting (SFL) duration (a marker of spontaneous pain). Thus, nociceptors express THIK1 that can be regulated by cutaneous inflammation. Finally, in vivo siRNA knockdown of THIK1 resulted in longer SFL duration than siRNA scramble-treated rats. Taken together our evidence suggests a potential involvement for THIK1 in pain processing following inflammation. Copyright © 2017 Elsevier Inc. All rights reserved.
Gong, Kerui; Kung, Ling-Hsuan; Magni, Giulia; Bhargava, Aditi; Jasmin, Luc
Glutamate in the peripheral nervous system is involved in neuropathic pain, yet we know little how nerve injury alters responses to this neurotransmitter in primary sensory neurons. We recorded neuronal responses from the ex-vivo preparations of the dorsal root ganglia (DRG) one week following a chronic constriction injury (CCI) of the sciatic nerve in adult rats. We found that small diameter DRG neurons (<30 µm) exhibited increased excitability that was associated with decreased membrane threshold and rheobase, whereas responses in large diameter neurons (>30 µm) were unaffected. Puff application of either glutamate, or the selective ionotropic glutamate receptor agonists alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and kainic acid (KA), or the group I metabotropic receptor (mGluR) agonist (S)-3,5-dihydroxyphenylglycine (DHPG), induced larger inward currents in CCI DRGs compared to those from uninjured rats. N-methyl-D-aspartate (NMDA)-induced currents were unchanged. In addition to larger inward currents following CCI, a greater number of neurons responded to glutamate, AMPA, NMDA, and DHPG, but not to KA. Western blot analysis of the DRGs revealed that CCI resulted in a 35% increase in GluA1 and a 60% decrease in GluA2, the AMPA receptor subunits, compared to uninjured controls. mGluR1 receptor expression increased by 60% in the membrane fraction, whereas mGluR5 receptor subunit expression remained unchanged after CCI. These results show that following nerve injury, small diameter DRG neurons, many of which are nociceptive, have increased excitability and an increased response to glutamate that is associated with changes in receptor expression at the neuronal membrane. Our findings provide further evidence that glutamatergic transmission in the periphery plays a role in nociception.
Wang, Yiheng; Ni, Hongxia; Zhang, Wenrui; Wang, Xiu; Zhang, Haishan
Local anesthetic may cause neurotoxicity in developing neurons. In this study, we examined the molecular mechanisms of microRNA-210 (miR-210) in regulating bupivacaine-induced dorsal root ganglia (DRG) neurotoxicity in vitro. Young mouse (P30) DRG explants were cultured in vitro and treated with 5 mM bupivacaine to induce neurotoxicity. QRT-PCR was used to evaluate the expression profiles of miRNAs within 24 h after bupivacaine treatment. MiR-210 was downregulated in DRG, and its effects on bupivacaine-induced neurotoxicity were evaluated by apoptosis and neurite growth assays, respectively. Putative downstream target of miR-210 in DRG, BDNF, was evaluated by dual-luciferase assay, qRT-PCR, and western blot, respectively. BDNF was then knocked down by siRNA to assess its associated effects in regulating DRG neurotoxicity. Within the initial 24 h after bupivacaine treatment, various patterns of miRNA expression were observed, whereas miR-210 was constantly upregulated. Application of miR-210 inhibitor efficiently downregulated endogenous miR-210, protected apoptosis and neurite retraction in bupivacaine damaged DRG neurons. Using dual-luciferase assay, qRT-PCR, and western blot, BDNF was confirmed to the downstream target of miR-210 in DRG. SiRNA-mediated BDNF downregulation reversed the effect of miR-210 downregulation in DRG neurotoxicity. MiR-210, through the regulation of BDNF, plays important role in anesthetics-induced DRG neurotoxicity.
Gu, Zirong; Imai, Fumiyasu; Kim, In Jung; Fujita, Hiroko; Katayama, Kei ichi; Mori, Kensaku; Yoshihara, Yoshihiro; Yoshida, Yutaka
Cell adhesion molecules belonging to the immunoglobulin superfamily (IgSF) control synaptic specificity through hetero- or homophilic interactions in different regions of the nervous system. In the developing spinal cord, monosynaptic connections of exquisite specificity form between proprioceptive sensory neurons and motor neurons, however, it is not known whether IgSF molecules participate in regulating this process. To determine whether IgSF molecules influence the establishment of synaptic specificity in sensory-motor circuits, we examined the expression of 157 IgSF genes in the developing dorsal root ganglion (DRG) and spinal cord by in situ hybridization assays. We find that many IgSF genes are expressed by sensory and motor neurons in the mouse developing DRG and spinal cord. For instance, Alcam, Mcam, and Ocam are expressed by a subset of motor neurons in the ventral spinal cord. Further analyses show that Ocam is expressed by obturator but not quadriceps motor neurons, suggesting that Ocam may regulate sensory-motor specificity in these sensory-motor reflex arcs. Electrophysiological analysis shows no obvious defects in synaptic specificity of monosynaptic sensory-motor connections involving obturator and quadriceps motor neurons in Ocam mutant mice. Since a subset of Ocam+ motor neurons also express Alcam, Alcam or other functionally redundant IgSF molecules may compensate for Ocam in controlling sensory-motor specificity. Taken together, these results reveal that IgSF molecules are broadly expressed by sensory and motor neurons during development, and that Ocam and other IgSF molecules may have redundant functions in controlling the specificity of sensory-motor circuits.
Ji, Zhi-Gang; Ito, Shin; Honjoh, Tatsuya; Ohta, Hiroyuki; Ishizuka, Toru; Fukazawa, Yugo; Yawo, Hiromu
In vertebrate somatosensory systems, each mode of touch-pressure, temperature or pain is sensed by sensory endings of different dorsal root ganglion (DRG) neurons, which conducted to the specific cortical loci as nerve impulses. Therefore, direct electrical stimulation of the peripheral nerve endings causes an erroneous sensation to be conducted by the nerve. We have recently generated several transgenic lines of rat in which channelrhodopsin-2 (ChR2) transgene is driven by the Thy-1.2 promoter. In one of them, W-TChR2V4, some neurons were endowed with photosensitivity by the introduction of the ChR2 gene, coding an algal photoreceptor molecule. The DRG neurons expressing ChR2 were immunohistochemically identified using specific antibodies to the markers of mechanoreceptive or nociceptive neurons. Their peripheral nerve endings in the plantar skin as well as the central endings in the spinal cord were also examined. We identified that ChR2 is expressed in a certain population of large neurons in the DRG of W-TChR2V4. On the basis of their morphology and molecular markers, these neurons were classified as mechanoreceptive but not nociceptive. ChR2 was also distributed in their peripheral sensory nerve endings, some of which were closely associated with CK20-positive cells to form Merkel cell-neurite complexes or with S-100-positive cells to form structures like Meissner's corpuscles. These nerve endings are thus suggested to be involved in the sensing of touch. Each W-TChR2V4 rat showed a sensory-evoked behavior in response to blue LED flashes on the plantar skin. It is thus suggested that each rat acquired an unusual sensory modality of sensing blue light through the skin as touch-pressure. This light-evoked somatosensory perception should facilitate study of how the complex tactile sense emerges in the brain.
Fan, Yihong; Hooker, Bradley A; Garrison, Tiffany Runyan; El-Kouhen, Odile F; Idler, Kenneth B; Holley-Shanks, Rhonda R; Meyer, Michael D; Yao, Betty Bei
The behavioral effects evoked by cannabinoids are primarily mediated by the CB(1) and CB(2) cannabinoid receptor subtypes. In vitro pharmacology of cannabinoid receptors has been elucidated using recombinant expression systems expressing either CB(1) or CB(2) receptors, with limited characterization in native cell lines endogenously expressing both CB(1) and CB(2) receptors. In the current study, we report the molecular and pharmacological characterization of the F-11 cell line, a hybridoma of rat dorsal root ganglion neurons and mouse neuroblastoma (N18TG2) cells, reported to endogenously express both cannabinoid receptors. The present study revealed that both receptors are of mouse origin in F-11 cells, and describes the relative gene expression levels between the two receptors. Pharmacological characterization of the F-11 cell line using cannabinoid agonists and antagonists indicated that the functional responses to these cannabinoid ligands are mainly mediated by CB(1) receptors. The non-selective cannabinoid ligands CP 55,940 and WIN 55212-2 are potent agonists and their efficacies in adenylate cyclase and MAPK assays are inhibited by the CB(1) selective antagonist SR141716A (SR1), but not by the CB(2) selective antagonist SR144528 (SR2). The endocannabinoid ligand 2AG, although not active in adenylate cyclase assays, was a potent activator of MAPK signaling in F-11 cells. The analysis of CB(1) and CB(2) receptor gene expression and the characterization of cannabinoid receptor pharmacology in the F-11 cell line demonstrate that it can be used as a tool for interrogating the endogenous signal transduction of cannabinoid receptor subtypes.
Nazıroğlu, Mustafa; Özgül, Cemil; Çiğ, Bilal; Doğan, Salih; Uğuz, Abdulhadi Cihangir
Glutathione (GSH) is the most abundant thiol antioxidant in mammalian cells and maintains thiol redox in the cells. GSH depletion has been implicated in the neurobiology of sensory neurons. Because the mechanisms that lead to melastatin-like transient receptor potential 2 (TRPM2) channel activation/inhibition in response to glutathione depletion and 2-aminoethyldiphenyl borinate (2-APB) administration are not understood, we tested the effects of 2-APB and GSH on oxidative stress and buthionine sulfoximine (BSO)-induced TRPM2 cation channel currents in dorsal root ganglion (DRG) neurons of rats. DRG neurons were freshly isolated from rats and the neurons were incubated for 24 h with BSO. In whole-cell patch clamp experiments, TRPM2 currents in the rat were consistently induced by H(2)O(2) or BSO. TRPM2 channels current densities and cytosolic free Ca(2+) content of the neurons were higher in BSO and H(2)O(2) groups than in control. However, the current densities and cytosolic Ca(2+) release were also higher in the BSO + H(2)O(2) group than in the H(2)O(2) alone. When intracellular GSH is introduced by pipette TRPM2 channel currents were not activated by BSO, H(2)O(2) or rotenone. BSO and H(2)O(2)-induced Ca(2+) gates were blocked by the 2-APB. Glutathione peroxidase activity, lipid peroxidation and GSH levels in the DRG neurons were also modulated by GSH and 2-APB inhibition. In conclusion, we observed the protective role of 2-APB and GSH on Ca(2+) influx through a TRPM2 channel in intracellular GSH depleted DRG neurons. Since cytosolic glutathione depletion is a common feature of neuropathic pain and diseases of sensory neuron, our findings are relevant to the etiology of neuropathology in DRG neurons.
Roh, Kangsan; Kil, Eui-Joon; Lee, Minji; Auh, Chung-Kyun; Lee, Myung-Ah; Yeom, Chang-Hwan; Lee, Sukchan
Oxaliplatin is a platinum-based anticancer drug used to treat metastatic colorectal, breast, and lung cancers. While oxaliplatin kills cancer cells effectively, it exhibits several side effects of varying severity. Neuropathic pain is commonly experienced during treatment with oxaliplatin. Patients describe symptoms of paresthesias or dysesthesias that are triggered by cold (acute neuropathy), or as abnormal sensory or motor function (chronic neuropathy). In particular, we found that aluminum levels were relatively high in some cancer patients suffering from neuropathic pain based on clinical observations. Based on these findings, we hypothesized that aluminum accumulation in the dorsal root ganglion (DRG) in the course of oxaliplatin treatment exacerbates neuropathic pain. In mice injected with oxaliplatin (three cycles of 3 mg/kg i.p. daily for 5 days, followed by 5 days of rest), we detected cold allodynia using the acetone test, but not heat hyperalgesia using a hot plate. However, co-treatment with aluminum chloride (AlCl3∙6H2O; 7 mg/kg i.p. for 14 days: equivalent 0.78 mg/kg of elemental Al) and oxaliplatin (1 cycle of 3 mg/kg i.p. daily for 5 days, followed by 5 days of rest) synergistically induced cold allodynia as well as increased TRPAl mRNA and protein expression. Inductively Coupled Plasma Mass Spectrometry (ICP-MS) analysis showed a significant increase in aluminum concentrations in the DRG of mice treated with aluminum chloride and oxaliplatin compared to aluminum chloride alone. Similarly, in a mouse induced-tumor model, aluminum concentrations were increased in DRG tissue and tumor cells after oxaliplatin treatment. Taken together, these findings suggest that aluminum accumulation in the DRG may exacerbate neuropathic pain in oxaliplatin-treated mice. PMID:25928068
Menon-Johansson, A S; Berrow, N; Dolphin, A C
High-voltage-activated (HVA) calcium channel currents (IBa) were recorded from acutely replated cultured dorsal root ganglion (DRG) neurons. IBa was irreversibly inhibited by 56.9 +/- 2.7% by 1 microM omega-conotoxin-GVIA (omega-CTx-GVIA), whereas the 1,4-dihydropyridine antagonist nicardipine was ineffective. The selective gamma-aminobutyric acidB (GABAB) agonist, (-)-baclofen (50 microM), inhibited the HVA IBa by 30.7 +/- 5.4%. Prior application of omega-CTx-GVIA completely occluded inhibition of the HVA IBa by (-)-baclofen, indicating that in this preparation (-)-baclofen inhibits N-type current. To investigate which G protein subtype was involved, cells were replated in the presence of anti-G protein antisera. Under these conditions the antibodies were shown to enter the cells through transient pores created during the replating procedure. Replating DRGs in the presence of anti-G(o) antiserum, raised against the C-terminal decapeptide of the G alpha o subunit, reduced (-)-baclofen inhibition of the HVA IBa, whereas replating DRGs in the presence of the anti-Gi antiserum did not. Using anti-G alpha o antisera (1:2000) and confocal laser microscopy, G alpha o localisation was investigated in both unreplated and replated neurons. G alpha o immunoreactivity was observed at the plasma membrane, neurites, attachment plaques and perinuclear region, and was particularly pronounced at points of cell-to-cell contact. The plasma membrane G alpha o immunoreactivity was completely blocked by preincubation with the immunising G alpha o undecapeptide (1 microgram.ml-1) for 1 h at 37 degrees C. A similar treatment also blocked recognition of G alpha o in brain membranes on immunoblots.(ABSTRACT TRUNCATED AT 250 WORDS)
Rahman, Md Habibur; Jha, Mithilesh Kumar; Kim, Jong-Heon; Nam, Youngpyo; Lee, Maan Gee; Go, Younghoon; Harris, Robert A; Park, Dong Ho; Kook, Hyun; Lee, In-Kyu; Suk, Kyoungho
The dorsal root ganglion (DRG) is a highly vulnerable site in diabetic neuropathy. Under diabetic conditions, the DRG is subjected to tissue ischemia or lower ambient oxygen tension that leads to aberrant metabolic functions. Metabolic dysfunctions have been documented to play a crucial role in the pathogenesis of diverse pain hypersensitivities. However, the contribution of diabetes-induced metabolic dysfunctions in the DRG to the pathogenesis of painful diabetic neuropathy remains ill-explored. In this study, we report that pyruvate dehydrogenase kinases (PDK2 and PDK4), key regulatory enzymes in glucose metabolism, mediate glycolytic metabolic shift in the DRG leading to painful diabetic neuropathy. Streptozotocin-induced diabetes substantially enhanced the expression and activity of the PDKs in the DRG, and the genetic ablation of Pdk2 and Pdk4 attenuated the hyperglycemia-induced pain hypersensitivity. Mechanistically, Pdk2/4 deficiency inhibited the diabetes-induced lactate surge, expression of pain-related ion channels, activation of satellite glial cells, and infiltration of macrophages in the DRG, in addition to reducing central sensitization and neuroinflammation hallmarks in the spinal cord, which probably accounts for the attenuated pain hypersensitivity. Pdk2/4-deficient mice were partly resistant to the diabetes-induced loss of peripheral nerve structure and function. Furthermore, in the experiments using DRG neuron cultures, lactic acid treatment enhanced the expression of the ion channels and compromised cell viability. Finally, the pharmacological inhibition of DRG PDKs or lactic acid production substantially attenuated diabetes-induced pain hypersensitivity. Taken together, PDK2/4 induction and the subsequent lactate surge induce the metabolic shift in the diabetic DRG, thereby contributing to the pathogenesis of painful diabetic neuropathy.
Xiao, Xing; Zhao, Xiao-Tao; Xu, Ling-Chi; Yue, Lu-Peng; Liu, Feng-Yu; Cai, Jie; Liao, Fei-Fei; Kong, Jin-Ge; Xing, Guo-Gang; Yi, Ming; Wan, You
Transient receptor potential vanilloid 1 (TRPV1) receptors are expressed in nociceptive neurons of rat dorsal root ganglions (DRGs) and mediate inflammatory pain. Nonspecific inhibition of protein-tyrosine phosphatases (PTPs) increases the tyrosine phosphorylation of TRPV1 and sensitizes TRPV1. However, less is known about tyrosine phosphorylation's implication in inflammatory pain, compared with that of serine/threonine phosphorylation. Src homology 2 domain-containing tyrosine phosphatase 1 (Shp-1) is a key phosphatase dephosphorylating TRPV1. In this study, we reported that Shp-1 colocalized with and bound to TRPV1 in nociceptive DRG neurons. Shp-1 inhibitors, including sodium stibogluconate and PTP inhibitor III, sensitized TRPV1 in cultured DRG neurons. In naive rats, intrathecal injection of Shp-1 inhibitors increased both TRPV1 and tyrosine-phosphorylated TRPV1 in DRGs and induced thermal hyperalgesia, which was abolished by pretreatment with TRPV1 antagonists capsazepine, BCTC, or AMG9810. Complete Freund's adjuvant (CFA)-induced inflammatory pain in rats significantly increased the expression of Shp-1, TRPV1, and tyrosine-phosphorylated TRPV1, as well as the colocalization of Shp-1 and TRPV1 in DRGs. Intrathecal injection of sodium stibogluconate aggravated CFA-induced inflammatory pain, whereas Shp-1 overexpression in DRG neurons alleviated it. These results suggested that Shp-1 dephosphorylated and inhibited TRPV1 in DRG neurons, contributing to maintain thermal nociceptive thresholds in normal rats, and as a compensatory mechanism, Shp-1 increased in DRGs of rats with CFA-induced inflammatory pain, which was involved in protecting against excessive thermal hyperalgesia.
Chen, Yu-Wen; Tzeng, Jann-Inn; Lin, Min-Fei; Hung, Ching-Hsia; Wang, Jhi-Joung
Exercise causes a variety of psychophysical effects (eg, alterations in pain sensation). Tissue injury induces mediator releases in the spinal cord resulting in pain hypersensitivity; however, the contribution of the dorsal root ganglion (DRG) is poorly understood. In this study, we tested if forced treadmill running can attenuate postoperative pain and alter substance P (SP) or proinflammatory cytokine level in the DRG by using a rat model of skin/muscle incision and retraction (SMIR). We evaluated mechanical sensitivity to von Frey stimuli (6 and 15 g) and expression of SP, interleukin-1β, and interleukin-6 in the DRG of sham-operated sedentary rats, SMIR sedentary rats, sham-operated rats with forced treadmill running, and SMIR rats with forced treadmill running. At postoperative day 8, trained rats ran for 5 days per week for 4 weeks on a treadmill 70 minutes/d with an intensity of 18 m/min. On postoperative day 6, SMIR sedentary rats displayed a significant mechanical hypersensitivity that persisted until postoperative day 35. By comparison, SMIR-operated rats, which received forced treadmill running, exhibited a quick recovery from mechanical hypersensitivity. SMIR sedentary rats showed an upregulation of SP, interleukin-1β, and interleukin-6 in the DRG at postoperative days 14 and 28, whereas SMIR-operated rats receiving forced treadmill running reversed this upregulation at postoperative day 28. We concluded that forced treadmill running alleviated persistent postincisional pain caused by SMIR surgery. This appears to be protective against postoperative pain, which probably relates to the downturn in excess SP, interleukin-1β, and interleukin-6 in the DRG. Controlling the expression of SP, interleukin-6, and interleukin-1β in the DRG can help manage postoperative pain. This finding could potentially help clinicians and physical therapists who seek to examine how exercise may attenuate postsurgical pain and its mechanism. Copyright © 2014 American Pain
Persson, Anna-Karin; Liu, Shujun; Faber, Catharina G; Merkies, Ingemar S J; Black, Joel A; Waxman, Stephen G
Small-fiber neuropathy (SFN) is characterized by injury to small-diameter peripheral nerve axons and intraepidermal nerve fibers (IENF). Although mechanisms underlying loss of IENF in SFN are poorly understood, available data suggest that it results from axonal degeneration and reduced regenerative capacity. Gain-of-function variants in sodium channel Na(V)1.7 that increase firing frequency and spontaneous firing of dorsal root ganglion (DRG) neurons have recently been identified in ∼30% of patients with idiopathic SFN. In the present study, to determine whether these channel variants can impair axonal integrity, we developed an in vitro assay of DRG neurite length, and examined the effect of 3 SFN-associated variant Na(V)1.7 channels, I228M, M932L/V991L (ML/VL), and I720K, on DRG neurites in vitro. At 3 days after culturing, DRG neurons transfected with I228M channels exhibited ∼20% reduced neurite length compared to wild-type channels; DRG neurons transfected with ML/VL and I720K variants displayed a trend toward reduced neurite length. I228M-induced reduction in neurite length was ameliorated by the use-dependent sodium channel blocker carbamazepine and by a blocker of reverse Na-Ca exchange. These in vitro observations provide evidence supporting a contribution of the I228M variant Na(V)1.7 channel to impaired regeneration and/or degeneration of sensory axons in idiopathic SFN, and suggest that enhanced sodium channel activity and reverse Na-Ca exchange can contribute to a decrease in length of peripheral sensory axons.
Regnier, Glenn; Bocksteins, Elke; Van de Vijver, Gerda; Snyders, Dirk J; van Bogaert, Pierre-Paul
Delayed rectifier voltage-gated K(+)(Kv) channels play an important role in the regulation of the electrophysiological properties of neurons. In mouse dorsal root ganglion (DRG) neurons, a large fraction of the delayed rectifier current is carried by both homotetrameric Kv2 channels and heterotetrameric channels consisting of Kv2 and silent Kv (KvS) subunits (i.e., Kv5-Kv6 and Kv8-Kv9). However, little is known about the contribution of Kv2-mediated currents during the postnatal development ofDRGneurons. Here, we report that the Stromatoxin-1 (ScTx)-sensitive fraction of the total outward K(+)current (IK) from mouseDRGneurons gradually decreased (~13%,P < 0.05) during the first month of postnatal development. Because ScTx inhibits both Kv2.1- and Kv2.2-mediated currents, this gradual decrease may reflect a decrease in currents containing either subunit. However, the fraction of Kv2.1 antibody-sensitive current that only reflects the Kv2.1-mediated currents remained constant during that same period. These results suggested that the fractional contribution of Kv2.2-mediated currents relative toIKdecreased with postnatal age. SemiquantitativeRT-PCRanalysis indicated that this decrease can be attributed to developmental changes in Kv2.2 expression as themRNAlevels of the Kv2.2 subunit decreased gradually between 1 and 4 weeks of age. In addition, we observed age-dependent fluctuations in themRNAlevels of the Kv6.3, Kv8.1, Kv9.1, and Kv9.3 subunits. These results support an important role of both Kv2 and KvS subunits in the postnatal maturation ofDRGneurons.
Park, Jin-Hee; Chae, Jisook; Roh, Kangsan; Kil, Eui-Joon; Lee, Minji; Auh, Chung-Kyun; Lee, Myung-Ah; Yeom, Chang-Hwan; Lee, Sukchan
Oxaliplatin is a platinum-based anticancer drug used to treat metastatic colorectal, breast, and lung cancers. While oxaliplatin kills cancer cells effectively, it exhibits several side effects of varying severity. Neuropathic pain is commonly experienced during treatment with oxaliplatin. Patients describe symptoms of paresthesias or dysesthesias that are triggered by cold (acute neuropathy), or as abnormal sensory or motor function (chronic neuropathy). In particular, we found that aluminum levels were relatively high in some cancer patients suffering from neuropathic pain based on clinical observations. Based on these findings, we hypothesized that aluminum accumulation in the dorsal root ganglion (DRG) in the course of oxaliplatin treatment exacerbates neuropathic pain. In mice injected with oxaliplatin (three cycles of 3 mg/kg i.p. daily for 5 days, followed by 5 days of rest), we detected cold allodynia using the acetone test, but not heat hyperalgesia using a hot plate. However, co-treatment with aluminum chloride (AlCl3∙6H2O; 7 mg/kg i.p. for 14 days: equivalent 0.78 mg/kg of elemental Al) and oxaliplatin (1 cycle of 3 mg/kg i.p. daily for 5 days, followed by 5 days of rest) synergistically induced cold allodynia as well as increased TRPAl mRNA and protein expression. Inductively Coupled Plasma Mass Spectrometry (ICP-MS) analysis showed a significant increase in aluminum concentrations in the DRG of mice treated with aluminum chloride and oxaliplatin compared to aluminum chloride alone. Similarly, in a mouse induced-tumor model, aluminum concentrations were increased in DRG tissue and tumor cells after oxaliplatin treatment. Taken together, these findings suggest that aluminum accumulation in the DRG may exacerbate neuropathic pain in oxaliplatin-treated mice.
EVERILL, B.; KOCSIS, J. D.
Whole-cell patch-clamp techniques were used to study the effects of nerve growth factor on voltage-dependent potassium conductance in normal and axotomized identified large cutaneous afferent dorsal root ganglion neurons (48–50 μm diameter) many of which probably give rise to myelinated Aβ fibers. K-currents were isolated by blocking Na- and Ca-currents with appropriate ion replacement and channel blockers. Separation of current components was achieved on the basis of response to variation in conditioning voltage. Cutaneous afferents were labeled by the retrograde marker hydroxy-stilbamide (FluoroGold) which was injected into the skin of the foot. The sciatic nerve was either ligated or crushed with fine forceps five to seven days later. Neurons were dissociated 14–17 days after injury. The cut ends of the sciatic nerves were positioned into polyethylene tubes, which were connected to mini-osmotic pumps filled with either nerve growth factor or sterile saline. Control neurons displayed a prominent sustained K-current and the transient potassium currents “A” and “D”. Nerve ligation, which blocks target reconnection resulted in near 50% reduction of total outward current; isolated sustained K-current and transient A-current were reduced by a comparable amount. Nerve crush, which allows regeneration to peripheral targets and exposure of the regenerating nerve to the distal nerve segment, resulted in a small reduction in sustained K-current but no reduction in transient A-current compared to controls. Levels of transient A-current and sustained K-current were maintained at control levels after nerve growth factor treatment. These results indicate that the large reduction in transient A-current, and in sustained K-current, observed in cutaneous afferent cell bodies after nerve ligation is prevented by application of nerve growth factor. PMID:11008179
Rahman, Md Habibur; Jha, Mithilesh Kumar; Kim, Jong-Heon; Nam, Youngpyo; Lee, Maan Gee; Go, Younghoon; Harris, Robert A.; Park, Dong Ho; Kook, Hyun; Lee, In-Kyu; Suk, Kyoungho
The dorsal root ganglion (DRG) is a highly vulnerable site in diabetic neuropathy. Under diabetic conditions, the DRG is subjected to tissue ischemia or lower ambient oxygen tension that leads to aberrant metabolic functions. Metabolic dysfunctions have been documented to play a crucial role in the pathogenesis of diverse pain hypersensitivities. However, the contribution of diabetes-induced metabolic dysfunctions in the DRG to the pathogenesis of painful diabetic neuropathy remains ill-explored. In this study, we report that pyruvate dehydrogenase kinases (PDK2 and PDK4), key regulatory enzymes in glucose metabolism, mediate glycolytic metabolic shift in the DRG leading to painful diabetic neuropathy. Streptozotocin-induced diabetes substantially enhanced the expression and activity of the PDKs in the DRG, and the genetic ablation of Pdk2 and Pdk4 attenuated the hyperglycemia-induced pain hypersensitivity. Mechanistically, Pdk2/4 deficiency inhibited the diabetes-induced lactate surge, expression of pain-related ion channels, activation of satellite glial cells, and infiltration of macrophages in the DRG, in addition to reducing central sensitization and neuroinflammation hallmarks in the spinal cord, which probably accounts for the attenuated pain hypersensitivity. Pdk2/4-deficient mice were partly resistant to the diabetes-induced loss of peripheral nerve structure and function. Furthermore, in the experiments using DRG neuron cultures, lactic acid treatment enhanced the expression of the ion channels and compromised cell viability. Finally, the pharmacological inhibition of DRG PDKs or lactic acid production substantially attenuated diabetes-induced pain hypersensitivity. Taken together, PDK2/4 induction and the subsequent lactate surge induce the metabolic shift in the diabetic DRG, thereby contributing to the pathogenesis of painful diabetic neuropathy. PMID:26769971
Joseph, Donald J.; Choudhury, Papiya; MacDermott, Amy B.
Synapses between nociceptive dorsal root ganglion (DRG) neurons and spinal cord dorsal horn neurons represent the first loci for transmission of painful stimuli. Our knowledge of the molecular organization and development of these synapses is sparse due, partly, to a lack of a reliable model system that reconstitutes synaptogenesis between these two neuronal populations. To address this issue, we have established an in vitro assay system consisting of separately purified DRG neurons and dorsal horn neurons on astrocyte micro-islands. Using immunocytochemistry, we have found that 97%, 93%, 98%, 96%, and 94% of DRG neurons on these microislands express markers often associated with nociceptive neurons including Substance P, TRPV1, calcitonin-gene related peptide (CGRP), TrKA, and peripherin, respectively. Triple labeling with these nociceptive-like markers, synaptic vesicle marker Vglut2 and using MAP2 as a dendritic marker revealed the presence of nociceptive-like markers at synaptic terminals. Using this immunocytochemical approach, we counted contact points as overlapping MAP2/Vglut2 puncta and showed that they increased with time in culture. Single and dual patch clamp recordings showed that overlapping Vglut2/MAP2 puncta observed after a few days in culture are likely to be functional synapses between DRG and dorsal horn neurons in our in vitro assay system. Taken together, these data suggest our co-culture microisland model system consists of mostly nociceptive-like DRG neurons that express presynaptic markers and form functional synapses with their dorsal horn partners. Thus, this model system may have direct application for studies on factors regulating development of nociceptive DRG/dorsal horn synapses. PMID:20385165
Mari, João Fernando; Saito, José Hiroki; Neves, Amanda Ferreira; Lotufo, Celina Monteiro da Cruz; Destro-Filho, João-Batista; Nicoletti, Maria do Carmo
Microelectrode Arrays (MEA) are devices for long term electrophysiological recording of extracellular spontaneous or evocated activities on in vitro neuron culture. This work proposes and develops a framework for quantitative and morphological analysis of neuron cultures on MEAs, by processing their corresponding images, acquired by fluorescence microscopy. The neurons are segmented from the fluorescence channel images using a combination of segmentation by thresholding, watershed transform, and object classification. The positioning of microelectrodes is obtained from the transmitted light channel images using the circular Hough transform. The proposed method was applied to images of dissociated culture of rat dorsal root ganglion (DRG) neuronal cells. The morphological and topological quantitative analysis carried out produced information regarding the state of culture, such as population count, neuron-to-neuron and neuron-to-microelectrode distances, soma morphologies, neuron sizes, neuron and microelectrode spatial distributions. Most of the analysis of microscopy images taken from neuronal cultures on MEA only consider simple qualitative analysis. Also, the proposed framework aims to standardize the image processing and to compute quantitative useful measures for integrated image-signal studies and further computational simulations. As results show, the implemented microelectrode identification method is robust and so are the implemented neuron segmentation and classification one (with a correct segmentation rate up to 84%). The quantitative information retrieved by the method is highly relevant to assist the integrated signal-image study of recorded electrophysiological signals as well as the physical aspects of the neuron culture on MEA. Although the experiments deal with DRG cell images, cortical and hippocampal cell images could also be processed with small adjustments in the image processing parameter estimation.
Hone, Arik J; Meyer, Erin L; McIntyre, Melissa; McIntosh, J Michael
The α6-containing nicotinic acetylcholine receptors (nAChRs) have recently been implicated in diseases of the central nervous system (CNS), including Parkinson's disease and substance abuse. In contrast, little is known about the role of α6* nAChRs in the peripheral nervous system (where the asterisk denotes the possible presence of additional subunits). Dorsal root ganglia (DRG) neurons are known to express nAChRs with a pharmacology consistent with an α7, α3β4*, and α4β2* composition. Here we present evidence that DRG neurons also express α6* nAChRs. We used RT-PCR to show the presence of α6 subunit transcripts and patch-clamp electrophysiology together with subtype-selective α-conotoxins to pharmacologically characterize the nAChRs in rat DRG neurons. α-Conotoxin BuIA (500 nM) blocked acetylcholine-gated currents (I(ACh)) by 90.3 ± 3.0%; the recovery from blockade was very slow, indicating a predominance of α(x)β4* nAChRs. Perfusion with either 300 nM BuIA[T5A;P6O] or 200 nM MII[E11A], α-conotoxins that target the α6β4* subtype, blocked I(ACh) by 49.3 ± 5 and 46.7 ± 8%, respectively. In these neurons, I(ACh) was relatively insensitive to 200 nM ArIB[V11L;V16D] (9.4±2.0% blockade) or 500 nM PnIA (23.0±4% blockade), α-conotoxins that target α7 and α3β2*/α6β2* nAChRs, respectively. We conclude that α6β4* nAChRs are among the subtypes expressed by DRG, and to our knowledge, this is the first demonstration of α6β4* in neurons outside the CNS.
Masuoka, Takayoshi; Kudo, Makiko; Yamashita, Yuka; Yoshida, Junko; Imaizumi, Noriko; Muramatsu, Ikunobu; Nishio, Matomo; Ishibashi, Takaharu
The transient receptor potential vanilloid 1 (TRPV1) channel is highly expressed in a subset of sensory neurons in the dorsal root ganglia (DRG) and trigeminal ganglia of experimental animals, responsible for nociception. Many researches have revealed that some TRPV1-positive neurons co-express the transient receptor potential ankyrin 1 (TRPA1) channel whose activities are closely modulated by TRPV1 channel. However, it is less investigated whether the activities of TRPV1 channel are modulated by the presence of TRPA1 channel in primary sensory neurons. This study clarified the difference in electrophysiological responses induced by TRPV1 channel activation between TRPA1-positive and TRPA1-negative DRG. TRPV1 and TRPA1 channel activations were evoked by capsaicin (1 μM), a TRPV1 agonist, and allyl isothiocyanate (AITC; 500 μM), a TRPA1 agonist, respectively. Capsaicin perfusion for 15 s caused a large inward current without a desensitization phase at a membrane potential of -70 mV in AITC-insensitive DRG (current density; 29.6 ± 5.6 pA/pF, time constant of decay; 12.8 ± 1.8 s). The capsaicin-induced currents in AITC-sensitive DRG had a small current density (12.7 ± 2.9 pA/pF) with a large time constant of decay (24.3 ± 5.4 s). In calcium imaging with Fura-2, the peak response by capsaicin was small and duration reaching the peak response was long in AITC-sensitive neurons. These electrophysiological differences were completely eliminated by HC-030031, a TRPA1 antagonist, in an extracellular solution or 10 mM EGTA, a Ca(2+) chelator, in an internal solution. Capsaicin perfusion for 120 s desensitized the inward currents after a transient peak. The decay during capsaicin perfusion was notably slow in AITC-sensitive DRG; ratio of capsaicin-induced current 60 s after the treatment per the peak current in AITC-sensitive neurons (78 ± 9%) was larger than that in AITC-insensitive neurons (48 ± 5%). The capsaicin-induced current in the desensitization phase was
Lee, Kwan Yeop; Charbonnet, Marcel; Gold, Michael S.
Despite evidence that high-affinity GABAA receptor subunit mRNA and protein are present in dorsal root ganglia (DRG), low affinity currents dominate those detected in acutely dissociated DRG neurons in vitro. This observation raises the possibility that high affinity receptors are normally trafficked out of the DRG toward central and peripheral terminals. We therefore hypothesized that with time in culture, there would be an increase in high-affinity GABAA currents in DRG neurons. To test this hypothesis, we studied dissociated DRG neurons 2 hrs (acute) and 24 hrs (cultured) after plating with whole cell patch clamp techniques, western blot and qRT-PCR analysis. GABAA current density increases dramatically with time in culture in association with the emergence of two persistent currents with EC50’s of 0.25 ± 0.01 μM and 3.2 ± 0.02 μM for GABA activation. In a subpopulation of neurons, there was also an increase in the potency of GABA activation of the transient current from an EC50 of 78.16 ± 10.1 μM to 9.56 ± 1.3 μM with time in culture. A fraction of the high affinity current was potentiated by δ-subunit agonist 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridine-3-ol (THIP). δ-subunit immunoreactivity was largely restricted to the cytosolic fraction in acute but the membrane fraction in cultured DRG neurons with no detectable change in δ-subunit mRNA. However, the emergence of a high affinity current blocked by THIP and insensitive to bicuculline was detected in a subpopulation of cultured neurons as well in association with an increase in ρ2 and 3-subunit mRNA in cultured DRG neurons. Our results suggest that high-affinity δ-subunit containing GABAA receptors are normally trafficked out of the DRG where they are targeted to peripheral and central processes. They also highlight that the interpretation of data obtained from cultured DRG neurons should be made with caution. PMID:22366297
van den Wijngaard, Paul W J; Sinnige, Mark P; Roobeek, Ilja; Reumer, Annet; Schoonheim, Peter J; Mol, Jos N M; Wang, Mei; De Boer, Albertus H
Germination of seeds proceeds in general in two phases, an initial imbibition phase and a subsequent growth phase. In grasses like barley, the latter phase is evident as the emergence of the embryonic root (radicle). The hormone abscisic acid (ABA) inhibits germination because it prevents the embryo from entering and completing the growth phase. Genetic and physiological studies have identified many steps in the ABA signal transduction cascade, but how it prevents radicle elongation is still not clear. For elongation growth to proceed, uptake of osmotically active substances (mainly K(+)) is essential. Therefore, we have addressed the question of how the activity of K(+) permeable ion channels in the plasma membrane of radicle cells is regulated under conditions of slow (+ABA) and rapid germination (+fusicoccin). We found that ABA arrests radicle growth, inhibits net K(+) uptake and reduces the activity of K(+) (in) channels as measured with the patch-clamp technique. In contrast, fusicoccin (FC), a well-known stimulator of germination, stimulates radicle growth, net K(+) uptake and reduces the activity of K(+) (out) channels. Both types of channels are under the control of 14-3-3 proteins, known as integral components of signal transduction pathways and instrumental in FC action. Intriguingly, 14-3-3 affected both channels in an opposite fashion: whereas K(+) (in) channel activity was fully dependent upon 14-3-3 proteins, K(+) (out) channel activity was reduced by 14-3-3 proteins by 60%. Together with previous data showing that 14-3-3 proteins control the activity of the plasma membrane H(+)-ATPase, this makes 14-3-3 a prime candidate for molecular master regulator of the cellular osmo-pump. Regulation of the osmo-pump activity by ABA and FC is an important mechanism in controlling the growth of the embryonic root during seed germination.
The diversity of neurons in sympathetic ganglia and dorsal root ganglia (DRG) provides intriguing systems for the analysis of neuronal differentiation. Cell surface receptors for the GDNF family ligands (GFLs) glial cell-line-derived neurotrophic factor (GDNF), neurturin and artemin, are expressed in subpopulations of these neurons prompting the question regarding their involvement in neuronal subtype specification. Mutational analysis in mice has demonstrated the requirement for GFL signalling during embryonic development of cholinergic sympathetic neurons as shown by the loss of expression from the cholinergic gene locus in ganglia from mice deficient for ret, the signal transducing subunit of the GFL receptor complex. Analysis in mutant animals and transgenic mice overexpressing GFLs demonstrates an effect on sensitivity to thermal and mechanical stimuli in DRG neurons correlating at least partially with the altered expression of transient receptor potential ion channels and acid-sensitive cation channels. Persistence of targeted cells in mutant ganglia suggests that the alterations are caused by differentiation effects and not by cell loss. Because of the massive effect of GFLs on neurite outgrowth, it remains to be determined whether GFL signalling acts directly on neuronal specification or indirectly via altered target innervation and access to other growth factors. The data show that GFL signalling is required for the specification of subpopulations of sensory and autonomic neurons. In order to comprehend this process fully, the role of individual GFLs, the transduction of the GFL signals, and the interplay of GFL signalling with other regulatory pathways need to be deciphered. PMID:18629541
Simopoulos, Thomas T; Kraemer, Jan; Nagda, Jyotsna V; Aner, Musa; Bajwa, Zahid H
We aimed to prospectively evaluate the response and safety of pulsed and continuous radiofrequecy lesioning of the dorsal root ganglion/segmental nerves in patients with chronic lumbosacral radicular pain. Seventy-six patients with chronic lumbosacral radicular pain refractory to conventional therapy met the inclusion criteria and were randomly assigned to one of 2 types of treatment, pulsed radiofrequency lesioning of the dorsal root ganglion/segmental nerve or pulsed radiofrequency followed immediately by continuous radiofrequency. Patients were carefully evaluated for neurologic deficits and side effects. The response was evaluated at 2 months and was then tracked monthly. A Kaplan-Meier analysis was used to illustrate the probability of success over time and a Box-Whisker analysis was applied to determine the mean duration of a successful analgesic effect. Two months after undergoing radiofrequency treatment, 70% of the patients treated with pulsed radiofrequency and 82% treated with pulsed and continuous radiofrequency had a successful reduction in pain intensity. The average duration of successful analgesic response was 3.18 months (+/- 2.81) in the group treated with pulsed radiofrequency and 4.39 months (+/-3.50) in those patients treated with pulsed and continuous radiofrequency lesioning. A Kaplan-Meier analysis illustrated that in both treatment groups the chance of success approached 50% in each group at 3 months. The vast majority of patients had lost any beneficial effects by 8 months. There was no statistical difference between the 2 treatment groups. No side effects or neurological deficits were found in either group. Pulsed mode radiofrequency of the dorsal root ganglion of segmental nerves appears to be a safe treatment for chronic lumbosacral radicular pain. A significant number of patients can derive at least a short-term benefit. The addition of heat via continuous radiofrequency does not offer a significant advantage. A randomized controlled
Wickley, Peter J.; Yuge, Ryo; Russell, Mary S.; Zhang, Hongyu; Sulak, Michael A.; Damron, Derek S.
Background The activity of transient receptor potential vanilloid subtype-1 (TRPV1) receptors, key nociceptive transducers in dorsal root ganglion sensory neurons, is enhanced by protein kinase C ε (PKCε) activation. The intravenous anesthetic propofol has been shown to activate PKCε. Our objectives were to examine whether propofol modulates TRPV1 function in dorsal root ganglion neurons via activation of PKCε. Methods Lumbar dorsal root ganglion neurons from wild-type and PKCε-null mice were isolated and cultured for 24 h. Intracellular free Ca2+ concentration was measured in neurons by using fura-2 acetoxymethyl ester. The duration of pain-associated behaviors was also assessed. Phosphorylation of PKCε and TRPV1 and the cellular translocation of PKCε from cytosol to membrane compartments were assessed by immunoblot analysis. Results In wild-type neurons, repeated stimulation with capsaicin (100 nM) progressively decreased the transient rise in intracellular free Ca2+ concentration. After desensitization, exposure to propofol rescued the Ca2+ response. The resensitizing effect of propofol was absent in neurons obtained from PKCε-null mice. Moreover, the capsaicin-induced desensitization of TRPV1 was markedly attenuated in the presence of propofol in neurons from wild-type mice but not in neurons from PKCε-null mice. Propofol also prolonged the duration of agonist-induced pain associated behaviors in wild-type mice. In addition, propofol increased phosphorylation of PKCε as well as TRPV1 and stimulated translocation of PKCε from cytosolic to membrane fraction. Discussion Our results indicate that propofol modulates TRPV1 sensitivity to capsaicin and that this most likely occurs through a PKCε-mediated phosphorylation of TRPV1. PMID:20808213
Phuket, Thanawath Ratanadilok Na; Covarrubias, Manuel
The dorsal root ganglion (DRG) contains heterogeneous populations of sensory neurons including primary nociceptive neurons and C-fibers implicated in pain signaling. Recent studies have demonstrated DRG hyperexcitability associated with downregulation of A-type K(+) channels; however, the molecular correlate of the corresponding A-type K(+) current (I(A)) has remained hypothetical. Kv4 channels may underlie the I(A) in DRG neurons. We combined electrophysiology, molecular biology (Whole-Tissue and Single-Cell RT-PCR) and immunohistochemistry to investigate the molecular basis of the I(A) in acutely dissociated DRG neurons from 7- to 8-day-old rats. Whole-cell recordings demonstrate a robust tetraethylammonium-resistant (20 mM) and 4-aminopyridine-sensitive (5 mM) I(A). Matching Kv4 channel properties, activation and inactivation of this I(A) occur in the subthreshold range of membrane potentials and the rate of recovery from inactivation is rapid and voltage-dependent. Among Kv4 transcripts, the DRG expresses significant levels of Kv4.1 and Kv4.3 mRNAs. Also, single small-medium diameter DRG neurons ( approximately 30 mum) exhibit correlated frequent expression of mRNAs encoding Kv4.1 and Nav1.8, a known nociceptor marker. In contrast, the expressions of Kv1.4 and Kv4.2 mRNAs at the whole-tissue and single-cell levels are relatively low and infrequent. Kv4 protein expression in nociceptive DRG neurons was confirmed by immunohistochemistry, which demonstrates colocalization of Kv4.3 and Nav1.8, and negligible expression of Kv4.2. Furthermore, specific dominant-negative suppression and overexpression strategies confirmed the contribution of Kv4 channels to I(A) in DRG neurons. Contrasting the expression patterns of Kv4 channels in the central and peripheral nervous systems, we discuss possible functional roles of these channels in primary sensory neurons.
Gao, L L; McMullan, S; Djouhri, L; Acosta, C; Harper, A A; Lawson, S N
The hyperpolarization-activated current (Ih) has been implicated in nociception/pain, but its expression levels in nociceptors remained unknown. We recorded Ih magnitude and properties by voltage clamp from dorsal root ganglion (DRG) neurons in vivo, after classifying them as nociceptive or low-threshold-mechanoreceptors (LTMs) and as having C-, Aδ- or Aα/β-conduction velocities (CVs). For both nociceptors and LTMs, Ih amplitude and Ih density (at −100 mV) were significantly positively correlated with CV. Median Ih magnitudes and Ih density in neuronal subgroups were respectively: muscle spindle afferents (MSAs): −4.6 nA, −33 pA pF−1; cutaneous Aα/β LTMs: −2.2 nA, −20 pA pF−1; Aβ-nociceptors: −2.6 nA, −21 pA pF−1; both Aδ-LTMs and nociceptors: −1.3 nA, ∼−14 pA pF−1; C-LTMs: −0.4 nA, −7.6 pA pF−1; and C-nociceptors: −0.26 nA, −5 pA pF−1. Ih activation slow time constants (slow τ values) were strongly correlated with fast τ values; both were shortest in MSAs. Most neurons had τ values consistent with HCN1-related Ih; others had τ values closer to HCN1+HCN2 channels, or HCN2 in the presence of cAMP. In contrast, median half-activation voltages (V0.5) of −80 to −86 mV for neuronal subgroups suggest contributions of HCN2 to Ih. τ values were unrelated to CV but were inversely correlated with Ih and Ih density for all non-MSA LTMs, and for Aδ-nociceptors. From activation curves ∼2–7% of Ih would be activated at normal membrane potentials. The high Ih may be important for excitability of A-nociceptors (responsible for sharp/pricking-type pain) and Aα/β-LTMs (tactile sensations and proprioception). Underlying HCN expression in these subgroups therefore needs to be determined. Altered Ih expression and/or properties (e.g. in chronic/pathological pain states) may influence both nociceptor and LTM excitability. PMID:22753545
Castro, Joel; Harrington, Andrea M; Garcia-Caraballo, Sonia; Maddern, Jessica; Grundy, Luke; Zhang, Jingming; Page, Guy; Miller, Paul E; Craik, David J; Adams, David J; Brierley, Stuart M
α-Conotoxin Vc1.1 is a small disulfide-bonded peptide from the venom of the marine cone snail Conus victoriae. Vc1.1 has antinociceptive actions in animal models of neuropathic pain, but its applicability to inhibiting human dorsal root ganglion (DRG) neuroexcitability and reducing chronic visceral pain (CVP) is unknown. We determined the inhibitory actions of Vc1.1 on human DRG neurons and on mouse colonic sensory afferents in healthy and chronic visceral hypersensitivity (CVH) states. In mice, visceral nociception was assessed by neuronal activation within the spinal cord in response to noxious colorectal distension (CRD). Quantitative-reverse-transcription-PCR, single-cell-reverse-transcription-PCR and immunohistochemistry determined γ-aminobutyric acid receptor B (GABABR) and voltage-gated calcium channel (CaV2.2, CaV2.3) expression in human and mouse DRG neurons. Vc1.1 reduced the excitability of human DRG neurons, whereas a synthetic Vc1.1 analogue that is inactive at GABABR did not. Human DRG neurons expressed GABABR and its downstream effector channels CaV2.2 and CaV2.3. Mouse colonic DRG neurons exhibited high GABABR, CaV2.2 and CaV2.3 expression, with upregulation of the CaV2.2 exon-37a variant during CVH. Vc1.1 inhibited mouse colonic afferents ex vivo and nociceptive signalling of noxious CRD into the spinal cord in vivo, with greatest efficacy observed during CVH. A selective GABABR antagonist prevented Vc1.1-induced inhibition, whereas blocking both CaV2.2 and CaV2.3 caused inhibition comparable with Vc1.1 alone. Vc1.1-mediated activation of GABABR is a novel mechanism for reducing the excitability of human DRG neurons. Vc1.1-induced activation of GABABR on the peripheral endings of colonic afferents reduces nociceptive signalling. The enhanced antinociceptive actions of Vc1.1 during CVH suggest it is a novel candidate for the treatment for CVP. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a
Hall, K E; Sima, A A; Wiley, J W
1. Whole-cell, high-threshold, voltage-dependent calcium currents (ICa) were enhanced in acutely dissociated, capsaicin-sensitive dorsal root ganglion neurones from diabetic Bio Bred/Worchester (BB/W) rats, compared with those from age-matched, non-diabetic controls. The magnitude of the enhancement increased with the duration of diabetes, and reached significance at diabetic durations of 6 months (diabetic: 6.3 +/- 0.4 nA; current density (CD), 157 +/- 12 pA pF-1; means +/- S.E.M., n = 9, P < 0.01; control: 3.9 +/- 0.6 nA; CD, 116 +/- 11 pA pF-1; n = 18) and 8 months (diabetic: 7.6 +/- 0.4 nA; CD, 177 +/- 25 pA pF-1; n = 11, P < 0.005; control: 5.1 +/- 0.5 nA; CD, 111 +/- 26 pA pF-1; n = 15). Low-threshold, voltage-dependent ICa were also enhanced in neurones from animals diabetic for 8 months (diabetic: 2.5 +/- 0.7 nA, n = 4, P < 0.05; control: 0.7 +/- 0.5 nA, n = 6). 2. The ICa enhancement was prevented by long-term treatment of diabetic animals with an aldose reductase inhibitor (ARI; peak ICa at 6 months: 4.41 +/- 0.48 nA, n = 2; at 8 months: 4.32 +/- 0.60 nA, n = 9). 3. The ICa enhancement was not due to a shift in the voltage dependence of either the current-voltage relationship or steady-state inactivation. 4. The L channel antagonist nifedipine and preferential N channel antagonist omega-conotoxin GVIA (omega-CgTX) caused a greater inhibition of high-threshold ICa in diabetic neurones compared with controls (nifedipine: control: 25 +/- 3%, n = 26; diabetic: 36 +/- 7%, n = 11; omega-CgTX: control: 40 +/- 4%, n = 21; diabetic: 50 +/- 7%, n = 7). Diabetic neurones also demonstrated a significantly greater residual current (2.44 +/- 0.34 nA, n = 7) in the presence of both antagonists vs. controls (1.28 +/- 0.30 nA, n = 8, P < 0.05), suggesting that N-, L- and additional non-N-, non-L-type high-threshold ICa were enhanced. Images Figure 2 PMID:7473199
Xing, Jihong; Lu, Jian; Li, Jianhua
The responsiveness of sensory neurons to muscle metabolites is altered under the conditions of insufficient limb blood supply in some diseases, such as peripheral artery disease. The purpose of this study was to examine ATP-induced current with activation of purinergic P2X subtypes P2X₃ and P2X₂/₃ in dorsal root ganglion (DRG) neurons of control limbs and limbs with 24 h of femoral artery occlusion using whole cell patch-clamp methods. Also, dual-labeling immunohistochemistry was employed to determine existence of P2X₃ expression in DRG neurons of thin-fiber afferents. DRG neurons from 4- to 6-wk-old rats were labeled by injecting the fluorescence tracer DiI into the hindlimb muscles 4-5 days before the recording experiments. Transient (P2X₃), mixed (P2X₃ and P2X₂/₃), and sustained (P2X₂/₃) current responses to α,β-methylene ATP (a P2X receptor agonist) are observed in small and medium DRG neurons, and size distribution of DRG neurons is similar in control and occluded limbs. However, the peak current amplitude of DRG neuron induced by stimulation of P2X₃ and/or P2X₂/₃ is larger in occluded limbs than that in control limbs. Moreover, the percentage of DRG neurons with P2X₃ transient currents is greater after arterial occlusion compared with control. In addition, a rapid desensitization was observed in DRG neurons with transient currents, but not with sustained currents in control and occluded groups. Furthermore, results from immunofluorescence experiments show that femoral artery occlusion primarily augments P2X₃ expression within DRG neurons projecting C-fiber afferents. Overall, these findings suggest that 1) greater ATP-induced currents with activation of P2X₃ and P2X₂/₃ are developed when hindlimb arterial blood supply is deficient under ischemic conditions and 2) increased P2X₃ expression is largely observed in C-fibers of DRG neurons after hindlimb vascular insufficiency.
Takenami, Tamie; Hiruma, Hiromi; Kaneko, Haruka; Okamoto, Hirotsugu; Kawakami, Tadashi
Sodium bisulfite (NaHSO3) was clinically used as a preservative agent for local anesthetics but was later suspected to be neurotoxic. However, recent studies reported that NaHSO3 reduces the neurotoxicity of local anesthetics. The purpose of this study was to examine the effects of NaHSO3 with and without procaine on axonal transport in cultured mouse dorsal root ganglion (DRG) neurons. Experiment 1 served to determine the dose-dependent effects of NaHSO3 on axonal transport (DRG neurons were treated with 0.01, 0.1, 1, 10, or 20 mM of NaHSO3), whereas experiment 2 investigated the effect of 0.1 mM NaHSO3 on the action of local anesthetics on axonal transport (DRG neurons were treated with 1 mM procaine alone, or with 0.1 mM NaHSO3 plus 1 mM procaine). As an additional experiment, DRG neurons were also treated with 1 mM chloroprocaine alone, or with 0.1 mM NaHSO3 plus 1 mM chloroprocaine. In these experiments, we analyzed the percent change in the number of anterogradely and retrogradely transported organelles and recorded changes in neurite morphology using video-enhanced microscopy. In experiment 1, NaHSO3 at more than 1 mM caused cell membrane damage and complete inhibition of axonal transport, whereas 0.1 mM NaHSO3 maintained axonal transport at 40% to 60% of control with intact cell membrane. In experiment 2, 1 mM procaine alone maintained axonal transport at 90% to 100%. However, application of 1 mM procaine-0.1 mM NaHSO3 solution resulted in deformation of neurites and with complete cessation of axonal transport. Likewise, although 1 mM chloroprocaine maintain axonal transport at 80% to 100%, 1 mM chloroprocaine-0.1 mM NaHSO3 arrested axonal transport. NaHSO3 resulted in a dose-dependent damage to the cell membrane and axonal transport, especially when used in combination with procaine or chloroprocaine.
Bocksteins, Elke; Van de Vijver, Gerda; Van Bogaert, Pierre-Paul; Snyders, Dirk J
Delayed rectifier voltage-gated K(+) (K(V)) channels are important determinants of neuronal excitability. However, the large number of K(V) subunits poses a major challenge to establish the molecular composition of the native neuronal K(+) currents. A large part (∼60%) of the delayed rectifier current (I(K)) in small mouse dorsal root ganglion (DRG) neurons has been shown to be carried by both homotetrameric K(V)2.1 and heterotetrameric channels of K(V)2 subunits with silent K(V) subunits (K(V)S), while a contribution of K(V)1 channels has also been demonstrated. Because K(V)3 subunits also generate delayed rectifier currents, we investigated the contribution of K(V)3 subunits to I(K) in small mouse DRG neurons. After stromatoxin (ScTx) pretreatment to block the K(V)2-containing component, application of 1 mM TEA caused significant additional block, indicating that the ScTx-insensitive part of I(K) could include K(V)1, K(V)3, and/or M-current channels (KCNQ2/3). Combining ScTx and dendrotoxin confirmed a relevant contribution of K(V)2 and K(V)2/K(V)S, and K(V)1 subunits to I(K) in small mouse DRG neurons. After application of these toxins, a significant TEA-sensitive current (∼19% of total I(K)) remained with biophysical properties that corresponded to those of K(V)3 currents obtained in expression systems. Using RT-PCR, we detected K(V)3.1-3 mRNA in DRG neurons. Furthermore, Western blot and immunocytochemistry using K(V)3.1-specific antibodies confirmed the presence of K(V)3.1 in cultured DRG neurons. These biophysical, pharmacological, and molecular results demonstrate a relevant contribution (∼19%) of K(V)3-containing channels to I(K) in small mouse DRG neurons, supporting a substantial role for K(V)3 subunits in these neurons.
Lu, Tan; Zhang, Chao; Chai, Mingxiang; An, Yongbo
Recent studies showed that Isoquercetin (Iso), a novel extracts of plants and fruits could protect neuronal cells from neurotoxicity and neuro-inflammation. However, its role in acute spinal cord injury (ASCI) have not been elucidated. In the present study, we investigated whether Iso could prevent Tunicamycin (TUN)-induced rat dorsal root ganglion (DRG) neurons from apoptosis and endoplasmic reticulum (ER) stress. DRG neurons were pre-treated with different doses of Iso for 24h (h) and then were stimulated with TUN (0.75μg/ml) for 24h. The cytotoxic effects and apoptosis were detected by MTT assay and TUNEL staining. The protein and mRNA expression levels were detected by Real Time PCR and Western blot, respectively. The localization of cleaved caspase-12 was evaluated by immunofluorescent staining and Western blot. The activation of caspase were measured by colorimetric assays and Western blot. Lactate Dehydrogenase (LDH) and Malondialdehyde (MDA) leakage were detected by the LDH or MDA Detection Kit PLUS. Iso protected TUN-associated DRG neurons from being damaged by cytotoxicity and apoptosis in a dose-dependent manner. Increased LDH and MDA leakage and proportion of TUNEL-positive cells, activation of caspase-3 and -9, increased Bcl-2 Assaciated X protein (Bax)/B cell lymphoma/lewkmia-2 (Bcl-2) ratio and mRNA levels of p53 Upregulated Modulator Of Apoptosis (PUMA) and DP5, and mitochondrial Cytochrome c release. Additionally, Iso down-regulated mRNA levels of ER stress genes, such as glucose-regulated protein 78 (GRP78), GRP94, C/EBP homologous protein (CHOP), and cleaved caspase-12 in TUN-induced DRG neurons. Moreover, Iso blocked the activation of three key branches of unfolded protein response in DRG neurons, including phosphorylation of pancreatic ER stress kinase (PERK), eukaryotic initiation factor 2 alpha (eIF2α), inositol-requiring enzyme 1α (IRE1α), and transcription factor 6 (ATF6). Collectively, Iso prevented TUN-induced DRG neurons apoptosis by
Siemionow, Krzysztof; Klimczak, Aleksandra; Brzezicki, Grzegorz; Siemionow, Maria; McLain, Robert F
After undergoing L5 hemilaminectomy, chromic gut suture was placed onto the DRG and the animals were sacrificed at various time-points. The purpose of this study was to identify the effects of inflammation on satellite cells (SCs) of the dorsal root ganglion (DRG) by analyzing glial fibrillary acidic protein (GFAP) expression in of the DRG at various time points. SCs are neuroglial cells that closely interact with nerve cells of the DRG. The role of SC remains unknown GFAP expression increases in response to CNS injury. Loss of GFAP has impaired Schwann cell proliferation and delayed nerve regeneration after injury. Sixty rats underwent a left L5 hemilaminectomy. In Group I, a chromic-gut suture was place topically on the DRG (n = 30), Group II was the sham surgery group (n = 30). DRGs were harvested at 6, 24, 48, 72 hours, and 7 days after surgery. In Group III, 6 control rats were killed and their bilateral L5 DRG harvested. The harvested DRG were analyzed using light microscopy for SC immunoreactivity, using GFAP, HIS-36, TNF-alpha, IL-1alpha, IL-1beta, IL-6 monoclonal antibodies. One hundred thirty-two DRGs were harvested for analysis. Naïve controls and neurons did not express GFAP. The SC sheath expressed GFAP as early as 6 hours postchromic gut application. In Group I, GFAP expression steadily increased after chromic-gut application with 100% of SC soma and SC sheaths being GFAP positive at 7 days. The contralateral DRG demonstrated delayed GFAP expression, with 83% of SC soma and SC sheaths were GFAP positive at 7 days. In Group II, 89% of sacs expressed GFAP by 7 compared to 79% in the contralateral undisturbed DRG. Under physiologic conditions, the expression of GFAP by SCs is undetectable. As the inflammatory process develops, GFAP expression steadily increases with 100% of SCs being GFAP immunoreactive 7 days after chromic gut application. These data suggest that SCs are the primary source of GFAP in the DRG. We hypothesize that SC play an important
ZHAO, LEI; LI, LI; MA, KE-TAO; WANG, YANG; LI, JING; SHI, WEN-YAN; ZHU, HE; ZHANG, ZHONG-SHUANG; SI, JUN-QIANG
The ability of non-steroidal anti-inflammatory drugs (NSAIDs) to modulate γ-aminobutyrate (GABA)-activated currents via Ca2+-activated Cl− channels in rat dorsal root ganglion neurons (DRG), was examined in the present study. During the preparation of DRG neurons harvested from Sprague-Dawley rats, the whole-cell recording technique was used to record the effect of NSAIDs on GABA-activated inward currents, and the expression levels of the TMEM16A and TMEM16B subunits were revealed. In the event that DRG neurons were pre-incubated for 20 sec with niflumic acid (NFA) and 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB) prior to the administration of GABA, the GABA-induced inward currents were diminished markedly in the majority of neurons examined (96.3%). The inward currents induced by 100 µmol/l GABA were attenuated by (0±0.09%; neurons = 4), (5.32±3.51%; neurons = 6), (21.3±4.00%; neurons = 5), (33.8±5.20%; neurons = 17), (52.2±5.10%; neurons = 4) and (61.1±4.12%; neurons = 12) by 0.1, 1, 3, 10, 30 and 100 µmol/l NFA, respectively. The inward currents induced by 100 µmol/l GABA were attenuated by (13.8±6%; neurons = 6), (23.2±14.7%; neurons = 6) and (29.7±9.1%; neurons = 9) by 3, 10 and 30 µmol/l NPPB, respectively. NFA and NPPB dose-dependently inhibited GABA-activated currents with half maximal inhibitory concentration (IC50) values of 6.7 and 11 µmol/l, respectively. The inhibitory effect of 100 µmol/l NFA on the GABA-evoked inward current were also strongly inhibited by nitrendipine (NTDP; an L-type calcium channel blocker), 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid tetrakis (a highly selective calcium chelating reagent), caffeine (a widely available Ca2+ consuming drug) and calcium-free extracellular fluid, in a concentration-dependent manner. Immunofluorescent staining indicated that TMEM16A and TMEM16B expression was widely distributed in DRG neurons. The results suggest that NSAIDs may be able to regulate Ca2
Reguzzoni, Marcella; Sangiorgi, Simone; Reverberi, Claudio; Borsani, Elisa; Rodella, Luigi F.; Dario, Alessandro; Tomei, Giustino; Dell’Orbo, Carlo
Since the dorsal root ganglia represent the first structure of pain modulation, they are the target of the newest therapies of neuropathic pain. Between these, pulsed radiofrequency (PRF) has been described among the promising non-invasive methods. Although the results encourage the clinical use of this procedure, their mechanism of action is still unclear. Aim of our study was to analyze acute effects of PRF on the rat lumbar ganglion and on nervous fibres running inside it. Clinical works describe PRF treatment as a techinque without any visible neurological deficit. The few disposable histological works are contractictory: some describe no signs of cellular damage and some demonstrate visible intracellular modifications. A total of 20 male Wistar rats were deeply anesthesized. Ten were positioned in a stereotactic system, and exposed to PRF at 2 Hz for 30 s after exposition of paravertebral muscles and positioning of a stimulation needle on left L4 ganglion. The other ten were used as controls. After 1 h, the left dorsal root ganglions L3, L4, L5 of the 20 animals were explanted, fixed in 2.5% Karnowsky solution and prepared for light and transmission electron microscopy. At light microscopy no differences between treated and control animals were observed; at transmission electron microscopy, instead, it was possible to observe that T gangliar cells contained an abnormal abundant smooth reticulum with enlarged cisternae and numerous vacuoles; myelinated axons presented pathological features and their myelin coverage was not adherent. Instead, unmyelinated axons appeared normal in shape and dimension and the Schwann cells surrounding it had intact plasmamembrane. Our results, obtained at acute stage, reveal that the PRF procedure should destroy the myelin envelope of nervous fibres. Further future studies, at chronic stage, should give other information on the prognosis of the myelinic damage. PMID:19172311
Qu, Yu-Juan; Zhang, Xiao; Fan, Zhen-Zhen; Huai, Juan; Teng, Yong-Bo; Zhang, Yang; Yue, Shou-Wei
The aim of this study was to investigate the relationships among TRPV4, p38, and neuropathic pain in a rat model of chronic compression of the dorsal root ganglion. Mechanical allodynia appeared after CCD surgery, enhanced via the intrathecal injection of 4α-phorbol 12,13-didecanoate (4α-PDD, an agonist of TRPV4) and anisomycin (an agonist of p38), but was suppressed by Ruthenium Red (RR, an inhibitor of TRPV4) and SB203580 (an inhibitor of p38). The protein expressions of p38 and P-p38 were upregulated by 4α-PDD and anisomycin injection but reduced by RR and SB203580. Moreover, TRPV4 was upregulated by 4α-PDD and SB203580 and downregulated by RR and anisomycin. In DRG tissues, the numbers of TRPV4- or p38-positive small neurons were significantly changed in CCD rats, increased by the agonists, and decreased by the inhibitors. The amplitudes of ectopic discharges were increased by 4α-PDD and anisomycin but decreased by RR and SB203580. Collectively, these results support the link between TRPV4 and p38 and their intermediary role for neuropathic pain in rats with chronic compression of the dorsal root ganglion. PMID:27366753
Burland, Marion; Paris, Lambert; Quintana, Patrice; Bec, Jean-Michel; Diouloufet, Lucie; Sar, Chamroeun; Boukhaddaoui, Hassan; Charlot, Benoit; Braga Silva, Jefferson; Chammas, Michel; Sieso, Victor; Valmier, Jean; Bardin, Fabrice
The effect of a 645 nm Light Emitting Diode (LED) light irradiation on the neurite growth velocity of adult Dorsal Root Ganglion (DRG) neurons with peripheral axon injury 4-10 days before plating and without previous injury was investigated. The real amount of light reaching the neurons was calculated by taking into account the optical characteristics of the light source and of media in the light path. The knowledge of these parameters is essential to be able to compare results of the literature and a way to reduce inconsistencies. We found that 4 min irradiation of a mean irradiance of 11.3 mW/cm(2) (corresponding to an actual irradiance reaching the neurons of 83 mW/cm(2)) induced a 1.6-fold neurite growth acceleration on non-injured neurons and on axotomized neurons. Although the axotomized neurons were naturally already in a rapid regeneration process, an enhancement was found to occur while irradiating with the LED light, which may be promising for therapy applications. Dorsal Root Ganglion neurons (A) without previous injury and (B) subjected to a conditioning injury.
Qu, Yu-Juan; Zhang, Xiao; Fan, Zhen-Zhen; Huai, Juan; Teng, Yong-Bo; Zhang, Yang; Yue, Shou-Wei
The aim of this study was to investigate the relationships among TRPV4, p38, and neuropathic pain in a rat model of chronic compression of the dorsal root ganglion. Mechanical allodynia appeared after CCD surgery, enhanced via the intrathecal injection of 4α-phorbol 12,13-didecanoate (4α-PDD, an agonist of TRPV4) and anisomycin (an agonist of p38), but was suppressed by Ruthenium Red (RR, an inhibitor of TRPV4) and SB203580 (an inhibitor of p38). The protein expressions of p38 and P-p38 were upregulated by 4α-PDD and anisomycin injection but reduced by RR and SB203580. Moreover, TRPV4 was upregulated by 4α-PDD and SB203580 and downregulated by RR and anisomycin. In DRG tissues, the numbers of TRPV4- or p38-positive small neurons were significantly changed in CCD rats, increased by the agonists, and decreased by the inhibitors. The amplitudes of ectopic discharges were increased by 4α-PDD and anisomycin but decreased by RR and SB203580. Collectively, these results support the link between TRPV4 and p38 and their intermediary role for neuropathic pain in rats with chronic compression of the dorsal root ganglion.
Expressing Constitutively Active Rheb in Adult Dorsal Root Ganglion Neurons Enhances the Integration of Sensory Axons that Regenerate Across a Chondroitinase-Treated Dorsal Root Entry Zone Following Dorsal Root Crush
Wu, Di; Klaw, Michelle C.; Kholodilov, Nikolai; Burke, Robert E.; Detloff, Megan R.; Côté, Marie-Pascale; Tom, Veronica J.
While the peripheral branch of dorsal root ganglion neurons (DRG) can successfully regenerate after injury, lesioned central branch axons fail to regrow across the dorsal root entry zone (DREZ), the interface between the dorsal root and the spinal cord. This lack of regeneration is due to the limited regenerative capacity of adult sensory axons and the growth-inhibitory environment at the DREZ, which is similar to that found in the glial scar after a central nervous system (CNS) injury. We hypothesized that transduction of adult DRG neurons using adeno-associated virus (AAV) to express a constitutively-active form of the GTPase Rheb (caRheb) will increase their intrinsic growth potential after a dorsal root crush. Additionally, we posited that if we combined that approach with digestion of upregulated chondroitin sulfate proteoglycans (CSPG) at the DREZ with chondroitinase ABC (ChABC), we would promote regeneration of sensory axons across the DREZ into the spinal cord. We first assessed if this strategy promotes neuritic growth in an in vitro model of the glial scar containing CSPG. ChABC allowed for some regeneration across the once potently inhibitory substrate. Combining ChABC treatment with expression of caRheb in DRG significantly improved this growth. We then determined if this combination strategy also enhanced regeneration through the DREZ after dorsal root crush in adult rats in vivo. After unilaterally crushing C4-T1 dorsal roots, we injected AAV5-caRheb or AAV5-GFP into the ipsilateral C5-C8 DRGs. ChABC or PBS was injected into the ipsilateral dorsal horn at C5-C8 to digest CSPG, for a total of four animal groups (caRheb + ChABC, caRheb + PBS, GFP + ChABC, GFP + PBS). Regeneration was rarely observed in PBS-treated animals, whereas short-distance regrowth across the DREZ was observed in ChABC-treated animals. No difference in axon number or length between the ChABC groups was observed, which may be related to intraganglionic inflammation induced by the
Expressing Constitutively Active Rheb in Adult Dorsal Root Ganglion Neurons Enhances the Integration of Sensory Axons that Regenerate Across a Chondroitinase-Treated Dorsal Root Entry Zone Following Dorsal Root Crush.
Wu, Di; Klaw, Michelle C; Kholodilov, Nikolai; Burke, Robert E; Detloff, Megan R; Côté, Marie-Pascale; Tom, Veronica J
While the peripheral branch of dorsal root ganglion neurons (DRG) can successfully regenerate after injury, lesioned central branch axons fail to regrow across the dorsal root entry zone (DREZ), the interface between the dorsal root and the spinal cord. This lack of regeneration is due to the limited regenerative capacity of adult sensory axons and the growth-inhibitory environment at the DREZ, which is similar to that found in the glial scar after a central nervous system (CNS) injury. We hypothesized that transduction of adult DRG neurons using adeno-associated virus (AAV) to express a constitutively-active form of the GTPase Rheb (caRheb) will increase their intrinsic growth potential after a dorsal root crush. Additionally, we posited that if we combined that approach with digestion of upregulated chondroitin sulfate proteoglycans (CSPG) at the DREZ with chondroitinase ABC (ChABC), we would promote regeneration of sensory axons across the DREZ into the spinal cord. We first assessed if this strategy promotes neuritic growth in an in vitro model of the glial scar containing CSPG. ChABC allowed for some regeneration across the once potently inhibitory substrate. Combining ChABC treatment with expression of caRheb in DRG significantly improved this growth. We then determined if this combination strategy also enhanced regeneration through the DREZ after dorsal root crush in adult rats in vivo. After unilaterally crushing C4-T1 dorsal roots, we injected AAV5-caRheb or AAV5-GFP into the ipsilateral C5-C8 DRGs. ChABC or PBS was injected into the ipsilateral dorsal horn at C5-C8 to digest CSPG, for a total of four animal groups (caRheb + ChABC, caRheb + PBS, GFP + ChABC, GFP + PBS). Regeneration was rarely observed in PBS-treated animals, whereas short-distance regrowth across the DREZ was observed in ChABC-treated animals. No difference in axon number or length between the ChABC groups was observed, which may be related to intraganglionic inflammation induced by the
Xu, Changshui; Xu, Wenyuan; Xu, Hong; Xiong, Wei; Gao, Yun; Li, Guilin; Liu, Shuangmei; Xie, Jinyan; Tu, Guihua; Peng, Haiying; Qiu, Shuyi; Liang, Shangdong
Tissue injury or inflammation of the nervous system may result in chronic neuropathic pain characterized by sensitivity to painful stimuli. P2X(3) receptors play a crucial role in facilitating pain transmission. Puerarin is an active compound of a traditional Chinese medicine Ge-gen, and Ge-gen soup has anti-inflammatory effects. The present research investigated the role of puerarin in the signalling of chronic neuropathic pain mediated by P2X(3) receptors of rat dorsal root ganglion neurons. Chronic constriction injury (CCI) rat model was adopted. Sprague-Dawley rats were randomly divided into blank control group (Ctrl), sham group (Sham), puerarin-treated control group (Ctrl+PUE), chronic constriction injury (CCI) group and puerarin-treated CCI group (CCI+PUE). Mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) were measured by the von-Frey test and the Hargreaves' test respectively. The stain values of P2X(3) protein and mRNA in L4/L5 dorsal root ganglion (DRG) were detected by immunohistochemistry, western blot and in situ hybridization. At day 4-7 after the operation of CCI rats, MWT and TWL in group CCI and CCI+PUE were lower than those in group Ctrl, Sham and Ctrl+PUE, while there was no difference among group Ctrl, Sham and Ctrl+PUE. At day 7-10 after operation, MWT and TWL in group CCI+PUE was higher than those in group CCI, but there was no significant difference between group CCI+PUE and group Ctrl (p>0.05). At day 14 after operation, the stain values of P2X(3) proteins and mRNAs in L4/L5 DRG of group CCI were higher than those in group Ctrl, Sham, Ctrl+PUE and CCI+PUE, while the stain values of P2X(3) proteins and mRNAs in group CCI+PUE were significantly decreased compared with those in group CCI. Therefore, puerarin may alleviate neuropathic pain mediated by P2X(3) receptors in dorsal root ganglion neurons.
Xiangming, Liu; Su, Chen; Shijin, Yin; Zhinan, Mei
Using whole-cell patch clamp technique on the membrane of freshly isolated dorsal root ganglion (DRG) neurons, the effects of dragon's blood resin and its important component loureirin B on tetrodotoxin-sensitive (TTX-S) voltage-gated sodium currents were observed. The results show that both blood resin and loureirin B could suppress TTX-S voltage-gated sodium currents in a dose-dependent way. The peak current amplitudes and the steady-state activation and inactivation curves are also made to shift by 0.05% blood resin and 0.2 mmol/L loureirin B. These results demonstrate that the effects of blood resin on TTX-S sodium current may contribute to loureirin B in blood resin. Perhaps the analgesic effect of blood resin is caused partly by loureirin B directly interfering with the nociceptive transmission of primary sensory neurons.
Deer, Timothy R.; Levy, Robert M.; Kramer, Jeffery; Poree, Lawrence; Amirdelfan, Kasra; Grigsby, Eric; Staats, Peter; Burton, Allen W.; Burgher, Abram H.; Obray, Jon; Scowcroft, James; Golovac, Stan; Kapural, Leonardo; Paicius, Richard; Kim, Christopher; Pope, Jason; Yearwood, Thomas; Samuel, Sam; McRoberts, W. Porter; Cassim, Hazmer; Netherton, Mark; Miller, Nathan; Schaufele, Michael; Tavel, Edward; Davis, Timothy; Davis, Kristina; Johnson, Linda; Mekhail, Nagy
Abstract Animal and human studies indicate that electrical stimulation of dorsal root ganglion (DRG) neurons may modulate neuropathic pain signals. ACCURATE, a pivotal, prospective, multicenter, randomized comparative effectiveness trial, was conducted in 152 subjects diagnosed with complex regional pain syndrome or causalgia in the lower extremities. Subjects received neurostimulation of the DRG or dorsal column (spinal cord stimulation, SCS). The primary end point was a composite of safety and efficacy at 3 months, and subjects were assessed through 12 months for long-term outcomes and adverse events. The predefined primary composite end point of treatment success was met for subjects with a permanent implant who reported 50% or greater decrease in visual analog scale score from preimplant baseline and who did not report any stimulation-related neurological deficits. No subjects reported stimulation-related neurological deficits. The percentage of subjects receiving ≥50% pain relief and treatment success was greater in the DRG arm (81.2%) than in the SCS arm (55.7%, P < 0.001) at 3 months. Device-related and serious adverse events were not different between the 2 groups. Dorsal root ganglion stimulation also demonstrated greater improvements in quality of life and psychological disposition. Finally, subjects using DRG stimulation reported less postural variation in paresthesia (P < 0.001) and reduced extraneous stimulation in nonpainful areas (P = 0.014), indicating DRG stimulation provided more targeted therapy to painful parts of the lower extremities. As the largest prospective, randomized comparative effectiveness trial to date, the results show that DRG stimulation provided a higher rate of treatment success with less postural variation in paresthesia intensity compared to SCS. PMID:28030470
Szallasi, A; Bíró, T; Szabó, T; Modarres, S; Petersen, M; Klusch, A; Blumberg, P M; Krause, J E; Sterner, O
1. A [3H]-resiniferatoxin (RTX) binding assay utilizing rat spinal cord membranes was employed to identify novel vanilloids in a collection of natural products of fungal origin. Of the five active compounds found (scutigeral, acetyl-scutigeral, ovinal, neogrifolin, and methyl-neogrifolin), scutigeral (Ki=19 microM), isolated from the edible mushroom Albatrellus ovinus, was selected for further characterization. 2. Scutigeral induced a dose-dependent 45Ca uptake by rat dorsal root ganglion neurons with an EC50 of 1.6 microM, which was fully inhibited by the competitive vanilloid receptor antagonist capsazepine (IC50=5.2 microM). 3. [3H]-RTX binding isotherms were shifted by scutigeral (10-80 microM) in a competitive manner. The Schild plot of the data had a slope of 0.8 and gave an apparent Kd estimate for scutigeral of 32 microM. 4. Although in the above assays scutigeral mimicked capsaicin, it was not pungent on the human tongue up to a dose of 100 nmol per tongue, nor did it provoke protective wiping movements in the rat (up to 100 microM) upon intraocular instillation. 5. In accord with being non-pungent, scutigeral (5 microM) did not elicit a measurable inward current in isolated rat dorsal root ganglion neurons under voltage-clamp conditions. It did, however, reduce the proportion of neurons (from 61 to 15%) that responded to a subsequent capsaicin (1 microM) challenge. In these neurons, scutigeral both delayed (from 27 to 72 s) and diminished (from 5.0 to 1.9 nA) the maximal current evoked by capsaicin. 6. In conclusion, scutigeral and its congeners form a new chemical class of vanilloids, the triprenyl phenols. Scutigeral promises to be a novel chemical lead for the development of orally active, non-pungent vanilloids.
Moen, Aurora; Haugen, Fred; Gjerstad, Johannes
Introduction. Lumbar radicular pain following intervertebral disc herniation may be associated with a local inflammatory response induced by nucleus pulposus (NP) cells. Methods. In anaesthetized Lewis rats, extracellular single unit recordings of wide dynamic range (WDR) neurons in the dorsal horn and qPCR were used to explore the effect of NP application onto the dorsal nerve roots (L3–L5). Results. A clear increase in C-fiber response was observed following NP conditioning. In the NP tissue, the expression of interleukin-1β (IL-1β), colony stimulating factor 1 (Csf1), fractalkine (CX3CL1), and the fractalkine receptor CX3CR1 was increased. Minocycline, an inhibitor of microglial activation, inhibited the increase in neuronal activity and attenuated the increase in IL-1β, Csf1, CX3L1, and CX3CR1 expression in NP tissue. In addition, the results demonstrated an increase in the expression of TNF, CX3CL1, and CX3CR1 in the dorsal root ganglions (DRGs). Conclusion. Hyperexcitability in the pain pathways and the local inflammation after disc herniation may involve upregulation of CX3CL1 signaling in both the NP and the DRG. PMID:28116212
Ugartechea-Chirino, Yamel; Swarup, Ranjan; Swarup, Kamal; Péret, Benjamin; Whitworth, Morag; Bennett, Malcolm; Bougourd, Sue
Background and Aims The root meristem of the Arabidopsis thaliana mature embryo is a highly organized structure in which individual cell shape and size must be regulated in co-ordination with the surrounding cells. The objective of this study was to determine the role of the AUX1 LAX family of auxin import carriers during the establishment of the embryonic root cell pattern. Methods The radicle apex of single and multiple aux1 lax mutant mature embryos was used to evaluate the effect of this gene family upon embryonic root organization and root cap size, cell number and cell size. Key Results It was demonstrated here that mutations within the AUX1 LAX family are associated with changes in cell pattern establishment in the embryonic quiescent centre and columella. aux1 lax mutants have a larger radicle root cap than the wild type and this is associated with a significant increase in the root-cap cell number, average cell size, or both. Extreme disorganization of the radicle apex was observed among quadruple aux1 lax1 lax2 lax3 mutant embryos, but not in single aux1 null or in lax1, lax2 and lax3 single mutants, indicating redundancy within the AUX1 LAX family. Conclusions It was determined that the AUX1 LAX family of auxin influx facilitators participates in the establishment of cell pattern within the apex of the embryonic root in a gene-redundant fashion. It was demonstrated that aux1 lax mutants are affected in cell proliferation and cell growth within the radicle tip. Thus AUX1 LAX auxin importers emerge as new players in morphogenetic processes involved in patterning during embryonic root formation. PMID:19952011
Ogawa, Nobuhiro; Kawai, Hiromichi; Terashima, Tomoya; Kojima, Hideto; Oka, Kazuhiro; Chan, Lawrence; Maegawa, Hiroshi
Neuropathic pain can be a debilitating condition. Many types of drugs that have been used to treat neuropathic pain have only limited efficacy. Recent studies indicate that pro-inflammatory mediators including tumor necrosis factor α (TNF-α) are involved in the pathogenesis of neuropathic pain. In the present study, we engineered a gene therapy strategy to relieve neuropathic pain by silencing TNF-α expression in the dorsal root ganglion (DRG) using lentiviral vectors expressing TNF short hairpin RNA1-4 (LV-TNF-shRNA1-4) in mice. First, based on its efficacy in silencing TNF-α in vitro, we selected shRNA3 to construct LV-TNF-shRNA3 for in vivo study. We used L5 spinal nerve transection (SNT) mice as a neuropathic pain model. These animals were found to display up-regulated mRNA expression of activating transcription factor 3 (ATF3) and neuropeptide Y (NPY), injury markers, and interleukin (IL)-6, an inflammatory cytokine in the ipsilateral L5 DRG. Injection of LV-TNF-shRNA3 onto the proximal transected site suppressed significantly the mRNA levels of ATF3, NPY and IL-6, reduced mechanical allodynia and neuronal cell death of DRG neurons. These results suggest that lentiviral-mediated silencing of TNF-α in DRG relieves neuropathic pain and reduces neuronal cell death, and may constitute a novel therapeutic option for neuropathic pain. PMID:24642694
Chung, Gehoon; Kim, Tae-Hyung; Shin, Hyewon; Chae, Eunhee; Yi, Hanju; Moon, Hongsik; Kim, Hyun Jin; Kim, Joong Soo; Jung, Sung Jun; Oh, Seog Bae
In this study, we determined mode of action of a novel carbamoyloxy arylalkanoyl arylpiperazine compound (SKL-NP) on hyperpolarization-activated cyclic nucleotide-gated (HCN) channel currents (I(h)) that plays important roles in neuropathic pain. In small or medium-sized dorsal root ganglion (DRG) neurons (<40 µm in diameter) exhibiting tonic firing and prominent I(h), SKL-NP inhibited I(h) and spike firings in a concentration dependent manner (IC(50)=7.85 µM). SKL-NP-induced inhibition of I(h) was blocked by pretreatment of pertussis toxin (PTX) and N-ethylmaleimide (NEM) as well as 8-Br-cAMP, a membrane permeable cAMP analogue. These results suggest that SKL-NP modulates I(h) in indirect manner by the activation of a Gi-protein coupled receptor that decreases intracellular cAMP concentration. Taken together, SKL-NP has the inhibitory effect on HCN channel currents (I(h)) in DRG neurons of rats.
Liu, Ning; Varma, Sushama; Shooter, Eric M; Tolwani, Ravi J
The Trembler-J (TrJ) mouse, containing a point mutation in the peripheral myelin protein 22 gene, is characterized by severe hypomyelination and is a representative model of Charcot-Marie-Tooth 1A disease/Dejerine-Sottas Syndrome. Previous studies have shown that protein kinase inhibitor K252a enhances wild-type Schwann cell myelination in culture. We used a dorsal root ganglion (DRG) explant culture system from the heterozygous TrJ/+ mouse to investigate if myelination could be enhanced by K252a. The TrJ/+ DRG explant cultures replicated some important features of the TrJ/+ mouse, showing reduced myelin protein accumulation, thinner myelin sheaths, and shortened myelin internodes. K252a increased myelin protein accumulation and myelin sheath thickness but did not substantially increase myelin internode length. Furthermore, the TrJ/+ DRG explant culture and sciatic nerves continued to respond to K252a during the stage when myelination is complete in the wild type. A general tyrosine kinase inhibitor, genistein, but not inhibitors of serine/threonine protein kinase inhibitors, had a similar effect to K252a. K252a is therefore able to partially overcome hypomyelination by enhancing mutant Schwann cell myelin formation in the TrJ/+ mouse.
Background Cerebral palsy (CP) may cause severe spasticity, requiring neurosurgical procedures. The most common neurosurgical procedures are continuous infusion of intrathecal baclofen and selective dorsal rhizotomy. Both are invasive and complex procedures. We hypothesized that a percutaneous radiofrequency lesion of the dorsal root ganglion (RF-DRG) could be a simple and safe alternative treatment. We undertook a pilot study to test this hypothesis. Methods We performed an RF-DRG procedure in 17 consecutive CP patients with severe hip flexor/adductor spasms accompanied by pain or care-giving difficulties. Six children were systematically evaluated at baseline, and 1 month and 6 months after treatment by means of the Modified Ashworth Scale (MAS), Gross Motor Function Measure (GMFM) and a self-made caregiver's questionnaire. Eleven subsequent children were evaluated using a Visual Analogue Scale (VAS) for spasticity, pain and ease of care. Results A total of 19 RF-DRG treatments were performed in 17 patients. We found a small improvement in muscle tone measured by MAS, but no effect on the GMFM scale. Despite this, the caregivers of these six treated children unanimously stated that the quality of life of their children had indeed improved after the RF-DRG. In the subsequent 11 children we found improvements in all VAS scores, in a range comparable to the conventional treatment options. Conclusion RF-DRG is a promising new treatment option for severe spasticity in CP patients, and its definitive effectiveness remains to be defined in a randomised controlled trial. PMID:20569438
Demirdaş, Arif; Nazıroğlu, Mustafa; Övey, Ishak Suat
Calcium ions (Ca(2+)) are important second messengers in neurons. Ketamine (KETAM) is an anesthetic and analgesic, with psychotomimetic effects and abuse potential. KETAM modulates the entry of Ca(2+) in neurons through glutamate receptors, but its effect on transient receptor potential melastatin 2 (TRPM2) and transient receptor potential vanilloid 1 (TRPV1) channels has not been clarified. This study investigated the short-term effects of KETAM on oxidative stress and TRPM2 and TRPV1 channel gating in hippocampal and dorsal root ganglion (DRG) neurons of rats. Freshly isolated hippocampal and DRG neurons were incubated for 24 h with KETAM (0.3 mM). The TRPM2 channel antagonist, N-(p-amylcinnamoyl)anthranilic acid (ACA), inhibited cumene hydroperoxide and ADP-ribose-induced TRPM2 currents in the neurons, and capsazepine (CPZ) inhibited capsaicin-induced TRPV1 currents. The TRPM2 and TRPV1 channel current densities and intracellular free calcium ion concentration of the neurons were lower in the neurons exposed to ACA and CPZ compared to the control neurons, respectively. However, the values were not further decreased by the KETAM + CPZ and KETAM + ACA treatments. KETAM decreased lipid peroxidation levels in the neurons but increased glutathione peroxidase activity. In conclusion, short-term KETAM treatment decreased oxidative stress levels but did not seem to influence TRPM2- and TRPV1-mediated Ca(2+) entry.
Jin, Yunju; Kim, Jun
Recent studies have demonstrated that nitric oxide (NO) activates transient receptor potential vanilloid subtype 1 (TRPV1) via S-nitrosylation of the channel protein. NO also modulates various cellular functions via activation of the soluble guanylyl cyclase (sGC)/protein kinase G (PKG) pathway and the direct modification of proteins. Thus, in the present study, we investigated whether NO could indirectly modulate the activity of TRPV1 via a cGMP/PKG-dependent pathway in cultured rat dorsal root ganglion (DRG) neurons. NO donors, sodium nitroprusside (SNP) and S-nitro-N-acetylpenicillamine (SNAP), decreased capsaicin-evoked currents (Icap). NO scavengers, hemoglobin and 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (CPTIO), prevented the inhibitory effect of SNP on Icap. Membrane-permeable cGMP analogs, 8-bromoguanosine 3', 5'-cyclic monophosphate (8bromo-cGMP) and 8-(4chlorophenylthio)-guanosine 3',5'-cyclic monophosphate (8-pCPT-cGMP), and the guanylyl cyclase stimulator YC-1 mimicked the effect of SNP on Icap. The PKG inhibitor KT5823 prevented the inhibition of Icap by SNP. These results suggest that NO can downregulate the function of TRPV1 through activation of the cGMP/PKG pathway in peripheral sensory neurons. PMID:22802704
Pan, Hai-Li; Zhang, Yu-Qiu; Zhao, Zhi-Qi
It has been demonstrated that lysophosphatidic acid (LPA) released from injury tissue and transient receptor potential vanilloid 1 (TRPV1) receptor are implicated in the induction of chronic pain. In the present study we examined whether an interaction between LPA receptor LPA(1) and TRPV1 in dorsal root ganglion (DRG) neurons contributes to the development of bone cancer pain. Bone cancer was established by injection of mammary gland carcinoma cells into the rat tibia. Following the development of bone cancer pain, the TRPV1 expression and capsaicin-evoked currents were up-regulated in rat DRG neurons at L(4-6) segments. Immunohistochemistry staining revealed a high co-localization of LPA(1) with TRPV1 in DRG neurons. In isolated DRG neurons, whole-cell patch recording showed that capsaicin-induced currents were potentiated by LPA in a dose-dependent manner. The potentiation was blocked by either LPA(1) antagonist, protein kinase C (PKC) inhibitor or PKCε inhibitor, but not by protein kinase A (PKA) inhibitor or Rho inhibitor. In the behavioral tests, both mechanical allodynia and thermal hyperalgesia in bone cancer rats were attenuated by LPA(1) antagonist. LPA potentiates TRPV1 current via a PKCε-dependent pathway in DRG neurons of rats with bone cancer, which may be a novel peripheral mechanism underlying the induction of bone cancer pain.
Kolarcik, Christi L.; Bourbeau, Dennis; Azemi, Erdrin; Rost, Erika; Zhang, Ling; Lagenaur, Carl F.; Weber, Douglas J.; Cui, X. Tracy
The spinal cord (SC) and dorsal root ganglion (DRG) are target implantation regions for neural prosthetics, but the tissue-electrode interface in these regions is not well-studied. To improve our understanding of these locations, we characterized the tissue reactions around implanted electrodes. L1, an adhesion molecule shown to maintain neuronal density and reduce gliosis in brain tissue, was then evaluated in SC and DRG implants. Following L1 immobilization onto neural electrodes, the bioactivities of the coatings were verified in vitro using neuron, astrocyte and microglia cultures. Non-modified and L1-coated electrodes were implanted into adult rats for 1 or 4 weeks. Hematoxylin and eosin staining along with cell-type specific antibodies were used to characterize the tissue response. In the SC and DRG, cells aggregated at the electrode-tissue interface. Microglia staining was more intense around the implant site and decreased with distance from the interface. Neurofilament staining in both locations was decreased or absent around the implant when compared to surrounding tissue. With L1, neurofilament staining was significantly increased while neuronal cell death decreased. Our results indicate that L1-modified electrodes may result in an improved chronic neural interface and will be evaluated in recording and stimulation studies. PMID:22750248
Qiu, Fang; Qiu, Chun-Yu; Liu, Yu-Qiang; Wu, Dan; Li, Jia-Da; Hu, Wang-Ping
Acid-sensing ion channels (ASICs), as key sensors for extracellular protons, are expressed in nociceptive sensory neurons and contribute to signalling pain caused by tissue acidosis. ASICs are also the subject of various factors. Here, we further provide evidence that the activity of ASICs is potentiated by the activation of 5-HT₂ receptors in rat dorsal root ganglion neurons. A specific 5-HT₂ receptor agonist, α-methyl-5-HT, dose-dependently enhanced proton-gated currents with an EC₅₀ of 0.13 ± 0.07 nM. The α-methyl-5-HT enhancing effect on proton-gated currents was blocked by cyproheptadine, a 5-HT₂ receptor antagonist, and removed by intracellular dialysis of either GDP-β-S or protein kinase C inhibitor GF109203X. Moreover, α-methyl-5-HT altered acid-evoked membrane excitability of rat DRG neurons and caused a significant increase in the amplitude of the depolarization and the number of spikes induced by acid stimuli. Finally, α-methyl-5-HT increased nociceptive responses to injection of acetic acid in rats. These results suggest that α-methyl-5-HT up-regulates the activity of ASICs via 5-HT₂ receptor and protein kinase C dependent signal pathways in rat primary sensory neurons and this potentiation contributed to acid- mediated pain in tissue injury and inflammation.
Quan, Xin; Guo, Kai; Wang, Yuqing; Huang, Liangliang; Chen, Beiyu; Ye, Zhengxu; Luo, Zhuojing
In a primary spinal cord injury, the amount of mechanical compression insult that the neurons experience is one of the most critical factors in determining the extent of the injury. The ultrastructural changes that neurons undergo when subjected to mechanical compression are largely unknown. In the present study, using a compression-driven instrument that can simulate mechanical compression insult, we applied mechanical compression stimulation at 0.3, 0.5, and 0.7 MPa to dorsal root ganglion (DRG) neurons for 10 min. Combined with atomic force microscopy, we investigated nanoscale changes in the membrane-skeleton, cytoskeleton alterations, and apoptosis induced by mechanical compression injury. The results indicated that mechanical compression injury leads to rearrangement of the membrane-skeleton compared with the control group. In addition, mechanical compression stimulation induced apoptosis and necrosis and also changed the distribution of the cytoskeleton in DRG neurons. Thus, the membrane-skeleton may play an important role in the response to mechanical insults in DRG neurons. Moreover, sudden insults caused by high mechanical compression, which is most likely conducted by the membrane-skeleton, may induce necrosis, apoptosis, and cytoskeletal alterations.
Chen, Su; Liu, Xiangming
Using patch clamp technique the effects of dragon's blood and its component loureirin B on tetrodotoxin-sensitive sodium channel currents in dorsal root ganglion cells were observed. The experimental data were simulated with Hodgkin-Huxley model and the corresponding parameters were estimated. In addition, computer-simulated neuron action potentials in the absence and presence of drugs were produced using Hodgkin-Huxley model. The results show that the conductance of tetrodotoxin-sensitive sodium channel was fitted with m3h model well, the half-activated potentials of the sodium channel in the presence of drugs were shifted to the depolarizing direction and the threshold intensity of the cells in the presence of drugs was increased. These results demonstrate that dragon's blood and loureirin B did not resemble the tetrodotoxin which inhibited tetrodotoxin-sensitive sodium channel currents completely. Perhaps the analgesic effects of dragon's blood were partly caused by loureirin B affecting the activation, blocking the action potential generation and interfering with the transmission of painful signals into the central nervous system.
Hu, Zhaotun; Zhou, Xi; Chen, Jia; Tang, Cheng; Xiao, Zhen; Ying, Dazhong; Liu, Zhonghua; Liang, Songping
Selenocosmia jiafu is a medium-sized theraphosid spider and an attractive source of venom, because it can be bred in captivity and it produces large amounts of venom. We performed reversed-phase high-performance liquid chromatography (RP-HPLC) and matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) analyses and showed that S. jiafu venom contains hundreds of peptides with a predominant mass of 3000–4500 Da. Patch clamp analyses indicated that the venom could inhibit voltage-gated Na+, K+ and Ca2+ channels in rat dorsal root ganglion (DRG) neurons. The venom exhibited inhibitory effects on tetrodotoxin-resistant (TTX-R) Na+ currents and T-type Ca2+ currents, suggesting the presence of antagonists to both channel types and providing a valuable tool for the investigation of these channels and for drug development. Intra-abdominal injection of the venom had severe toxic effects on cockroaches and caused death at higher concentrations. The LD50 was 84.24 μg/g of body weight in the cockroach. However, no visible symptoms or behavioral changes were detected after intraperitoneal injection of the venom into mice even at doses up to 10 mg/kg body weight. Our results provide a basis for further case-by-case investigations of peptide toxins from this venom. PMID:24603666
Yan, Jun; Hu, Shufen; Zou, Kang; Xu, Min; Wang, Qianliang; Miao, Xiuhua; Yu, Shan Ping; Xu, Guang-Yin
The pathogenesis of pain in lumbar disc herniation (LDH) remains poorly understood. We have recently demonstrated that voltage-gated sodium channels (VGSCs) in dorsal root ganglion (DRG) neurons were sensitized in a rat model of LDH. However, the detailed molecular mechanism for sensitization of VGSCs remains largely unknown. This study was designed to examine roles of the endogenous hydrogen sulfide synthesizing enzyme cystathionine β-synthetase (CBS) in sensitization of VGSCs in a previously validated rat model of LDH. Here we showed that inhibition of CBS activity by O-(Carboxymethyl) hydroxylamine hemihydrochloride (AOAA) significantly attenuated pain hypersensitivity in LDH rats. Administration of AOAA also reduced neuronal hyperexcitability, suppressed the sodium current density, and right-shifted the V1/2 of the inactivation curve, of hindpaw innervating DRG neurons, which is retrogradely labeled by DiI. In vitro incubation of AOAA did not alter the excitability of acutely isolated DRG neurons. Furthermore, CBS was colocalized with NaV1.7 and NaV1.8 in hindpaw-innervating DRG neurons. Treatment of AOAA markedly suppressed expression of NaV1.7 and NaV1.8 in DRGs of LDH rats. These data suggest that targeting the CBS-H2S signaling at the DRG level might represent a novel therapeutic strategy for chronic pain relief in patients with LDH. PMID:27905525
van Velsen, Valery; van Helmond, Noud; Chapman, Kenneth B
Chronic neuropathic pain is often refractory to conventional medical treatments and leads to significant disability and socio-economic burden. Dorsal root ganglion (DRG) stimulation has recently emerged as a treatment for persistent neuropathic pain, but creating a strain relief loop on the S1 level has thus far been a challenging technical component of DRG lead placement. We describe a refined technique for strain relief loop formation on the S1 level using a transforaminal approach that we employed in a 45-year old patient with intractable foot pain. We successfully placed a strain relief loop in the sacral space in a predictable and easily reproducible manner using a transforaminal anchorless approach. The patient experienced a decrease in visual analogue pain score (85%), and improvement in function during the trial period, and proceeded with permanent implantation. The described sacral transforaminal strain relief loop formation technique appears to be a more reliable and predictable technique of DRG lead placement in the sacrum than those previously documented. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
Nazıroğlu, M; Çelik, Ö; Özgül, C; Çiğ, B; Doğan, S; Bal, R; Gümral, N; Rodríguez, A B; Pariente, J A
We aimed to investigate the protective effects of melatonin and 2.45 GHz electromagnetic radiation (EMR) on brain and dorsal root ganglion (DRG) neuron antioxidant redox system, Ca(2+) influx, cell viability and electroencephalography (EEG) records in the rat. Thirty two rats were equally divided into four different groups namely group A1: Cage control, group A2: Sham control, group B: 2.45 GHz EMR, group C: 2.45 GHz EMR+melatonin. Groups B and C were exposed to 2.45 GHz EMR during 60 min/day for 30 days. End of the experiments, EEG records and the brain cortex and DRG samples were taken. Lipid peroxidation (LP), cell viability and cytosolic Ca(2+) values in DRG neurons were higher in group B than in groups A1 and A2 although their concentrations were increased by melatonin, 2-aminoethyldiphenyl borinate (2-APB), diltiazem and verapamil supplementation. Spike numbers of EEG records in group C were lower than in group B. Brain cortex vitamin E concentration was higher in group C than in group B. In conclusion, Melatonin supplementation in DRG neurons and brain seems to have protective effects on the 2.45 GHz-induced increase Ca(2+) influx, EEG records and cell viability of the hormone through TRPM2 and voltage gated Ca(2+) channels. Copyright © 2011 Elsevier Inc. All rights reserved.
Zhang, Yan; Jia, Yan-Yan; Guo, Jin-Lei; Liu, Pei-Qing; Jiang, Jian-Min
The (−)-gallocatechin-3-gallate (GCG) concentration in some tea beverages can account for as much as 50% of the total catechins. It has been shown that catechins have analgesic properties. Voltage-gated sodium channels (Nav) mediate neuronal action potentials. Tetrodotoxin inhibits all Nav isoforms, but Nav1.8 and Nav1.9 are relatively tetrodotoxin-resistant compared to other isoforms and functionally linked to nociception. In this study, the effects of GCG on tetrodotoxin-resistant Na+ currents were investigated in rat primary cultures of dorsal root ganglion neurons via the whole-cell patch-clamp technique. We found that 1 μM GCG reduced the amplitudes of peak current density of tetrodotoxin-resistant Na+ currents significantly. Furthermore, the inhibition was accompanied by a depolarizing shift of the activation voltage and a hyperpolarizing shift of steady-state inactivation voltage. The percentage block of GCG (1 μM) on tetrodotoxin-resistant Na+ current was 45.1% ± 1.1% in 10 min. In addition, GCG did not produce frequency-dependent block of tetrodotoxin-resistant Na+ currents at stimulation frequencies of 1 Hz, 2 Hz and 5 Hz. On the basis of these findings, we propose that GCG may be a potential analgesic agent. PMID:23652835
Ishihara, A.; Ohira, Y.; Roy, R. R.; Nagaoka, S.; Sekiguchi, C.; Hinds, W. E.; Edgerton, V. R.
The cross-sectional areas and succinate dehydrogenase activities of L5 dorsal root ganglion neurons in rats were determined after 14 days of spaceflight and after nine days of recovery. The mean and distribution of the cross-sectional areas were similar to age-matched, ground-based controls for both the spaceflight and for the spaceflight plus recovery groups. The mean succinate dehydrogenase activity was significantly lower in spaceflight compared to aged-matched control rats, whereas the mean succinate dehydrogenase activity was similar in age-matched control and spaceflight plus recovery rats. The mean succinate dehydrogenase activity of neurons with cross-sectional areas between 1000 and 2000 microns2 was lower (between 7 and 10%) in both the spaceflight and the spaceflight plus recovery groups compared to the appropriate control groups. The reduction in the oxidative capacity of a subpopulation of sensory neurons having relatively large cross-sectional areas immediately following spaceflight and the sustained depression for nine days after returning to 1 g suggest that the 0 g environment induced significant alterations in proprioceptive function.
Meng, Bo; Shen, Lu-Lu; Shi, Xiao-Ting; Gong, Yong-Sheng; Fan, Xiao-Fang; Li, Jun; Cao, Hong
Type 2 diabetic mellitus (T2DM) has reached pandemic status and shows no signs of abatement. Diabetic neuropathic pain (DNP) is generally considered to be one of the most common complications of T2DM, which is also recognized as one of the most difficult types of pain to treat. As one kind of peripheral neuropathic pain, DNP manifests typical chronic neuralgia symptoms, including hyperalgesia, allodynia, autotomy, and so on. The injured dorsal root ganglion (DRG) is considered as the first stage of the sensory pathway impairment, whose neurons display increased frequency of action potential generation and increased spontaneous activities. These are mainly due to the changed properties of voltage-gated sodium channels (VGSCs) and the increased sodium currents, especially TTX-R sodium currents. Curcumin, one of the most important phytochemicals from turmeric, has been demonstrated to effectively prevent and/or ameliorate diabetic mellitus and its complications including DNP. The present study demonstrates that the TTX-R sodium currents of small-sized DRG neurons isolated from DNP rats are significantly increased. Such abnormality can be efficaciously ameliorated by curcumin.
Sharp, Andrew A.; Fedorovich, Yuri
Our understanding of the role of somatosensory feedback in regulating motility during chicken embryogenesis and fetal development in general has been hampered by the lack of an approach to selectively alter specific sensory modalities. In adult mammals, pyridoxine overdose has been shown to cause a peripheral sensory neuropathy characterized by a loss of both muscle and cutaneous afferents, but predominated by a loss of proprioception. We have begun to explore the sensitivity of the nervous system in chicken embryos to the application of pyridoxine on embryonic days 7 and 8, after sensory neurons in the lumbosacral region become post-mitotic. Upon examination of the spinal cord, DRG and peripheral nerves, we find that pyridoxine causes a loss of TrkC-positive neurons, a decrease in the diameter of the muscle innervating nerve tibialis, and a reduction in the number of large diameter axons in this nerve. However, we found no change in the number of Substance P or CGRP-positive neurons, the number of motor neurons or the diameter or axonal composition of the femoral cutaneous nerve. Therefore, pyridoxine causes a peripheral sensory neuropathy in embryonic chickens largely consistent with its effects in adult mammals. However, the lesion may be more restricted to proprioception in the chicken embryo. Therefore, pyridoxine lesion induced during embryogenesis in the chicken embryo can be used to asses how the loss of sensation, largely proprioception, alters spontaneous embryonic motility and subsequent motor development. PMID:25592428
Ma, Zhong; Hasenstein, Karl H.
Vertical orientation of emerging roots typically is the first response of plants to gravity. Although root gravitropism has been studied extensively, no conclusive data on the onset of gravisensing exist. We determined the inception of gravisensitivity in flax (Linum usitatissimum) roots by clinorotating germinating seeds after various periods of static orientation (gravistimulation) of imbibed seeds. Gravitropic competency was established about 8 h after imbibition, 11 h prior to germination. The time was determined based on 50% of the newly emerged roots curving in the direction of the gravity vector during static imbibition, despite subsequent clinorotation. The threshold value was affected by the orientation of the seeds. Upward orientation of the micropyle/radicle reduced the number of graviresponding roots to about one-half. Prolonged clinorotation weakened the graviresponse. Gravisensing was accompanied by the development of amyloplasts, but the actin cytoskeleton was not involved because imbibition in Latrunculin B did not affect the onset of gravisensitivity or germination, and the development of F-actin in untreated controls was observed only after the onset of gravisensitivity. PMID:16377740
Boateng, EK; Novejarque, A; Pheby, T; Rice, ASC; Huang, W
Background Heterogeneity is increasingly recognized in clinical presentation of neuropathic pain (NP), but less often recognized in animal models. Neurochemical dysregulation in rodent dorsal root ganglia (DRG) is associated with peripheral nerve trauma, but poorly studied in non-traumatic NP conditions. Methods This study aimed to investigate the temporal expressions of activating transcription factor-3 (ATF-3), growth-associated protein-43 (GAP-43), neuropeptide Y (NPY) and galanin in traumatic and non-traumatic rat models of neuropathies associated with NP. Expressions of these markers were examined in the DRG at different time points following tibial nerve transection (TNT) injury and antiretroviral drug stavudine (d4T) administration using immunohistochemistry. The development of sensory gain following these insults was assessed by measuring limb withdrawal to a punctate mechanical stimulus. Results Both TNT-injured and d4T-treated rats developed hindpaw mechanical hypersensitivity. Robust expressions of ATF-3, GAP-43, NPY and galanin in both small- and large-sized L5 DRG neurons were observed in the DRG from TNT-injured rats. In contrast, d4T-treated rats did not exhibit any significant neurochemical changes in the DRG. Conclusions Taken together, the results suggest that ATF-3, GAP-43, NPY and galanin are likely indicators of nerve trauma-associated processes and not generic markers for NP. These experiments also demonstrate distinct expression patterns of neurochemical markers in the DRG and emphasize the mechanistic difference between nerve trauma and antiretroviral drug-associated NP. PMID:25070481
Desarmenien, M.; Feltz, P.; Headley, P. M.; Santangelo, F.
1 In anticipation that centrally active gamma-aminobutyric acid (GABA)-mimetic drugs may be clinically useful, derivatives of GABA with an imine link (Schiff base) to a lipophilic carrier have recently been prepared. The present paper concerns the actions of [alpha(4-chlorophenyl)5-fluoro, 2-hydroxy benzilidene-amino]-4-butanoate Na+, SL 75 102. 2 To test one aspect of the GABA-mimetic properties of SL 75 102, this compound was compared with GABA for activity on intracellularly-recorded neurones in rat dorsal root ganglia in vitro. On these neurones GABA, administered either by microiontophoresis or direct into the superfusion medium, causes a depolarization, due to an increased chloride conductance, followed by a period of desensitization. 3 The actions of Sl 75 102 were in nearly all respects identical to those of GABA; parameters examined were the effects on membrane potential and input conductance, desensitization, dose-response characteristics and sensitivity to the GABA antagonists, bicuculline and picrotoxin. 4 SL 75 102 was less potent than GABA (mean relative potency 0.03:1). 5 SL 75 102 therefore appears to be a weak agonist at GABA receptors of these neurones. PMID:7214101
Ohshiro, Hiroyuki; Ogawa, Shinji; Shinjo, Katsuhiro
Sensory information is conveyed to the central nervous system by primary afferent neurons within dorsal root ganglia (DRG), which synapse onto neurons of the dorsal horn of the spinal cord. This synaptic connection is central to the processing of both sensory and pain stimuli. Here, we describe a model system to monitor synaptic transmission between DRG neurons and dorsal horn neurons that is compatible with high-throughput screening. This co-culture preparation comprises DRG and dorsal horn neurons and utilizes Ca(2+) imaging with the indicator dye Fura-2 to visualize synaptic transmission. Addition of capsaicin to co-cultures stimulated DRG neurons and led to activation of dorsal horn neurons as well as increased intracellular Ca(2+) concentrations. This effect was dose-dependent and absent when DRG neurons were omitted from the culture. NMDA receptors are a critical component of synapses between DRG and dorsal horn neurons as MK-801, a use-dependent non-competitive antagonist, prevented activation of dorsal horn neurons following capsaicin treatment. This model system allows for rapid and efficient analysis of noxious stimulus-evoked Ca(2+) signal transmission and provides a new approach both for investigating synaptic transmission in the spinal cord and for screening potential analgesic compounds.
Su, Jie; Gao, Tianle; Shi, Tiejun; Xiang, Qiong; Xu, Xiaojun; Wiesenfeld-Hallin, Zsuzsanna; Hökfelt, Tomas; Svensson, Camilla I
The mechanisms underlying rheumatoid arthritis (RA)-induced pain are still not fully elucidated, and accumulating data indicate that peripheral inflammation is not the only factor driving pain in these patients. The focus of our work is to investigate the molecular basis for long-term alterations in nociceptive pathways induced by polyarthritis using the collagen antibody-induced arthritis (CAIA) mouse model. In this model, mechanical hypersensitivity outlasts the joint inflammation by weeks. Here we examined expression levels of neuropeptides, ion channels, and nerve injury markers associated with neuropathic and/or inflammatory pain in dorsal root ganglia (DRGs) and spinal cord both during the peak of inflammation (day 15) and when the inflammation has resolved but the hypersensitivity persists (days 45-47). No apparent differences were observed in substance P, calcitonin gene-related peptide, or neuropeptide Y protein expression in DRGs and spinal cord of CAIA mice. However, the neuropeptide galanin, the ATP-gated ion channel P2X3, and calcium channel subunit α2δ1 were significantly increased in the CAIA DRGs as compared to controls, both 15 and 47 days after induction of arthritis. On day 15 there was an increase in expression of two factors associated with nerve injury and cell stress, activating transcription factor 3 and growth-associated protein 43 in DRGs, whereby the latter was still dramatically upregulated after 47 days. In conclusion, this study suggests that long-term joint inflammation has an impact on DRG neurons that resembles both inflammation and nerve injury-induced pain states. Thus, antibody-driven inflammation generates a pain state with a unique neurochemical profile. © 2015 Wiley Periodicals, Inc.
Bossowska, A; Majewski, M
Conantokin G (CTG), isolated from the venom of the marine cone snail Conus geographus, is an antagonist of N-methyl-d-aspartate receptors (NMDARs), the activation of which, especially those located on the central afferent terminals and dorsal horn neurons, leads to hypersensitivity and pain. Thus, CTG blocking of NMDARs, has an antinociceptive effect, particularly in the case of neurogenic pain treatment. As many urinary bladder disorders are caused by hyperactivity of sensory bladder innervation, it seems useful to estimate the influence of CTG on the plasticity of sensory neurons supplying the organ. Retrograde tracer Fast Blue (FB) was injected into the urinary bladder wall of six juvenile female pigs. Three weeks later, intramural bladder injections of CTG (120 microg per animal) were carried out in all animals. After a week, dorsal root ganglia of interest were harvested from all animals and neurochemical characterization of FB+ neurons was performed using a routine double-immunofluorescence labeling technique on 10-microm-thick cryostat sections. CTG injections led to a significant decrease in the number of FB+ neurons containing substance P (SP), pituitary adenylate cyclase activating polypeptide (PACAP), somatostatin (SOM), calbindin (CB) and nitric oxide synthase (NOS) when compared with healthy animals (20% vs. 45%, 13% vs. 26%, 1.3% vs. 3%, 1.2 vs. 4% and 0.9% vs. 6% respectively) and to an increase in the number of cells immunolabelled for galanin (GAL, 39% vs. 6.5%). These data demonstrated that CTG changed the chemical coding of bladder sensory neurons, thus indicating that CTG could eventually be used in the therapy of selected neurogenic bladder illnesses.
Takahashi, Yuzuru; Ohtori, Seiji; Takahashi, Kazuhisa
Referred pain due to lumbar disc disorders can be analyzed using the stereoscopic structure of the peripheral sensory nervous system. The rostrocaudal structure has been clarified. The dorsoventral structure of the lumbar spine would be useful for mapping areas of pain perception in spinal disorders. The neurotracer 1,1-dioctadecyl-3,3,3,3-tetramethylindocarbocyanine perchlorate (DiI) was applied to the lateral portion of the L5/6 intervertebral disc in rats to examine the dorsoventral organization of the sensory nervous system in the lumbar spine and related tissues. Fluorogold (FG) was applied to reference sites located at the spinous process of the L5 vertebra, the L5/6 facet joint, the psoas muscle at the L5 level, or the rectus abdominis muscle at the pubic symphysis. FG was also applied to the lateral portion of the disc (DiI application site) at L5 or at the L5 level as controls for the double labeling. Labeled neurons were counted in dorsal root ganglia (DRGs) from L1 through L4. The proportion of neurons double-labeled with DiI and FG in the total number of DiI-labeled and FG-labeled neurons was 32.9% in the control group; 0% in the spinous process, 0.6% in the facet joint, 2.3% in the psoas muscle, and 0.1% in the rectus abdominis muscle. DRG neurons with dichotomizing afferent fibers were most prevalent (2.3%) between the lateral disc and the psoas muscle at the groin; they were rare or absent between the disc and other reference sites. Dorsoventral organization of the primary sensory system in the lumbar body trunk was suggested from the proportion of DRG neurons with dichotomizing afferent fibers innervating the lumbar disc and other tissues. The present findings provide a pathomechanism of groin referred pain in lumbar disc disorders.
Maratou, Klio; Wallace, Victoria C J; Hasnie, Fauzia S; Okuse, Kenji; Hosseini, Ramine; Jina, Nipurna; Blackbeard, Julie; Pheby, Timothy; Orengo, Christine; Dickenson, Anthony H; McMahon, Stephen B; Rice, Andrew S C
To elucidate the mechanisms underlying peripheral neuropathic pain in the context of HIV infection and antiretroviral therapy, we measured gene expression in dorsal root ganglia (DRG) of rats subjected to systemic treatment with the anti-retroviral agent, ddC (Zalcitabine) and concomitant delivery of HIV-gp120 to the rat sciatic nerve. L4 and L5 DRGs were collected at day 14 (time of peak behavioural change) and changes in gene expression were measured using Affymetrix whole genome rat arrays. Conventional analysis of this data set and Gene Set Enrichment Analysis (GSEA) was performed to discover biological processes altered in this model. Transcripts associated with G protein coupled receptor signalling and cell adhesion were enriched in the treated animals, while ribosomal proteins and proteasome pathways were associated with gene down-regulation. To identify genes that are directly relevant to neuropathic mechanical hypersensitivity, as opposed to epiphenomena associated with other aspects of the response to a sciatic nerve lesion, we compared the gp120+ddC-evoked gene expression with that observed in a model of traumatic neuropathic pain (L5 spinal nerve transection), where hypersensitivity to a static mechanical stimulus is also observed. We identified 39 genes/expressed sequence tags that are differentially expressed in the same direction in both models. Most of these have not previously been implicated in mechanical hypersensitivity and may represent novel targets for therapeutic intervention. As an external control, the RNA expression of three genes was examined by RT-PCR, while the protein levels of two were studied using western blot analysis.
Maratou, Klio; Wallace, Victoria C.J.; Hasnie, Fauzia S.; Okuse, Kenji; Hosseini, Ramine; Jina, Nipurna; Blackbeard, Julie; Pheby, Timothy; Orengo, Christine; Dickenson, Anthony H.; McMahon, Stephen B.; Rice, Andrew S.C.
To elucidate the mechanisms underlying peripheral neuropathic pain in the context of HIV infection and antiretroviral therapy, we measured gene expression in dorsal root ganglia (DRG) of rats subjected to systemic treatment with the anti-retroviral agent, ddC (Zalcitabine) and concomitant delivery of HIV-gp120 to the rat sciatic nerve. L4 and L5 DRGs were collected at day 14 (time of peak behavioural change) and changes in gene expression were measured using Affymetrix whole genome rat arrays. Conventional analysis of this data set and Gene Set Enrichment Analysis (GSEA) was performed to discover biological processes altered in this model. Transcripts associated with G protein coupled receptor signalling and cell adhesion were enriched in the treated animals, while ribosomal proteins and proteasome pathways were associated with gene down-regulation. To identify genes that are directly relevant to neuropathic mechanical hypersensitivity, as opposed to epiphenomena associated with other aspects of the response to a sciatic nerve lesion, we compared the gp120 + ddC-evoked gene expression with that observed in a model of traumatic neuropathic pain (L5 spinal nerve transection), where hypersensitivity to a static mechanical stimulus is also observed. We identified 39 genes/expressed sequence tags that are differentially expressed in the same direction in both models. Most of these have not previously been implicated in mechanical hypersensitivity and may represent novel targets for therapeutic intervention. As an external control, the RNA expression of three genes was examined by RT-PCR, while the protein levels of two were studied using western blot analysis. PMID:18606552
Sørensen, Bodil; Tandrup, Trine; Koltzenburg, Martin; Jakobsen, Johannes
The role of the p75 neurotrophin receptor for neuronal survival after nerve crush was studied in L5 dorsal root ganglia (DRG) of knockout mice and controls with assumption-free stereological methods. Numbers of neuronal A- and B-cells were obtained using the optical fractionator and optical disector techniques. At birth, the total number of DRG neurons was 10,000 +/- 2,600 in control mice compared with 5,100 +/- 1,300 in p75 knockout mice. During postnatal development, 1,400 neuronal B-cell bodies were lost in p75 knockouts (2P < 0.05) and 1,100 in controls (NS), whereas the A-cell population remained stable. After a sciatic nerve crush, the total neuron loss in controls was 15.4% +/- 3.5% (2P < 0.05) and 22.7% +/- 5.1% (2P < 0.05) at days 14 and 42, respectively. In contrast, there was no loss in total number of neurons after crush in p75 knockout mice. Neuronal A-cell number was unchanged after the crush in p75 knockouts as well as in controls at both times. At 14 days, the population of B-cells was reduced by 24.8% +/- 3.6% in controls and by 6.1% +/- 3.5% in p75 knockouts, this difference being significant (2P < 0.001). At 42 days, the B-cell loss was 29.6% +/- 5.5% in controls and 4.2% +/- 6.4% in p75 knockouts (2P < 0.001). In conclusion, the lack of the p75 receptor results in neuronal DRG cells that are resistant to nerve injury, pointing to a role for the receptor in apoptosis. Copyright 2003 Wiley-Liss, Inc.
Schaeffer, Véronique; Meyer, Laurence; Patte-Mensah, Christine; Eckert, Anne; Mensah-Nyagan, Ayikoe G
Neurosteroids are synthesized either by glial cells, by neurons, or within the context of neuron-glia cross-talk. Various studies suggested neurosteroid involvement in the control of neurodegeneration but there is no evidence showing that the natural protection of nerve cells against apoptosis directly depends on their own capacity to produce neuroprotective neurosteroids. Here, we investigated the interactions between neurosteroidogenesis and apoptosis occurring in sensory structures of rats subjected to neuropathic pain generated by sciatic nerve chronic constriction injury (CCI). Using the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL), we observed no apoptotic cells in the spinal cord up to 30 days after CCI although pain symptoms such as mechano-allodynia, thermal and mechanical hyperalgesia were evidenced with the Hargreaves's behavioral and von Frey filament tests. In contrast, double-labeling experiments combining TUNEL and immunostaining with antibodies against glutamine synthetase or neuronal nuclei protein revealed apoptosis occurrence in satellite glial cells (SGC) (not in neurons) of CCI rat ipsilateral dorsal root ganglia (DRG) at day 30 after injury. Pulse-chase experiments coupled with high performance liquid chromatography and flow scintillation detection showed that, among numerous biosynthetic pathways converting [(3)H]pregnenolone into various [(3)H]neurosteroids, only [(3)H]estradiol formation was selectively modified and upregulated in DRG of CCI rats. Consistently, immunohistochemical investigations localized aromatase (estradiol-synthesizing enzyme) in DRG neurons but not in SGC. Pharmacological inhibition of aromatase caused apoptosis of CCI rat DRG neurons. Altogether, our results suggest that endogenously produced neurosteroids such as estradiol may be pivotal for the protection of DRG sensory neurons against sciatic nerve CCI-induced apoptosis.
Yu, Tianchao; Lin, Wanchun
Application of general anesthetics may induce neurotoxicity in dorsal root ganglia (DRG) neurons. In this study, we examined the possible protective mechanism and associated signaling pathways of small-molecule glycogen synthase kinase-3 (GSK-3) inhibitor, SB216763, in bupivacaine-injured mouse DRG neurons in vitro. In vitro DRG explant of 6-week old mice was treated with 5mM bupivacaine to induce neurotoxicity. The explants were also pre-treated with SB216763 for 72h. Neural protection of SB216763 on bupivacaine-injured DRG neurons was investigated by TUNEL assay, neurite outgrowth assay and western blot assay, respectively. Possible downstream gene of GSK-3 signaling pathway, protein kinase C (PKC) was knocked down by siRNA in DRG explant. Its function in regulating GSK-3 inhibition induced DRG neural protection was also examined by TUNEL, neurite outgrowth and western blot assays. Pre-treatment of SB216763 significantly ameliorated bupivacaine induced apoptosis and neurite loss in DRG neurons. Western blot showed that, in addition to the decrease of phosphorylated-GSK-3 α/β protein, SB216763 increased PKC and decreased caspase-3 (Casp-3) in bupivacaine-injured DRG neurons. SiRNA-mediated PKC knockdown was able to reverse the neural protection of SB216763 in bupivacaine-injured DRG neurons. Western blot showed that PKC knockdown increased phosphorylated-GSK-3 α/β and Casp-3 protein in DRG neurons, confirming that PKC was directly involved in GSK-3-inhibition induced neural protection in DRG. GSK-3 inhibitor SB216763, through PKC, is effective in protecting anesthetics-induced neurotoxicity in DRG. Copyright © 2016. Published by Elsevier Masson SAS.
Fang, Xiao; Shi, Xiao-Han; Huang, Li-Bin; Rong, Wei-Fang; Ma, Bei
The study was aimed to investigate the changes in mechanical pain threshold in the condition of chronic inflammatory pain after transient receptor potential vanilloid 1 (TRPV1) gene was knockout. Hind-paw intraplantar injection of complete freund's adjuvant (CFA, 20 μL) produced peripheral inflammation in wild-type and TRPV1 knockout female mice. The mechanical pain thresholds were measured during the 8 days after injection and pre-injection by using Von-Frey hair. Nine days after injection, mice were killed and the differences of expression of c-Fos and P2X3 receptor in the dorsal root ganglia (DRG) and spinal cord dorsal horn were examined by Western blotting between the two groups. Compared with that in wild-type mice, the mechanical pain threshold was increased significantly in TRPV1 knockout mice (P < 0.05); 3 days after CFA injection, the baseline mechanical pain threshold in the TRPV1 knockout mice group was significantly higher than that in the wild-type mice group (P < 0.05); The result of Western blotting showed that the expression of c-Fos protein both in DRG and spinal cord dorsal horn of TRPV1 knockout mice group was decreased significantly compared with that in wild-type mice group (P < 0.01, P < 0.05), while the expression of P2X3 receptor in DRG of TRPV1 knockout mice group was increased significantly compared with that in wild-type mice group (P < 0.05). Our findings indicate that TRPV1 may influence the peripheral mechanical pain threshold by mediating the expression of c-Fos protein both in DRG and spinal cord dorsal horn and changing the expression of P2X3 receptor in DRG.
Bossowska, A; Majewski, M
Botulinum toxin type A (BTX) is a potent neurotoxin, which in recent years has been effectively applied in experimental treatments of many neurogenic disorders of the urinary bladder. BTX is a selective, presynaptically-acting blocking agent of acetylcholine release from nerve terminals what, in turn, leads to the cessation of somatic motor and/or parasympathetic transmission. However, application of this toxin in urological practice is still in the developmental stages and the full mechanism of its action remain elusive. Thus, the present study was aimed at investigating the neurochemical characterization of dorsal root ganglion (DRG) neurons supplying the porcine urinary bladder after BTX treatment. Retrograde tracer Fast Blue (FB) was injected into the urinary bladder wall in six juvenile female pigs and three weeks later, intramural bladder injections of BTX (100 IU per animal) were carried out in all the animals. After a week, DRG from L1 to Cql were harvested from the pigs and neurochemical characterization of FB+ neurons was performed using double- labeling immunofluorescence technique on 10-microm-thick cryostat sections. BTX injections led to a significant decrease in the number of FB+ neurons containing substance P (SP), calcitonin gene-related peptide (CGRP), calbindin (CB), somatostatin (SOM) and neuronal nitric oxide synthase (nNOS) when compared with that found in the healthy animals (19% vs. 45%, 18% vs. 36%, 0.6% vs. 3%, 0.4 vs. 4% and 0.1% vs. 6%, respectively) These data demonstrated that BTX changed the chemical coding of bladder sensory neurons, and therefore this drug should be taken into consideration when it planning experimental therapy of selected neurogenic bladder disorders.
Lyu, Chuang; Lyu, Gong-Wei; Martinez, Aurora; Shi, Tie-Jun Sten
Background The proapoptotic molecule BAX, plays an important role in mitochondrial apoptotic pathway. Dorsal root ganglion (DRG) neurons depend on neurotrophic factors for survival at early developmental stages. Withdrawal of neurotrophic factors will induce apoptosis in DRG neurons, but this type of cell death can be delayed or prevented in neonatal Bax knockout (KO) mice. In adult animals, evidence also shows that DRG neurons are less dependent upon neurotrophic factors for survival. However, little is known about the effect of Bax deletion on the survival of normal and denervated DRG neurons in adult mice. Methods A unilateral sciatic nerve transection was performed in adult Bax KO mice and wild-type (WT) littermates. Stereological method was employed to quantify the number of lumbar-5 DRG neurons 1 month post-surgery. Nerve injury-induced autotomy behavior was also examined on days 1, 3, and 7 post-surgery. Results There were significantly more neurons in contralateral DRGs of KO mice as compared with WT mice. The number of neurons was reduced in ipsilateral DRGs in both KO and WT mice. No changes in size distributions of DRG neuron profiles were detected before or after nerve injury. Injury-induced autotomy behavior developed much earlier and was more serious in KO mice. Conclusion Although postnatal death or loss of DRG neurons is partially prevented by Bax deletion, this effect cannot interfere with long-term nerve injury-induced neuronal loss. The exaggerated self-amputation behavior observed in the mutant mice indicates that Bax deficiency may enhance the development of spontaneous pain following nerve injury. PMID:28919807
Liuzzi, F J; Scoville, S A; Bufton, S M
Estrogen status has profound effects on cutaneous sensitivity in adult female rats. The presence of alpha-estrogen receptor mRNA and protein in NGF-dependent, adult female rat dorsal root ganglion (DRG) neurons raises the possibility that estrogen modulates cutaneous sensation by acting directly on primary afferent neurons, perhaps by altering their sensitivity to NGF. The present study examined the effect of long-term (90 days) daily injections of an estrogen preparation, Premarin (Wyeth-Ayerst, Radnor, PA), to ovariectomized adult rats on lumbar DRG high-affinity NGF receptor, trkA, mRNA levels, and on beta-preprotachykinin (beta-PPT) mRNA levels, which have been shown to be regulated by NGF. Two doses were used in the experiments, the higher dose being 10 times that of the lower dose. Such injections had an effect opposite that reported for short-term, acute estrogen treatment on DRG trkA mRNA levels. The current data show that long-term daily estrogen treatment decreases trkA mRNA levels by 36%. After 90 days of estrogen treatment, no dose effect was evident. Moreover, as would be expected if beta-PPT gene expression is regulated by NGF through the trkA receptor, long-term estrogen treatment decreased DRG neuronal beta-PPT mRNA levels by about 30%. As with trkA, there was no dose effect evident after 90 days of estrogen treatment. These data suggest the possibility that estrogen modulates DRG neuropeptide gene expression and, perhaps, cutaneous sensitivity by regulating NGF receptor gene expression. Copyright 1999 Academic Press.
Özgül, C; Nazıroğlu, M
N-acetylcysteine (NAC) is a thiol-containing (sulphydryl donor) antioxidant, which contributes to regeneration of glutathione (GSH) and also acts through a direct reaction with free radicals. Thiol depletion has been implicated in the neurobiology of sensory neurons and pain. We reported recently an activator role of intracellular GSH depletion on calcium influx through transient receptor potential melastatin-like 2 (TRPM2) channels in rat dorsal root ganglion (DRG). NAC may have a protective role on calcium influx through regulation of TRPM2 channels in the neurons. Therefore, we tested the effects of NAC on TRPM2 channel currents in cytosolic GSH depleted DRG in rats. DRG neurons were freshly isolated from rats and the neurons were incubated for 24 h with buthionine sulfoximine (BSO). In whole-cell patch clamp experiments, TRPM2 currents in the DRG incubated with BSO were gated by H(2)O(2). TRPM2 channels current densities, cytosolic free Ca(2+) content, and lipid peroxidation values in the neurons were higher in H(2)O(2) and BSO + H(2)O(2) group than in controls; however GSH and GSH peroxidase (GSH-Px) values were decreased. BSO + H(2)O(2)-induced TRPM2 channel gating was totally inhibited by extracellular NAC and partially inhibited by 2-aminoethyl diphenylborinate. GSH-Px activity, lipid peroxidation and GSH levels in the DRG neurons were also modulated by NAC. In conclusion, we observed a modulator role of NAC on Ca(2+) influx through a TRPM2 channel in intracellular GSH depleted DRG neurons. NAC incubation before BSO exposure appears to be more protective than NAC incubation after BSO exposure. Since cytosolic thiol group depletion is a common feature of neuropathic pain, our findings are relevant to the etiology and treatment of pain neuropathology in DRG neurons. Copyright © 2012 Elsevier Inc. All rights reserved.
Nazıroğlu, Mustafa; Özgül, Cemil; Çelik, Ömer; Çiğ, Bilal; Sözbir, Ercan
Exposure to oxidative stress causes health problems, including sensory neuron neuropathy and pain. Rotenone is a toxin used to generate intracellular oxidative stress in neurons. However, the mechanism of toxicity in dorsal root ganglion (DRG) neurons has not been characterized. Melastatin-like transient receptor potential 2 (TRPM2) channel activation and inhibition in response to oxidative stress, ADP-ribose (ADPR), flufenamic acid (FFA) and 2-aminoethoxydiphenyl borate (2-APB) in DRG neurons are also not clear. We tested the effects of FFA and 2-APB on ADPR and rotenone-induced TRPM2 cation channel activation in DRG neurons of rats. DRG neurons were freshly isolated from rats and studied with the conventional whole-cell patch-clamp technique. Rotenone, FFA and 2-APB were extracellularly added through the patch chamber, and ADPR was applied intracellularly through the patch pipette. TRPM2 cation currents were consistently induced by ADPR and rotenone. Current densities of the neurons were higher in the ADPR and rotenone groups than in control. The time courses (gating times) in the neurons were longer in the rotenone than in the ADPR group. ADPR and rotenone-induced TRPM2 currents were totally blocked by 2-APB and partially blocked by FFA. In conclusion, TRPM2 channels were constitutively activated by ADPR and rotenone, and 2-APB and FFA induced an inhibitory effect on TRPM2 cation channel currents in rat DRG neurons. Since oxidative stress is a common feature of neuropathic pain and diseases of sensory neurons, the present findings have broad application to the etiology of neuropathic pain and diseases of DRG neurons.
Demirdaş, Arif; Nazıroğlu, Mustafa; Övey, İshak Suat
Overload of Ca(2+) entry and excessive oxidative stress in neurons are the two main causes of depression. Activation of transient receptor potential (TRP) vanilloid type 1 (TRPV1) and TRP melastatin 2 (TRPM2) during oxidative stress has been linked to neuronal survival. Duloxetine (DULOX) in neurons reduced the effects of Ca(2+) entry and reactive oxygen species (ROS) through glutamate receptors, and this reduction of effects may also occur through TRPM2 and TRPV1 channels. In order to better characterize the actions of DULOX in peripheral pain and hippocampal oxidative injury through modulation of TRPM2 and TRPV1, we tested the effects of DULOX on apoptosis and oxidative stress in the hippocampal and dorsal root ganglion (DRG) neurons of rats. Freshly isolated hippocampal and DRG neurons were incubated for 24 h with DULOX. In whole-cell patch-clamp and intracellular-free calcium ([Ca(2+)]) concentration (Fura-2) experiments, cumene hydroperoxide and ADP-ribose-induced TRPM2 currents in the neurons were inhibited by N-(p-amylcinnamoyl) anthranilic acid (ACA) and capsaicin-induced TRPV1 currents were inhibited by capsazepine (CPZ) incubations. TRPM2 and TRPV1 channel current densities, [Ca(2+)] concentration, apoptosis, caspase 3, caspase 9, mitochondrial depolarization, and intracellular ROS production values in the neurons were lower in the DULOX group than in controls. In addition, the above values were further decreased by DULOX + CPZ and DULOX + ACA treatments. In conclusion, TRPM2 and TRPV1 channels are involved in Ca(2+) entry-induced neuronal death and modulation of the activity of these channels by DULOX treatment may account for their neuroprotective activity against apoptosis, excessive ROS production, and Ca(2+) entry.
Ritter, David M; Ho, Cojen; O'Leary, Michael E; Covarrubias, Manuel
Fast inactivation of heterologously expressed Kv3.4 channels is dramatically slowed upon phosphorylation of the channel's N-terminal (N-type) inactivation gate by protein kinase C (PKC). However, the presence and physiological importance of this exquisite modulation in excitable tissues were unknown. Here, we employed minimally invasive cell-attached patch-clamping, single-cell qPCR and specific siRNAs to unambiguously demonstrate that fast-inactivating Kv3.4 channels underlie a robust high voltage-activated A-type K(+) current (I(AHV)) in nociceptive dorsal root ganglion neurons from 7-day-old rats. We also show that PKC activation with phorbol 12,13-dibutyrate (PDBu) causes a 4-fold slowing of Kv3.4 channel inactivation and, consequently, accelerates the repolarization of the action potential (AP) by 22%, which shortens the AP duration by 14%. G-protein coupled receptor (GPCR) agonists eliminate I(AHV) fast inactivation in a membrane-delimited manner, suggesting a Kv3.4 channel signalling complex. Preincubation of the neurons with the PKC inhibitor bisindolylmaleimide II inhibits the effect of GPCR agonists and PDBu. Furthermore, activation of PKC via GPCR agonists recapitulates the effects of PDBu on the AP. Finally, transfection of the neurons with Kv3.4 siRNA prolongs the AP by 25% and abolishes the GPCR agonist-induced acceleration of the AP repolarization. These results show that Kv3.4 channels help shape the repolarization of the nociceptor AP, and that modulation of Kv3.4 channel N-type inactivation by PKC regulates AP repolarization and duration. We propose that the dramatic modulation of I(AHV) fast inactivation by PKC represents a novel mechanism of neural plasticity with potentially significant implications in the transition from acute to chronic pain.
Ma, W; St-Jacques, B; Rudakou, U; Kim, Y N
Persistent peripheral sensitization contributes to chronic pain. Plasticity of nociceptive dorsal root ganglion (DRG) neurons (nociceptors) induced by pro-inflammatory mediators contributes to sensitization. Prostaglandin E2 (PGE2) enriched in injured tissues is known not only directly to sensitize DRG neurons, but also to potentiate sensitizing effects of other pain mediators such as capsaicin and its receptor transient receptor potential vanilloid-1 (TRPV1). It remains unknown whether PGE2 potentiates TRPV1 activity by stimulating its synthesis, cell surface and axonal trafficking in DRG neurons. Combined biochemical, morphological, pharmacological and behavioral approaches have been used to address this issue in both in vitro and in vivo models. PGE2 increased TRPV1 externalization in cultured rat DRG neurons in a time- and concentration-dependent manner, an event blocked by an inhibitor of protein synthesis or anterograde export. EP1 and EP4, but not EP2 and EP3, mediated this event. EP1 agonist-induced TRPV1 externalization was suppressed by inhibitors of CaMKII, PLC, PKC and PKCε, while EP4 agonist-induced TRPV1 externalization by inhibitors of cAMP/PKA and ERK/MAPK. Pre-exposure to PGE2 potentiated release of calcitonin gene-related peptide from cultured DRG neurons evoked by subsequent capsaicin stimulation. This event was blocked by an inhibitor of protein synthesis or export, suggesting that PGE2-induced TRPV1 synthesis and externalization is coupled to enhanced TRPV1 activity. Pre-exposure to PGE2 not only prolonged tactile allodynia evoked by subsequent capsaicin challenge, but also increased TRPV1 levels in L4-6 DRG, sciatic nerves and plantar skin. Our data indicate that facilitating TRPV1 synthesis, cell surface and axonal trafficking is a novel mechanism underlying PGE2 potentiation of TRPV1 activity. © 2016 European Pain Federation - EFIC®.
Song, Ying; Zhang, Yong-Mei; Xu, Jie; Wu, Jing-Ru; Qin, Xia; Hua, Rong
The aim of the paper is to study the effect of spontaneous firing of injured dorsal root ganglion (DRG) neuron in chronic compression of DRG (CCD) model on excitability of wide dynamic range (WDR) neuron in rat spinal dorsal horn. In vivo intracellular recording was done in DRG neurons and in vivo extracellular recording was done in spinal WDR neurons. After CCD, incidence of spontaneous discharge and firing frequency enhanced to 59.46% and (4.30 ± 0.69) Hz respectively from 22.81% and (0.60 ± 0.08) Hz in normal control group (P < 0.05). Local administration of 50 nmol/L tetrodotoxin (TTX) on DRG neuron in CCD rats decreased the spontaneous activities of WDR neurons from (191.97 ± 45.20)/min to (92.50 ± 30.32)/min (P < 0.05). On the other side, local administration of 100 mmol/L KCl on DRG neuron evoked spontaneous firing in a reversible way (n = 5) in silent WDR neurons of normal rats. There was 36.36% (12/33) WDR neuron showing after-discharge in response to innocuous mechanical stimuli on cutaneous receptive field in CCD rats, while after-discharge was not seen in control rats. Local administration of TTX on DRG with a concentration of 50 nmol/L attenuated innocuous electric stimuli-evoked after-discharge of WDR neurons in CCD rats in a reversible manner, and the frequency was decreased from (263 ± 56.5) Hz to (117 ± 30) Hz (P < 0.05). The study suggests that the excitability of WDR neurons is influenced by spontaneous firings of DRG neurons after CCD.
Walwyn, Wendy; John, Scott; Maga, Matthew; Evans, Christopher J; Hales, Tim G
Recombinant micro and delta opioid receptors expressed in cell lines can form heterodimers with distinctive properties and trafficking. However, a role for opioid receptor heterodimerization in neurons has yet to be identified. The inhibitory coupling of opioid receptors to voltage-dependent Ca(2+) channels (VDCCs) is a relatively inefficient process and therefore provides a sensitive assay of altered opioid receptor function and expression. We examined micro-receptor coupling to VDCCs in dorsal root ganglion neurons of delta(+/+), delta(+/-), and delta(-/-) mice. Neurons deficient in delta receptors exhibited reduced inhibition of VDCCs by morphine and [D-Ala(2),Phe(4),Gly(5)-ol]-enkephalin (DAMGO). An absence of delta receptors caused reduced efficacy of DAMGO without affecting potency. An absence of delta receptors reduced neither the density of VDCCs nor their inhibition by either the GABA(B) receptor agonist baclofen or intracellular guanosine 5'-O-(3-thio)triphosphate. Flow cytometry revealed a reduction in micro-receptor surface expression in delta(-/-) neurons without altered DAMGO-induced internalization. There was no change in micro-receptor mRNA levels. D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH(2)-sensitive mu-receptor-coupling efficacy was fully restored to delta(+/+) levels in delta(-/-) neurons by expression of recombinant delta receptors. However, the dimerization-deficient delta-15 construct expressed in delta(-/-) neurons failed to fully restore the inhibitory coupling of micro-receptors compared with that seen in delta(+/+) neurons, suggesting that, although not essential for micro-receptor function, micro-delta receptor dimerization contributes to full micro-agonist efficacy. Because DAMGO exhibited a similar potency in delta(+/+) and delta(-/-) neurons and caused similar levels of internalization, the role for heterodimerization is probably at the level of receptor biosynthesis.
Ding, Xin-Li; Wang, Yong-Hui; Ning, Li-Ping; Zhang, Yang; Ge, Hong-You; Jiang, Hong; Wang, Rong; Yue, Shou-Wei
The aim of the present study was to test the hypothesis that the TRPV4-NO-cGMP-PKG cascade is involved in the maintenance of thermal hyperalgesia following chronic compression of the dorsal root ganglion (DRG) (the procedure hereafter termed CCD) in rats. CCD rats showed thermal hyperalgesia and increased nitrite production. Intrathecal administration of ruthenium red (TRPV4 antagonist, 0.1-1 nmol), TRPV4 antisense ODN (TRPV4 AS, 40 microg, daily for 7 days), N(G)-L-nitro-arginine methyl ester (l-NAME, inhibitor of NO synthase, 30-300 nmol), 1H-[1,2,4]-oxadiazolo [4,3-a] quinoxalin-1-one (ODQ, a soluble guanylate cyclase inhibitor, 50-100 nmol) or 8-(4-Chlorophenylthio) guanosine 3',5'-cyclic Monophosphothioate, Rp-Isomer sodium salt (Rp-8-pCPT-cGMPS, a PKG inhibitor, 25-50 nmol) induced a significant (P<0.001) and dose-dependent increase in the paw withdrawal latency (PWL) compared with control rats, respectively. Ruthenium red (1 nmol), TRPV4 AS (40 microg, daily for 7 days) or L-NAME (300 nmol) decreased nitrite (an index of nitric oxide formation) in the DRG of CCD rats. In addition, the phorbol ester 4alpha-phorbol 12,13-didecanoate (4alpha-PDD, TRPV4 synthetic activator, 1 nmol), co-administered with L-NAME (300 nmol), attenuated the suppressive effect of L-NAME on CCD-induced thermal hyperalgesia and nitrite production. Our data suggested that the TRPV4-NO-cGMP-PKG pathway could be involved in CCD-induced thermal hyperalgesia.
Li, Yan; Tatsui, Claudio Esteves; Rhines, Laurence D; North, Robert Y; Harrison, Daniel S; Cassidy, Ryan M; Johansson, Caj A; Kosturakis, Alyssa K; Edwards, Denaya D; Zhang, Hongmei; Dougherty, Patrick M
Here, it is shown that paclitaxel-induced neuropathy is associated with the development of spontaneous activity (SA) and hyperexcitability in dorsal root ganglion (DRG) neurons that is paralleled by increased expression of low-voltage-activated calcium channels (T-type; Cav3.2). The percentage of DRG neurons showing SA and the overall mean rate of SA were significantly higher at day 7 in rats receiving paclitaxel treatment than in rats receiving vehicle. Cav3.2 expression was increased in L4-L6 DRG and spinal cord segments in paclitaxel-treated rats, localized to small calcitonin gene-related peptide and isolectin B4 expressing DRG neurons and to glial fibrillary acidic protein-positive spinal cord cells. Cav3.2 expression was also co-localized with toll-like receptor 4 (TLR4) in both the DRG and the dorsal horn. T-type current amplitudes and density were increased at day 7 after paclitaxel treatment. Perfusion of the TLR4 agonist lipopolysaccharide directly activated DRG neurons, whereas this was prevented by pretreatment with the specific T-type calcium channel inhibitor ML218 hydrochloride. Paclitaxel-induced behavioral hypersensitivity to mechanical stimuli in rats was prevented but not reversed by spinal administration of ML218 hydrochloride or intravenous injection of the TLR4 antagonist TAK242. Paclitaxel induced inward current and action potential discharges in cultured human DRG neurons, and this was blocked by ML218 hydrochloride pretreatment. Furthermore, ML218 hydrochloride decreased firing frequency in human DRG, where spontaneous action potentials were present. In summary, Cav3.2 in concert with TLR4 in DRG neurons appears to contribute to paclitaxel-induced neuropathy.
Shypshyna, M S; Veselovsky, N S; Myasoedov, N F; Shram, S I; Fedulova, S A
The influence of long-term culturing (12 days in vitro) of dorsal root ganglion (DRG) and dorsal horn (DH) neurons with peptide Semax on the level of synaptic activity at co-cultures, as well as short-term plasticity in sensory synapses were studied. It has been shown that neuronal culturing with peptide at concentrations of 10 and 100 µM led to increasing the frequency of spontaneous glutamatergic postsynaptic currents in DH neurons to 71.7 ± 1.8% and 93.9 ± 3.1% (n = 6; P < 0.001); Semax has a not significant effect on the amplitude and frequency of miniature glutamatergic currents, but causes an increase of the amplitudes of spontaneous postsynaptic currents, as well as elevates the quantum content. The data show the increase of multivesicular glutamate release efficiency in neural networks of co-cultures following incubation with the peptide. Also Semax (10 and 100 µM) induces changes of the basic parameters of short-term plasticity in sensory synapses: (1) increasing the paired-pulse ratio from 0.53 ± 0.028 (n = 8) to 0.91 ± 0.072 (n = 6, P < 0.01) and 0.95 ± 0.026 (n = 7; P < 0.001); (2) reducing the ratio of the coefficients of variation (CV2/ CV1) from 1.49 ± 0.11 (n = 8) to 1.02 ± 0.09 (n = 6; P < 0.05) and 1.11 ± 0.13 (n = 7; P < 0.0) respectively. The results indicate a stimulating effect of Semax on the activity of glutamatergic synapses in neural networks of co-cultures, as well as the ability of the peptide to effectively modulate the short-term plasticity in sensory synapses.
Scheff, Nicole N.; Lu, Shao-Gang; Gold, Michael S.
Persistent inflammation results in an increase in the magnitude and duration of high K+-evoked Ca2+ transients in putative nociceptive cutaneous dorsal root ganglion (DRG) neurons. The purpose of the present study was to determine whether recruitment of Ca2+-induced Ca2+ release (CICR) contributes to these inflammation-induced changes. Acutely dissociated, retrogradely labeled cutaneous DRG neurons from naïve and complete Freund’s adjuvant inflamed adult male Sprague Dawley rats were studied with ratiometric microfluorimetry. Ryanodine only attenuated the duration but not magnitude of the high K+-evoked Ca2+ transient in neurons from inflamed rats. However, there was no significant impact of inflammation on the potency or efficacy of ryanodine-induced block of the caffeine-evoked Ca2+ transient, or the impact of sarco-endoplasmic reticulum ATPase (SERCA) inhibition on the high K+-evoked Ca2+ transient. Furthermore, while there was no change in the magnitude, an inflammation-induced increase in the duration of the caffeine-evoked Ca2+ transient was only observed with a prolonged caffeine application. In contrast to the high K+-evoked Ca2+ transient, there was no evidence of direct mitrochondrial involvement or that of the Ca2+ extrusion mechanism, the Na+/Ca2+ exchanger, on the caffeine-evoked Ca2+ transient, and block of SERCA only increased the duration of this transient. These results indicate the presence of Ca2+ regulatory domains in cutaneous nociceptive DRG neurons within which cytosolic Ca2+ increased via influx and release are highly segregated. Furthermore, our results suggest that changes in neither CICR machinery nor the coupling between Ca2+ influx and CICR are primarily responsible for the inflammation-induced changes in the evoked Ca2+ transient. PMID:23642703
Nazıroğlu, M; Çiğ, B; Özgül, C
A main component of St. John's Wort (Hypericum perforatum, HP) is hyperforin which has antioxidant properties in dorsal root ganglion (DRG) neurons, due to its ability to modulate NADPH oxidase and protein kinase C. Recent reports indicate that oxidative stress through NADPH oxidase activates TRPM2 channels. HP may be a useful treatment for Ca(2+) entry and oxidative stress through modulation of TRPM2 channels in the DRG. We aimed to investigate the protective role of HP on Ca(2+) entry and oxidative stress through TRPM2 channels in DRG neurons of rats. The native rat DRG neurons were used in whole-cell patch-clamp, Fura-2 and antioxidant experiments. Appropriate, nontoxic concentrations and incubation times for HP were determined in the DRG neurons by assessing cell viability. The H2O2-induced TRPM2 currents were inhibited by 2-aminoethyl diphenylborinate (2-APB) and N-(p-amylcinnamoyl)anthranilic acid (ACA). TRPM2 current densities and cytosolic free Ca(2+) concentration in the neurons were also reduced by HP (2 and 24h). In Fura-2 experiments, cytosolic Ca(2+) mobilization was reduced by voltage-gated calcium channel blockers (verapamil+diltiazem, V+D) and HP. Glutathione peroxidase activity and GSH values in the DRG were high in HP, 2-APB and V+D groups although lipid peroxidation level was low in the groups. In conclusion, we observed a protective role for HP on Ca(2+) entry through a TRPM2 channel in the DRG neurons. Since over-production of oxidative stress and Ca(2+) entry are implicated in the pathophysiology of neuropathic pain and neuronal inflammation, our findings may be relevant to the etiology and treatment of neuropathology in DRG neurons.
Morioka, Norimitsu; Yoshida, Yosuke; Nakamura, Yoki; Hidaka, Nobue; Hisaoka-Nakashima, Kazue; Nakata, Yoshihiro
Although brain-derived neurotrophic factor (BDNF) is localized in primary sensory neurons and has crucial roles in nociceptive transduction, the mechanisms involved in regulation of BDNF exon-specific mRNA expression in dorsal root ganglion (DRG) neurons have yet to be determined. Rat primary cultures of DRG neurons were stimulated with phorbol-12-myristate-13-acetate (PMA), a potent activator of protein kinase C (PKC), which resulted in the robust expression of both BDNF mRNA and protein. Among each BDNF mRNA exon, it was found that exons I, IV and VI were especially induced after PMA stimulation. The induction of these exons was significantly blocked by Gö6983 (a broad spectrum PKC inhibitor), Gö6976 (a conventional PKCs and PKCμ inhibitor), and rottlerin (a PKCδ inhibitor), but not by a PKCε inhibitor. The effect of PMA on exons I and VI was blocked by either U0126 (a MAP kinase kinase (MEK) inhibitor) or SB202190 (a p38 inhibitor), and PMA's effect on exon IV was inhibited by U0126 but not by SB202190. Furthermore, the activation of cAMP-responsive element-binding protein (CREB) was associated with the induction of exons I and IV, and the activation of nuclear factor-κB (NF-κB) contributed to the induction of exons I, IV and VI. These results show that the activation of PKCs induces the expression of BDNF mRNA exons I, IV and VI through exon-specific mechanisms, including extracellular signal-regulated kinase, p38, CREB and NF-κB, in cultured DRG neurons. These data suggest multiple pathways in the expression of BDNF in nociceptive sensory neurons.
Huang, Jianying; Yang, Yang; Zhao, Peng; Gerrits, Monique M; Hoeijmakers, Janneke G J; Bekelaar, Kim; Merkies, Ingemar S J; Faber, Catharina G; Dib-Hajj, Sulayman D; Waxman, Stephen G
Idiopathic small-fiber neuropathy (I-SFN), clinically characterized by burning pain in distal extremities and autonomic dysfunction, is a disorder of small-caliber nerve fibers of unknown etiology with limited treatment options. Functional variants of voltage-gated sodium channel Nav1.7, encoded by SCN9A, have been identified in approximately one-third of I-SFN patients. These variants render dorsal root ganglion (DRG) neurons hyperexcitable. Sodium channel Nav1.8, encoded by SCN10A, is preferentially expressed in small-diameter DRG neurons, and produces most of the current underlying the upstroke of action potentials in these neurons. We previously demonstrated two functional variants of Nav1.8 that either enhance ramp current or shift activation in a hyperpolarizing direction, and render DRG neurons hyperexcitable, in I-SFN patients with no mutations of SCN9A. We have now evaluated additional I-SFN patients with no mutations in SCN9A, and report a novel I-SFN-related Nav1.8 mutation I1706V in a patient with painful I-SFN. Whole-cell voltage-clamp recordings in small DRG neurons demonstrate that the mutation hyperpolarizes activation and the response to slow ramp depolarizations. However, it decreases fractional channels resistant to fast inactivation and reduces persistent currents. Current-clamp studies reveal that mutant channels decrease current threshold and increase the firing frequency of evoked action potentials within small DRG neurons. These observations suggest that the effects of this mutation on activation and ramp current are dominant over the reduced persistent current, and show that these pro-excitatory gating changes confer hyperexcitability on peripheral sensory neurons, which may contribute to pain in this individual with I-SFN.
Liu, Xiangming; Chen, Su; Zhang, Yuxia; Zhang, Fan
To clarify the modulation of dragon's blood on the tetrodotoxin-resistant (TTX-R) sodium currents in dorsal root ganglion (DRG) neurons and explore its corresponding material basis for the efficacy, using whole-cell patch clamp technique, the effects of dragon's blood and the combined effects of three components (cochinchinenin A, cochinchinenin B, and loureirin B) extracted from dragon's blood on the TTX-R sodium currents in acute-isolated DRG neurons of rats were observed. According to the operational definition of material basis for the efficacy of TCM established, the material basis of the modulation on the TTX-R sodium currents in DRG neurons of dragon's blood was judged from the experimental results. The drug interaction equation of Greco et al. was used to assess the interaction of the three components extracted from dragon's blood. This investigation demonstrated that dragon's blood suppressed the peak TTX-R sodium currents in a dose-dependent way and affected the activations of TTX-R sodium currents. The effects of the combination of cochinchinenin A, cochinchinenin B, and loureirin B were in good agreement with those of dragon's blood. Although the three components used alone could modulate TTX-R sodium currents, the concentrations of the three components used alone were respectively higher than those used in combination when the inhibition rates on the TTX-R sodium currents of them used alone and in combination were the same. The combined effects of the three components were synergistic. These results suggested that the interference with pain messages caused by the modulation of dragon's blood on TTX-R sodium currents in DRG neurons may explain some of the analgesic effect of dragon's blood and the corresponding material basis for the efficacy is the combination of cochinchinenin A, cochinchinenin B, and loureirin B.
Bai, Xue; Chen, Tianhua; Gao, Yang; Li, Hao; Li, Zhenzhong; Liu, Zhen
Polybrominated diphenyl ethers (PBDEs) exist extensively in the environment as contaminants, in which 2,2',3,3',4,4',5,5',6,6'-decabrominated diphenyl ether (BDE-209) is the most abundant PBDE found in human samples. BDE-209 has been shown to cause neurotoxicity of primary sensory neurons with few effective therapeutic options available. Here, cultured dorsal root ganglion (DRG) neurons were used to determine the therapeutic effects of insulin-like growth factor-1 (IGF-1) on BDE-209-induced neurotoxicity. The results showed that IGF-1 promoted neurite outgrowth and cell viability of DRG neurons with BDE-209-induced neurotoxicity. IGF-1 inhibited oxidative stress and apoptotic cell death caused by BDE-209 exposure. IGF-1 could reverse the decrease in growth-associated protein-43 (GAP-43) and calcitonin gene-related peptide (CGRP), but not neurofilament-200 (NF-200), expression resulting from BDE-209 exposure. The effects of IGF-1 could be blocked by the extracellular signal-regulated protein kinase (ERK1/2) inhibitor PD98059 and the phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002, either alone or in combination. IGF-1 may play an important role in neuroprotective effects on DRG neurons with BDE-209-induced neurotoxicity through inhibiting oxidative stress and apoptosis and regulating GAP-43 and CGRP expression of DRG neurons. Both ERK1/2 and PI3K/Akt signaling pathways were involved in the effects of IGF-1. Thus, IGF-1 might be one of the therapeutic agents on BDE-209-induced neurotoxicity.
Johnson, I P; Sears, T A
Evidence is emerging for a role of rough endoplasmic reticulum (RER) in the form of stress granules, the unfolded protein response and protein bodies in the response of neurons to injury and in neurodegenerative diseases. Here, we have studied the role of the peripheral target in regulating the RER and polyribosomes of Nissl bodies in axotomised adult cat dorsal root ganglion (DRG) neurons where axonal regeneration and peripheral target reinnervation was either allowed or denied. Retrograde labelling with horseradish peroxidise was used as an independent marker to enable selection of only those DRG neuronal cell bodies with axons in the injured intercostal nerves. Indications of polyribosomal dispersal were seen by 6h following axotomy, and by 24h the normal orderly arrangement of lamellae of RER in Nissl bodies had become disorganised. These ultrastructural changes preceded light microscopical chromatolysis by 1-3d. The retrograde response was maximal 8-32 d after axotomy. Clusters of debris-laden satellite cells/macrophages were present at this time but no ultrastructural evidence of neuronal apoptosis or necrosis was seen and there were no differences in the initial retrograde response according to the type of injury. By 64 d following axotomy with reinnervation, approximately half the labelled DRG neurons showed restoration of the orderly arrangement of RER and polyribosomes in their Nissl bodies. This was not seen after axotomy with reinnervation denied. We propose that the target-dependent changes in Nissl body ultrastructure described here are part of a continuum that can modify neuronal protein synthesis directed towards growth, maintenance or death of the neuron. This represents a possible structural basis for mediating the varied effects of neurotrophic interactions.
He, Xiao-fen; Wei, Jun-jun; Shou, Sheng-yun; Fang, Jian-qiao; Jiang, Yong-liang
Objective: To investigate the analgesic effects of electroacupuncture (EA) at 2 and 100 Hz on type 2 diabetic neuropathic pain (DNP) and on the expressions of the P2X3 receptor and calcitonin gene-related peptide (CGRP) in the dorsal root ganglion (DRG). Methods: Rat type 2 DNP was induced by a high calorie and high sugar diet fed for 7 weeks, plus a single intraperitoneal injection of streptozotocin (STZ) after 5 weeks. EA at 2 and 100 Hz was carried out once every day after 7 weeks for 7 consecutive days. Body weight, serum fasting insulin (FINS), fasting blood glucose (FBG), insulin sensitivity index (ISI), and paw withdrawal latency (PWL) were measured. The expressions of L4–L6 DRG P2X3 receptors and CGRP were assessed by immunofluorescence. Results: Type 2 DNP was successfully induced as shown by the increased body weight, FINS, and FBG, as well as the reduced ISI and PWL. Expressions of P2X3 receptors and CGRP in L4–L6 DRGs increased. EA at both 2 and 100 Hz relieved type 2 DNP, but the analgesic effect of EA was stronger at 2 Hz. P2X3 receptor expression decreased in L4–L6 DRGs following EA at 2 Hz and in L5 and L6 DRGs following EA at 100 Hz. EA at both 2 and 100 Hz down-regulated CGRP overexpression in L4–L6 DRGs. Conclusions: These findings indicate that EA at 2 Hz is a good option for the management of type 2 DNP. The EA effect may be related to its down-regulation of the overexpressions of the DRG P2X3 receptors and CGRP in this condition. PMID:28271659
Hoffman, E. Matthew; Schechter, Ruben; Miller, Kenneth E.
Most, if not all, dorsal root ganglion (DRG) neurons use the neurotransmitter glutamate. There are, however, conflicting reports of the percentages of DRG neurons that express glutaminase (GLS), the enzyme that synthesizes glutamate, ranging from 30% to 100% of DRG neurons. Defining DRG neuron populations by the expression of proteins like GLS, which indicates function, is routinely accomplished with immunolabeling techniques. Proper characterization of DRG neuron populations relies on accurate detection of such antigens. It is known intuitively that fixation can alter immunoreactivity (IR). In this study, we compared the effects of five formaldehyde concentrations between 0.25% and 4.0% (w/v) and five picric acid concentrations between 0.0% and 0.8% (w/v) on the IR of GLS, the voltage-gated sodium channel 1.8 (Nav1.8), and the capsaicin receptor TRPV1. We also compared the effects of five incubation time lengths from 2 to 192 hr, in primary antiserum on IR. Lowering formaldehyde concentration elevated IR for all three antigens, while raising picric acid concentration increased Nav1.8 and TRPV1 IR. Increasing IR improved detection sensitivity, which led to higher percentages of labeled DRG neurons. By selecting fixation conditions that optimized IR, we found that all DRG neurons express GLS, 69% of neurons express Nav1.8, and 77% of neurons express TRPV1, indicating that some previous studies may have underestimated the percentages of DRG neurons expressing these proteins. This manuscript contains online supplemental material at http://www.jhc.org. Please visit this article online to view these materials. (J Histochem Cytochem 58:329–344, 2010) PMID:20026672
Naziroğlu, Mustafa; Ozgül, Cemil
Transient receptor potential ion channel melastatin subtype 8 (TRPM8) is activated by cold temperature and cooling agents, such as menthol and icilin. Compounds containing peppermint are reported to reduce symptoms of environmental cold stress such as cold allodynia in dorsal root ganglion (DRG) neuron; however, the underlying mechanisms of action are unclear. We tested the effects of physiological heat (37°C), anthralic acid (ACA and 0.025 mM), 2-aminoethyl diphenylborinate (2-APB and 0.05) on noxious cold (10°C) and menthol (0.1 mM)-induced TRPM8 cation channel currents in the DRG neurons of rats. DRG neurons were freshly isolated from rats. In whole-cell patch clamp experiments, TRPM8 currents were consistently induced by noxious cold or menthol. TRPM8 channels current densities of the neurons were higher in cold and menthol groups than in control. When the physiological heat is introduced by chamber TRPM8 channel currents were inhibited by the heat. Noxious cold-induced Ca(2+) gates were blocked by the ACA although menthol-induced TRPM8 currents were not blocked by ACA and 2-APB. In conclusion, the results suggested that activation of TRPM8 either by menthol or nociceptive cold can activate TRPM8 channels although we observed the protective role of heat, ACA and 2-APB through a TRPM8 channel in nociceptive cold-activated DRG neurons. Since cold allodynia is a common feature of neuropathic pain and diseases of sensory neuron, our findings are relevant to the etiology of neuropathology in DRG neurons.
Wang, Xi-Jie; An, Shan-Shan; Cheng, Hong; Xu, San-Hua; Cheng, Jie; Lu, Wei; Gao, Rong; Xiao, Hang
A long-chain polypeptide BmKNJX11 was purified from the venom of Asian scorpion Buthus martensi Karsch (BmK) by a combination of gel filtration, ion-exchange chromatography, and reverse-phase high-performance liquid chromatography. The molecular mass was found to be 7036.85 Da by electrospray ionization mass spectrometry. The first 15 N-terminal amino acid sequence of BmKNJX11 was determined to be GRDAY IADSE NCTYT by Edman degradation. With whole cell recording, BmKNJX11 inhibited tetrodotoxin-sensitive voltage-gated sodium channels (TTX-S VGSC) in freshly isolated rat dorsal root ganglion (DRG) neurons in a concentration- and voltage-dependent manner. At a concentration of 40 mug/ml BmKNJX11 lowered the activation threshold and produced negative shifting of TTX-S sodium current (I(Na)) activation curve. In addition, BmKNJX11 induced shifting of the steady-state inactivation curve to the left, delayed the recovery of TTX-S I(Na) from inactivation, and also reduced the fraction of available sodium channels. These results suggested that BmKNJX11 might exert effects on VGSC by binding to a specific site. Considering that TTX-S VGSC expressed in DRG neurons play a critical role in nociceptive transmission, the interaction of BmKNJX11 with TTX-S VGSC might lead to a change in excitability of nociceptive afferent fibers, which may be involved in the observed peripheral pain expression.
Chen, Cheng; Bai, Xue; Bi, Yanwen; Liu, Guixiang; Li, Hao; Liu, Zhen; Liu, Huaxiang
Paclitaxel (PT)-induced neurotoxicity is a significant problem associated with successful treatment of cancers. Insulin-like growth factor-1 (IGF-1) is a neurotrophic factor and plays an important role in promoting axonal growth from dorsal root ganglion (DRG) neurons. Whether IGF-1 has protective effects on neurite growth, cell viability, neuronal apoptosis and neuronal phenotypes in DRG neurons with PT-induced neurotoxicity is still unclear. In this study, primary cultured rat DRG neurons were used to assess the effects of IGF-1 on DRG neurons with PT-induced neurotoxicity. The results showed that PT exposure caused neurite retraction in a dose-dependent manner. PT exposure caused a decrease of cell viability and an increase in the ratio of apoptotic cells which could be reversed by IGF-1. The percentage of calcitonin gene-related peptide immunoreactive (CGRP-IR) neurons and neurofilament (NF)-200-IR neurons, mRNA, and protein levels of CGRP and NF-200 decreased significantly after treatment with PT. IGF-1 administration had protective effects on CGRP-IR neurons, but not on NF-200-IR neurons. Either extracellular signal-regulated protein kinase (ERK1/2) inhibitor PD98059 or phosphatidylinositol 3-kinase (PI3 K) inhibitor LY294002 blocked the effect of IGF-1. The results imply that IGF-1 may attenuate apoptosis to improve neuronal cell viability and promote neurite growth of DRG neurons with PT-induced neurotoxicity. Moreover, these results support an important neuroprotective role of exogenous IGF-1 on distinct subpopulations of DRG neurons which is responsible for skin sensation. The effects of IGF-1 might be through ERK1/2 or PI3 K/Akt signaling pathways. These findings provide experimental evidence for IGF-1 administration to alleviate neurotoxicity of distinct subpopulations of DRG neurons induced by PT.
Hong, Shuangsong; Morrow, Thomas J.; Paulson, Pamela E.; Isom, Lori L.; Wiley, John W.
Diabetic neuropathy is a common form of peripheral neuropathy, yet the mechanisms responsible for pain in this disease are poorly understood. Alterations in the expression and function of voltage-gated tetrodotoxin-resistant (TTX-R) sodium channels have been implicated in animal models of neuropathic pain, including models of diabetic neuropathy. We investigated the expression and function of TTX-sensitive (TTX-S) and TTX-R sodium channels in dorsal root ganglion (DRG) neurons and the responses to thermal hyperalgesia and mechanical allodynia in streptozotocin-treated rats between 4–8 weeks after onset of diabetes. Diabetic rats demonstrated a significant reduction in the threshold for escape from innocuous mechanical pressure (allodynia) and a reduction in the latency to withdrawal from a noxious thermal stimulus (hyperalgesia). Both TTX-S and TTX-R sodium currents increased significantly in small DRG neurons isolated from diabetic rats. The voltage-dependent activation and steady-state inactivation curves for these currents were shifted negatively. TTX-S currents induced by fast or slow voltage ramps increased markedly in neurons from diabetic rats. Immunoblots and immunofluorescence staining demonstrated significant increases in the expression of Nav1.3 (TTX-S) and Nav1.7 (TTX-S) and decreases in the expression of Nav1.6 (TTX-S) and Nav1.8 (TTX-R) in diabetic rats. The level of serine/threonine phosphorylation of Nav1.6 and Nav1.8 increased in response to diabetes. In addition, increased tyrosine phosphorylation of Nav1.6 and Nav1.7 was observed in DRGs from diabetic rats. These results suggest that both TTX-S and TTX-R sodium channels play important roles and that differential phosphorylation of sodium channels involving both serine/threonine and tyrosine sites contributes to painful diabetic neuropathy. PMID:15123645
Lv, H; Chen, H; Xu, J J; Jiang, Y S; Shen, Y J; Zhou, S Z; Xu, H; Xiong, Y C
Reactive oxygen species (ROS) play a critical role in the pathogenesis of neuropathic pain, but few studies have examined the role of oxidative stress in the mirror-image neuropathic pain (MINP). The present study was to investigate the role of ROS in MINP caused by chronic compression of the dorsal root ganglion (DRG) (CCD) in a rat model. SD rats were randomly divided into sham group and CCD group. CCD was conducted to induce MINP. CCD rats were intraperitoneally injected with α-Phenyl-N-tert-butyl-nitrone (PBN) at 7 days after surgery. Paw withdrawal mechanical threshold (PWMT) was measured at -1, 1, 3, 5 and 7 days after surgery in sham group and CCD group, and at 8 time points after PBN injection. Rats were sacrificed at 3 and 7 days after surgery in sham group and CCD group and at 0.5 and 2 h after PBN injection, and the superoxide dismutase (SOD) and catalase activities, as well as hydrogen peroxide (H2O2) and malonaldehyde (MDA) contents were determined in the contralateral DRGs. Results showed bilateral PWMT reduced significantly in sham group and CCD group, but it returned to nearly normal level in sham group. MDA content, H2O2 content and SOD activity increased significantly, while catalase activity remained unchanged in CCD rats. PBN at 100 mg/kg significantly attenuated bilateral mechanical hyperalgesia accompanied by the improvement of oxidative stress in the contralateral DRGs. Our results demonstrate that ROS produced in the contralateral DRG are involved in the pathogenesis of CCD induced MINP, and ROS scavenger may be a promising drug for the therapy of MINP.
Yokota, Kenjiro; Tatebayashi, Hideharu; Matsuo, Tadashi; Shoge, Takashi; Motomura, Haruhiko; Matsuno, Toshiyuki; Fukuda, Akira; Tashiro, Nobutada
1. Several neuroleptics inhibited the 3 microM gamma-aminobutyric acid induced-chloride current (GABA-current) on dissociated rat dorsal root ganglion neurons in whole-cell patch-clamp investigations. 2. The IC(50) for clozapine, zotepine, olanzapine, risperidone and chlorpromazine were 6.95, 18.26, 20.30, 106.01 and 114.56 microM, respectively. The values for the inhibitory effects of neuroleptics on the GABA (3 microM)-current, which were calculated by the fitting Hill's equations where the concentrations represent the mean therapeutic blood concentrations, were ranked clozapine>zotepine>chlorpromazine>olanzapine>risperidone. These inhibitory effects, weighted with the therapeutic concentrations of neuroleptics, were correlated with the clinical incidences of seizure during treatment with neuroleptics. 3. Clozapine reduced the picrotoxin-inhibiton, and may compete with a ligand of the t-butylbicyclophosphorothionate (TBPS) binding site. 4. Haloperidol and quetiapine did not affect the peak amplitude of the GABA (3 microM)-current. However, haloperidol reduced the clozapine-inhibition, and may antagonize ligand binding to TBPS binding site. 5. Neuroleptics including haloperidol and quetiapine enhanced the desensitization of the GABA (3 microM)-current. However, haloperidol and quetiapine at 100 microM inhibited the desensitization at the beginning of application. 6. Blonanserin (AD-5423) at 30 and 50 microM potentiated the GABA (3 microM)-current to 170.1+/-6.9 and 192.0+/-10.6% of the control current, respectively. Blonanserin shifted GABA concentration-response curve leftward. Blonanserin only partly negatively interacted with diazepam. The blonanserin-potentiation was not reversed by flumazenil. Blonanserin is not a benzodiazepine receptor agonist. 7. The various effects of neuroleptics on the GABA-current may be related to the clinical effects including modifying the seizure threshold.
Yokota, Kenjiro; Tatebayashi, Hideharu; Matsuo, Tadashi; Shoge, Takashi; Motomura, Haruhiko; Matsuno, Toshiyuki; Fukuda, Akira; Tashiro, Nobutada
Several neuroleptics inhibited the 3 μM γ-aminobutyric acid induced-chloride current (GABA-current) on dissociated rat dorsal root ganglion neurons in whole-cell patch-clamp investigations. The IC50 for clozapine, zotepine, olanzapine, risperidone and chlorpromazine were 6.95, 18.26, 20.30, 106.01 and 114.56 μM, respectively. The values for the inhibitory effects of neuroleptics on the GABA (3 μM)-current, which were calculated by the fitting Hill's equations where the concentrations represent the mean therapeutic blood concentrations, were ranked clozapine>zotepine>chlorpromazine>olanzapine>risperidone. These inhibitory effects, weighted with the therapeutic concentrations of neuroleptics, were correlated with the clinical incidences of seizure during treatment with neuroleptics. Clozapine reduced the picrotoxin-inhibiton, and may compete with a ligand of the t-butylbicyclophosphorothionate (TBPS) binding site. Haloperidol and quetiapine did not affect the peak amplitude of the GABA (3 μM)-current. However, haloperidol reduced the clozapine-inhibition, and may antagonize ligand binding to TBPS binding site. Neuroleptics including haloperidol and quetiapine enhanced the desensitization of the GABA (3 μM)-current. However, haloperidol and quetiapine at 100 μM inhibited the desensitization at the beginning of application. Blonanserin (AD-5423) at 30 and 50 μM potentiated the GABA (3 μM)-current to 170.1±6.9 and 192.0±10.6% of the control current, respectively. Blonanserin shifted GABA concentration-response curve leftward. Blonanserin only partly negatively interacted with diazepam. The blonanserin-potentiation was not reversed by flumazenil. Blonanserin is not a benzodiazepine receptor agonist. The various effects of neuroleptics on the GABA-current may be related to the clinical effects including modifying the seizure threshold. PMID:11906969
Mika, J; Rojewska, E; Makuch, W; Przewlocka, B
A role of neuropeptides in neuropathic pain development has been implicated; however, the neuroimmune interactions that are involved in the underlying mechanisms may be more important than previously thought. To examine a potential role of relations between glia cells and neuropeptides in neuropathic pain, we performed competitive reverse-transcription polymerase chain reaction (RT-PCR) from the dorsal lumbar spinal cord and the dorsal root ganglion (DRG) after chronic constriction injury (CCI) in the rat sciatic nerve. The RT-PCR results indicated that complement component 1, q subcomponent (C1q) mRNA expression was higher than glial fibrillary acidic protein (GFAP) in the spinal cord 3 and 7 days post-CCI, suggesting that spinal microglia and perivascular macrophages are more activated than astrocytes. In parallel, we observed a strong upregulation of prodynorphin mRNA in the spinal cord after CCI, with no changes in the expression of proenkephalin or pronociceptin. Conversely, the expression of GFAP mRNA in the DRG was higher than C1q, which suggests that the satellite cells are activated shortly after injury, followed by the macrophages and polymorphonuclear leukocytes infiltrating the DRG. In the DRG, we also observed a very strong upregulation of prodynorphin (1387%) as well as pronociceptin (122%) and a downregulation of proenkephalin (47%) mRNAs. Interestingly, preemptive and repeated i.p. injection of minocycline reversed the activation of microglia/macrophages in the spinal cord and the trafficking of peripheral immune cells into the DRG, and markedly diminished the upregulation of prodynorphin and pronociceptin in the DRG. We thus provide novel findings that inhibition of C1q-positive cells by minocycline can diminish injury-induced neuropeptide changes in the DRG. This suggests that immune cells-derived pronociceptive factors may influence opioid peptide expression. Therefore, the injury-induced activation of microglia and leukocytes and the subsequent
Yoshikawa, Masaaki; Masuda, Tomoyuki; Kobayashi, Azusa; Senzaki, Kouji; Ozaki, Shigeru; Aizawa, Shin; Shiga, Takashi
The runt-related transcription factor Runx1 regulates cell-type specification and axonal projections of nociceptive dorsal root ganglion (DRG) neurons, whereas bone morphogenetic protein 4 (BMP4) is required for axonal growth during neuronal development. Although Runx1 has been shown to be involved in BMP4 signaling in non-neural tissues, the Runx1 function in BMP4-dependent regulation of neuronal development is unclear. To investigate interactions between Runx1 and BMP4 in neurite outgrowth, we cultured DRGs from wild-type and Runx1-deficient mouse embryos in the presence or absence of BMP4. Neurite outgrowth was decreased in BMP4-treated wild-type DRGs and untreated Runx1-deficient DRGs, suggesting the inhibitory effect of BMP4 and facilitatory effect of Runx1 on neurite outgrowth. In addition, the combination of BMP4 treatment and Runx1 deficiency increased neurite outgrowth, suggesting that Runx1 is required for BMP4-induced suppression of neurite outgrowth and that the loss of Runx1 results in a functional switch of BMP4 from neurite growth suppressing to neurite growth promoting. Both BMP4 treatment and Runx1 deficiency increased calcitonin gene-related peptide (CGRP)-positive neurons, and CGRP expression was not increased by BMP4 treatment in Runx1-deficient mice, suggesting that Runx1 contributes to BMP4-induced CGRP expression in DRG neurons. Thus, Runx1 contributes to BMP4 regulation of neurite outgrowth and CGRP expression in DRG and may control BMP4 functional switching during embryogenesis.
Aparicio, J G; Hopp, H; Choi, A; Mandayam Comar, J; Liao, V C; Harutyunyan, N; Lee, T C
Human retinal ganglion cells (RGCs) derived from pluripotent stem cells (PSCs) have anticipated value for human disease study, drug screening, and therapeutic applications; however, their full potential remains underdeveloped. To characterize RGCs in human embryonic stem cell (hESC) derived retinal organoids we examined RGC markers and surface antigen expression and made comparisons to human fetal retina. RGCs in both tissues exhibited CD184 and CD171 expression and distinct expression patterns of the RGC markers BRN3 and RBPMS. The retinal progenitor cells (RPCs) of retinal organoids expressed CD184, consistent with its expression in the neuroblastic layer in fetal retina. In retinal organoids CD184 expression was enhanced in RGC competent RPCs and high CD184 expression was retained on post-mitotic RGC precursors; CD171 was detected on maturing RGCs. The differential expression timing of CD184 and CD171 permits identification and enrichment of RGCs from retinal organoids at differing maturation states from committed progenitors to differentiating neurons. These observations will facilitate molecular characterization of PSC-derived RGCs during differentiation, critical knowledge for establishing the veracity of these in vitro produced cells. Furthermore, observations made in the retinal organoid model closely parallel those in human fetal retina further validating use of retinal organoid to model early retinal development.
Zhang, Xiulin; Beckel, Jonathan M; Daugherty, Stephanie L; Wang, Ting; Woodcock, Stephen R; Freeman, Bruce A; de Groat, William C
Effects of nitro-oleic acid (OA-NO2) on TRP channels were examined in guinea-pig dissociated dorsal root ganglia (DRG) neurons using calcium imaging and patch clamp techniques. OA-NO2 increased intracellular Ca2+ in 60–80% DRG neurons. 1-Oleoyl-2acetyl-sn-glycerol (OAG), a TRPC agonist, elicited responses in 36% of OA-NO2-sensitive neurons while capsaicin (TRPV1 agonist) or allyl-isothiocyanate (AITC, TRPA1 agonist) elicited responses in only 16% and 10%, respectively, of these neurons. A TRPV1 antagonist (diarylpiperazine, 5 μm) in combination with a TRPA1 antagonist (HC-030031, 30 μm) did not change the amplitude of the Ca2+ transients or percentage of neurons responding to OA-NO2; however, a reducing agent DTT (50 mm) or La3+ (50 μm) completely abolished OA-NO2 responses. OA-NO2 also induced a transient inward current associated with a membrane depolarization followed by a prolonged outward current and hyperpolarization in 80% of neurons. The reversal potentials of inward and outward currents were approximately −20 mV and −60 mV, respectively. Inward current was reduced when extracellular Na+ was absent, but unchanged by niflumic acid (100 μm), a Cl− channel blocker. Outward current was abolished in the absence of extracellular Ca2+ or a combination of two Ca2+-activated K+ channel blockers (iberiotoxin, 100 nm and apamin, 1 μm). BTP2 (1 or 10 μm), a broad spectrum TRPC antagonist, or La3+ (50 μm) completely abolished OA-NO2 currents. RT-PCR performed on mRNA extracted from DRGs revealed the expression of all seven subtypes of TRPC channels. These results support the hypothesis that OA-NO2 activates TRPC channels other than the TRPV1 and TRPA1 channels already known to be targets in rat and mouse sensory neurons and challenge the prevailing view that electrophilic compounds act specifically on TRPA1 or TRPV1 channels. The modulation of sensory neuron excitability via actions on multiple TRP channels can contribute to the anti-inflammatory effect
Wang, Li-Jie; Wang, Yang; Chen, Meng-Jie; Tian, Zhen-Pu; Lu, Bi-Han; Mao, Ke-Tao; Zhang, Liang; Zhao, Lei; Shan, Li-Ya; Li, Li; Si, Jun-Qiang
Niflumic acid (NFA) is a type of non-steroidal anti-inflammatory drug. Neuropathic pain is caused by a decrease in presynaptic inhibition mediated by γ-aminobutyric acid (GABA). In the present study, a whole-cell patch-clamp technique and intracellular recording were used to assess the effect of NFA on GABA-induced inward current in dorsal root ganglion (DRG) neurons of a chronic constriction injury (CCI) model. It was observed that 1-1,000 µmol/l GABA induced a concentration-dependent inward current in DRG neurons. Compared with pseudo-operated rats, the thermal withdrawal latency (TWL) of CCI rats significantly decreased (P<0.01); however, the TWLs of each NFA group (50 and 300 µmol/l) were significantly longer than that of the CCI group (P<0.01). In the CCI group, the response evoked by GABA (10(-6)-10(-3) mol/l) was reduced in a concentration dependent manner compared with a normal control group (P<0.01), and the current amplitudes of CCI rats activated by the same concentrations of GABA (10(-6)-10(-3) mol/l) were significantly decreased compared with the control group (P<0.05). The inward currents activated by 100 µmol/l GABA were suppressed by treatment with 1, 10 and 100 µmol/l NFA (5.32±3.51, 33.8±5.20, and 52.2±6.32%, respectively; P<0.05). The inverse potentials of GABA-induced currents were 9.87±1.32 and 9.64±1.24 mV with and without NFA, respectively (P<0.05). Pre-treatment with NFA exerted a strong inhibitory effect on the peak value of GABA-induced current, and the GABA-induced response was inhibited by the same concentrations of NFA (1, 10 and 100 µmol/l) in the control and CCI groups (P<0.05). The results suggest that NFA reduced the primary afferent depolarization (PAD) associated with neuropathic pain and mediated by the GABAA receptor. NFA may regulate neuropathic pain by inhibiting dorsal root reflexes, which are triggered PAD.
Wang, Li-Jie; Wang, Yang; Chen, Meng-Jie; Tian, Zhen-Pu; Lu, Bi-Han; Mao, Ke-Tao; Zhang, Liang; Zhao, Lei; Shan, Li-Ya; Li, Li; Si, Jun-Qiang
Niflumic acid (NFA) is a type of non-steroidal anti-inflammatory drug. Neuropathic pain is caused by a decrease in presynaptic inhibition mediated by γ-aminobutyric acid (GABA). In the present study, a whole-cell patch-clamp technique and intracellular recording were used to assess the effect of NFA on GABA-induced inward current in dorsal root ganglion (DRG) neurons of a chronic constriction injury (CCI) model. It was observed that 1–1,000 µmol/l GABA induced a concentration-dependent inward current in DRG neurons. Compared with pseudo-operated rats, the thermal withdrawal latency (TWL) of CCI rats significantly decreased (P<0.01); however, the TWLs of each NFA group (50 and 300 µmol/l) were significantly longer than that of the CCI group (P<0.01). In the CCI group, the response evoked by GABA (10−6-10−3 mol/l) was reduced in a concentration dependent manner compared with a normal control group (P<0.01), and the current amplitudes of CCI rats activated by the same concentrations of GABA (10−6-10−3 mol/l) were significantly decreased compared with the control group (P<0.05). The inward currents activated by 100 µmol/l GABA were suppressed by treatment with 1, 10 and 100 µmol/l NFA (5.32±3.51, 33.8±5.20, and 52.2±6.32%, respectively; P<0.05). The inverse potentials of GABA-induced currents were 9.87±1.32 and 9.64±1.24 mV with and without NFA, respectively (P<0.05). Pre-treatment with NFA exerted a strong inhibitory effect on the peak value of GABA-induced current, and the GABA-induced response was inhibited by the same concentrations of NFA (1, 10 and 100 µmol/l) in the control and CCI groups (P<0.05). The results suggest that NFA reduced the primary afferent depolarization (PAD) associated with neuropathic pain and mediated by the GABAA receptor. NFA may regulate neuropathic pain by inhibiting dorsal root reflexes, which are triggered PAD. PMID:28810599
Zhang, Xin-Sheng; Li, Xiao; Luo, Hai-Jie; Huang, Zhu-Xi; Liu, Cui-Cui; Wan, Qing; Xu, Shu-Wei; Wu, Shao-Ling; Ke, Song-Jian; Ma, Chao
Clinically, chronic low back pain and sciatica associated with lumbar disc herniation (LDH) is a common musculoskeletal disorder. Due to the unawareness of detailed mechanisms, it is difficult to get an effective therapy. The aim of the present study was to identify the role of the RAGE/STAT3 pathway in the dorsal root ganglion (DRG) on the formation and development of persistent pain hypersensitivity induced by LDH. Controlled animal study. University laboratory. After LDH induced by implantation of autologous nucleus pulposus (NP, harvested from animal tail) on the left L5 nerve root was established, mechanical thresholds and electrophysiological tests were conducted at relevant time points during an observation period of 28 days. Protein levels and localization of RAGE and p-STAT3 were performed by using Western blotting and immunohistochemistry, respectively. LDH induced persistent pain hypersensitivity, increased excitability of DRG neurons, and upregulated the expression of RAGE and p-STAT3 in the DRG. Consecutive injection of both RAGE antagonist FPS-ZM1 (i.t.) and STAT3 activity inhibitor S3I-201 (i.t.) inhibited the enhanced excitability of DRG neurons and mechanical allodynia induced by NP implantation. Furthermore, local knockdown of STAT3 by intrathecal injection of AAV-Cre-GFP into STAT3flox/flox mice markedly alleviated NP implantation-induced mechanical allodynia in mice. Importantly, the expression of p-STAT3 was colocalized with that of RAGE in the DRG and inhibition of RAGE with FPS-ZM1 prevented NP implantation-induced STAT3 activation. More underlying mechanism(s) of the role of the RAGE/STAT3 pathway on the formation and development of persistent pain hypersensitivity induced by LDH will be needed to be explored in future research. These findings suggest activation of the RAGE/STAT3 pathway plays a critical role in persistent pain induced by LDH, and this pathway may represent novel therapeutic targets for the treatment of LDH
Knezevic, Nebojsa Nick; Candido, Kenneth D; Cokic, Ivan; Krbanjevic, Aleksandar; Berth, Sarah L; Knezevic, Ivana
Epidural and intrathecal injections of methylprednisolone acetate (MPA) have become the most commonly performed interventional procedures in the United States and worldwide in the last 2 decades. However neuraxial MPA injection has been dogged by controversy regarding the presence of different additives used in commercially prepared glucocorticoids. We previously showed that MPA could be rendered 85% free of polyethylene glycol (PEG) by a simple physical separation of elements in the suspension. The objective of the present study was to explore a possible cytotoxic effect of commercially available MPA (with intact or reduced preservatives) on rat sensory neurons. We exposed primary dissociated rat dorsal root ganglia (DRG) sensory neurons to commercially available MPA for 24 hours with either the standard (commercial) concentration of preservatives or to different fractions following separation (MPA suspension whose preservative concentration had been reduced, or fractions containing higher concentrations of preservatives). Cells were stained with the TUNEL assay kit to detect apoptotic cells and images were taken on the Bio-Rad Laser Sharp-2000 system. We also detected expression of caspase-3, as an indicator of apoptosis in cell lysates. We exposed sensory neurons from rat DRG to different concentrations of MPA from the original commercially prepared vial. TUNEL assay showed dose-related responses and increased percentages of apoptotic cells with increasing concentrations of MPA. Increased concentrations of MPA caused 1.5 - 2 times higher caspase-3 expression in DRG sensory neurons than in control cells (ANOVA, P = 0.001). Our results showed that MPA with reduced preservatives caused significantly less apoptosis observed with TUNEL assay labeling (P < 0.001) and caspase-3 immunoblotting (P = 0.001) than in neurons exposed to MPA from a commercially prepared vial or "clear phase" that contained higher concentrations of preservatives. Even though MPA with reduced
Erriquez, Jessica; Bernascone, Silvia; Ciarletta, Monica; Filigheddu, Nicoletta; Graziani, Andrea; Distasi, Carla
Ghrelin is a hormone regulating energy homeostasis via interaction with its receptor, GHSR-1a. Ghrelin activities in dorsal root ganglia (DRG) cells are unknown. Herein we show that ghrelin induces a change of cytosolic calcium concentration in both glia and neurons of embryonic chick DRG. Both RT-PCR and binding studies performed with fluorescent ghrelin in the presence of either unlabeled ghrelin or GHSR-1a antagonist D-Lys(3)-GHRP-6, indicate that DRG cells express GHSR-1a. In glial cells the response is characterized by a rapid transient rise in [Ca(2+)](i) followed by a long lasting rise. The calcium elevation is dependent on calcium release from thapsigargin-sensitive intracellular stores and on activation of two distinct Ca(2+) entry pathways, a receptor activated calcium entry and a store operated calcium entry. Surprisingly, D-Lys(3)-GHRP-6 exerts several activities in the absence of exogenous ghrelin: (i) it activates calcium release from thapsigargin-sensitive intracellular stores and calcium entry via voltage-operated channels in non-neuronal cells; (ii) it inhibits calcium oscillations in non-neuronal cells exhibiting spontaneous Ca(2+) activity and iii) it promotes apoptosis of DRG cells, both neurons and glia. In summary, we provide the first evidence for ghrelin activity in DRG, and we also demonstrate that the widely used D-Lys(3)-GHRP-6 ghrelin antagonist features ghrelin independent activities.
Slappendel, R; Crul, B J; Braak, G J; Geurts, J W; Booij, L H; Voerman, V F; de Boo, T
The efficacy of radiofrequency lesion treatment of the cervical dorsal root ganglion (RF-DRG) in cervicobrachialgia was investigated in 61 patients by a randomized prospective double blinded study. Before lesion treatment the putative pain provoking spinal root was identified by diagnostic blocks with a local anesthetic agent. One group of patients (n = 32, group I) was treated with a radiofrequency lesion of 67 degrees C and in a control group (n = 29, group II) a temperature of 40 degrees C was applied. Three months after treatment a significant reduction in VAS scores was demonstrated in both groups. The outcome of the treatments was identical (VAS reduction: group I, 1.7; group II, 1.9; P = 0.001). In group I a VAS reduction of 3 or more occurred in 11/31 (34%) and in group II in 11/29 (38%) of patients. A VAS reduction of 2 or more occurred in group I in 15/31 (47%) and in group II in 15/29 (51%) of patients. This study suggests that treatment with 40 degrees C radiofrequency application of the dorsal root ganglion is equally effective as treatment at 67 degrees C. Further appraisal of this treatment is required.
Özdemir, Ümit Sinan; Nazıroğlu, Mustafa; Şenol, Nilgün; Ghazizadeh, Vahid
Oxidative stress and cytosolic Ca(2+) overload have important roles on apoptosis in dorsal root ganglion (DRG) neurons after spinal cord injury (SCI). Hypericum perforatum (HP) has an antioxidant property in the DRGs due to its ability to modulate NADPH oxidase and protein kinase C pathways. We aimed to investigate the protective property of HP on oxidative stress, apoptosis, and Ca(2+) entry through transient receptor potential melastatin 2 (TRPM2) and transient receptor potential vanilloid 1 (TRPV1) channels in SCI-induced DRG neurons of rats. Rats were divided into four groups as control, HP, SCI, and SCI + HP. The HP groups received 30 mg/kg HP for three concessive days after SCI induction. The SCI-induced TRPM2 and TRPV1 currents and cytosolic free Ca(2+) concentration were reduced by HP. The SCI-induced decrease in glutathione peroxidase and cell viability values were ameliorated by HP treatment, and the SCI-induced increase in apoptosis, caspase 3, caspase 9, cytosolic reactive oxygen species (ROS) production, and mitochondrial membrane depolarization values in DRG of SCI group were overcome by HP treatment. In conclusion, we observed a protective role of HP on SCI-induced oxidative stress, apoptosis, and Ca(2+) entry through TRPM2 and TRPV1 in the DRG neurons. Our findings may be relevant to the etiology and treatment of SCI by HP. Graphical Abstract Possible molecular pathways of involvement of Hypericum perforatum (HP) on apoptosis, oxidative stress, and calcium accumulation through TRPM2 and TRPV1 channels in DRG neurons of SCI-induced rats. The TRPM2 channel is activated by ADP-ribose and oxidative stress through activation of ADP-ribose pyrophosphate although it was inhibited by N-(p-amylcinnamoyl) anthranilic acid (ACA) and 2-aminoethyl diphenylborinate (2APB). The TRPV1 channel is activated by oxidative stress and capsaicin and it is blocked by capsazepine. Injury in the DRG can result in augmented ROS release, leading to Ca(2+) uptake through
Ghazizadeh, Vahid; Nazıroğlu, Mustafa
Incidence rates of epilepsy and use of Wi-Fi worldwide have been increasing. TRPV1 is a Ca(2+) permeable and non-selective channel, gated by noxious heat, oxidative stress and capsaicin (CAP). The hyperthermia and oxidant effects of Wi-Fi may induce apoptosis and Ca(2+) entry through activation of TRPV1 channel in epilepsy. Therefore, we tested the effects of Wi-Fi (2.45 GHz) exposure on Ca(2+) influx, oxidative stress and apoptosis through TRPV1 channel in the murine dorsal root ganglion (DRG) and hippocampus of pentylentetrazol (PTZ)-induced epileptic rats. Rats in the present study were divided into two groups as controls and PTZ. The PTZ groups were divided into two subgroups namely PTZ + Wi-Fi and PTZ + Wi-Fi + capsazepine (CPZ). The hippocampal and DRG neurons were freshly isolated from the rats. The DRG and hippocampus in PTZ + Wi-Fi and PTZ + Wi-Fi + CPZ groups were exposed to Wi-Fi for 1 hour before CAP stimulation. The cytosolic free Ca(2+), reactive oxygen species production, apoptosis, mitochondrial membrane depolarization, caspase-3 and -9 values in hippocampus were higher in the PTZ group than in the control although cell viability values decreased. The Wi-Fi exposure induced additional effects on the cytosolic Ca(2+) increase. However, pretreatment of the neurons with CPZ, results in a protection against epilepsy-induced Ca(2+) influx, apoptosis and oxidative damages. In results of whole cell patch-clamp experiments, treatment of DRG with Ca(2+) channel antagonists [thapsigargin, verapamil + diltiazem, 2-APB, MK-801] indicated that Wi-Fi exposure induced Ca(2+) influx via the TRPV1 channels. In conclusion, epilepsy and Wi-Fi in our experimental model is involved in Ca(2+) influx and oxidative stress-induced hippocampal and DRG death through activation of TRPV1 channels, and negative modulation of this channel activity by CPZ pretreatment may account for the neuroprotective activity against oxidative stress.
Despite considerable research, the mechanisms of neuropathic pain induced by excessive oxidative stress production and overload calcium ion (Ca(2+)) entry in dorsal root ganglion (DRG) remain substantially unidentified. The transient receptor potential melastatin 2 (TRPM2) and vanilloid 1 (TRPV1) channels are activated with different stimuli including oxidative stress. TRPM2 and TRPV1 have been shown to be involved in induction of neuropathic pain. However, the activation mechanisms of TRPM2 and TRPV1 via NADPH oxidase and protein kinase C (PKC) pathways are poorly understood. In this study, I investigated the roles of NADPH oxidase and PKC on Ca(2+) entry through TRPM2 and TRPV1 channels in in vitro DRG neurons of rats. Rat DRG neurons were used in whole-cell patch clamp experiments. The H2O2-induced TRPM2 current densities were decreased by N-(p-amylcinnamoyl)anthranilic acid (ACA), and dose-dependent capsaicin (CAP) and H2O2-induced TRPV1 currents were inhibited by capsazepine (CPZ). The TRPV1 channel is activated in the DRG neurons by 0.01 mM capsaicin but not 0.001 mM or 0.05 mM capsaicin. TRPM2 and TRPV1 currents were increased by the PKC activator, phorbol myristate acetate (PMA), although the currents were decreased by ACA, CPZ, and the PKC inhibitor, bisindolylmaleimide I (BIM). Both channel currents were further increased by PMA + H2O2 as compared to H2O2 only. In the combined presence of PMA + BIM, no TRPM2 or TRPV1 currents were observed. The CAP and H2O2-induced TRPM2 current densities were also decreased by the NADPH oxidase inhibitors apocynin and N-Acetylcysteine. In conclusion, these results demonstrate a protective role for NADPH oxidase and PKC inhibitors on Ca(2+) entry through TRPM2 and TRPV1 channels in DRG neurons. Since excessive oxidative stress production and Ca(2+) entry are implicated in the pathophysiology of neuropathic pain, the findings may be relevant to the etiology and treatment of neuropathology in DRG neurons.
Walwyn, Wendy; Evans, Christopher J; Hales, Tim G
Beta-arrestins bind to agonist-activated G-protein-coupled receptors regulating signaling events and initiating endocytosis. In beta-arrestin2-/- (beta arr2-/-) mice, a complex phenotype is observed that includes altered sensitivity to morphine. However, little is known of how beta-arrestin2 affects mu receptor signaling. We investigated the coupling of mu receptors to voltage-gated Ca2+ channels (VGCCs) in beta arr2+/+ and beta arr2-/- dorsal root ganglion neurons. A lack of beta-arrestin2 reduced the maximum inhibition of VGCCs by morphine and DAMGO (D-Ala2-N-Me-Phe4-glycol5-enkephalin) without affecting agonist potency, the onset of receptor desensitization, or the functional contribution of N-type VGCCs. The reduction in inhibition was accompanied by increased naltrexone-sensitive constitutive inhibitory coupling of mu receptors to VGCCs. Agonist-independent mu receptor inhibitory coupling was insensitive to CTAP (Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2), a neutral antagonist that inhibited the inverse agonist action of naltrexone. These functional changes were accompanied by diminished constitutive recycling and increased cell-surface mu receptor expression in beta arr2-/- compared with beta arr2+/+ neurons. Such changes could not be explained by the classical role of beta-arrestins in agonist-induced endocytosis. The localization of the nonreceptor tyrosine kinase c-Src appeared disrupted in beta arr2-/- neurons, and there was reduced activation of c-Src by DAMGO. Using the Src inhibitor PP2 [4-amino-5-(4-chlorophenyl)-(t-butyl)pyrazolo[3,4-d]pyrimidine], we demonstrated that defective Src signaling mimics the beta arr2-/- cellular phenotype of reduced mu agonist efficacy, increased constitutive mu receptor activity, and reduced constitutive recycling. We propose that beta-arrestin2 is required to target c-Src to constitutively active mu receptors, resulting in their internalization, providing another dimension to the complex role of beta-arrestin2 and c-Src in G
Van Zundert, J; Lamé, I E; de Louw, A; Jansen, J; Kessels, F; Patijn, J; van Kleef, M
Cervicogenic headache and cervicobrachialgia are frequent diagnoses of chronic cervical pain. After failure of conservative treatment, an interventional approach may be indicated in the absence of any indication for causal surgical treatment. The pulsed radiofrequency (PRF) technique exposes the nerve to a high-frequency electric field while the temperature of the electrode tip does not exceed 42°C. This method is thought to be nondestructive and almost free of neurologic side effects and complications. Our extended pilot study was performed to confirm the perceived efficacy of PRF for short- and long-term relief of chronic cervical pain. We carried out a clinical audit of the first 18 patients treated with PRF at the cervical dorsal root ganglion. An independent evaluator reviewed the medical records. Patients with good clinical results at 8 weeks were evaluated for long-term effect (> 6 months), based on a 7-point Likert scale. Thirteen patients (72%) showed short-term clinical success (≥ 50% pain relief). Mean follow-up was 19.4 months (SD 8.9 months), maximum 2.5 years. The duration of satisfactory pain relief (6 or 7 on the Likert scale) varied between 2 and over 30 months, with a mean duration of 9.2 months (SD 11.2 months). Kaplan-Meier analysis illustrated that 50% of patients experienced success 3 months after treatment. We could not identify predictive variables for clinical outcome. None of the patients reported post-treatment neuritis or other adverse events. To our knowledge, this is the first documented series of chronic cervical pain syndromes treated with PRF. Satisfactory pain relief of at least 50% was achieved in 13 of 18 (72%) patients at 8 weeks. More than one year after treatment, six patients (33%) continue to rate treatment outcome as good or very good. No side effects were reported. j.
Chen, Yong; Li, Guangwen; Huang, Li-Yen Mae
The purinergic P2X3 receptor (P2X3R) expressed in the dorsal root ganglion (DRG) sensory neuron and the P2X7 receptor (P2X7R) expressed in the surrounding satellite glial cell (SGC) are two major receptors participating in neuron-SGC communication in adult DRGs. Activation of P2X7Rs was found to tonically reduce the expression of P2X3Rs in DRGs, thus inhibiting the abnormal pain behaviors in adult rats. P2X receptors are also actively involved in sensory signaling in developing rodents. However, very little is known about the developmental change of P2X7Rs in DRGs and the interaction between P2X7Rs and P2X3Rs in those animals. We therefore examined the expression of P2X3Rs and P2X7Rs in postnatal rats and determined if P2X7R-P2X3R control exists in developing rats. We immunostained DRGs of immature rats and found that P2X3Rs were expressed only in neurons and P2X7Rs were expressed only in SGCs. Western blot analyses indicated that P2X3R expression decreased while P2X7R expression increased with the age of rats. Electrophysiological studies showed that the number of DRG neurons responding to the stimulation of the P2XR agonist, α,β-meATP, was higher and the amplitudes of α,β-meATP-induced depolarizations were larger in immature DRG neurons. As a result, P2X3R-mediated flinching responses were much more pronounced in immature rats than those found in adult rats. When we reduced P2X7R expression with P2X7R-siRNA in postnatal and adult rats, P2X3R-mediated flinch responses were greatly enhanced in both rat populations. These results show that the P2X7R expression increases as rats age. In addition, P2X7Rs in SGCs exert inhibitory control on the P2X3R expression and function in sensory neurons of immature rats, just as observed in adult rats. Regulation of P2X7R expression is likely an effective way to control P2X3R activity and manage pain relief in infants.
Yang, Y; Wu, H; Yan, J-Q; Song, Z-B; Guo, Q-L
Both tumor necrosis factor (TNF)-α and the angiotensin (Ang) II/angiotensin II receptor type 1 (AT1) axis play important roles in neuropathic pain and nociception. In the present study, we explored the interaction between the two systems by examining the mutual effects between TNF-α and the Ang II/AT1 receptor axis in dorsal root ganglion (DRG) neurons. Rat DRG neurons were treated with TNF-α in different concentrations for different lengths of time in the presence or absence of transcription inhibitor actinomycin D, TNF receptor 1 (TNFR1) inhibitor SPD304, β-catenin signaling inhibitor CCT031374, or different kinase inhibitors. TNF-α decreased the AT1 receptor mRNA level as well as the AT1a receptor promoter activity in a dose-dependent manner within 30 h, which led to dose-dependent inhibition of Ang II-binding AT1 receptor level on the cell membrane. Actinomycin D (1 mg/ml), SPD304 (50 μM), p38 mitogen-activated protein kinase (MAPK) inhibitor PD169316 (25 μM), and CCT031374 (50 μM) completely abolished the inhibitory effect of TNF-α on AT1 receptor expression. TNF-α dose-dependently increased soluble β-catenin and phosphorylated GSK-3β levels, which was blocked by SPD304 and PD169316. In DRG neurons treated with AT2 receptor agonist CGP421140, or Ang II with or without AT1 receptor antagonist losartan or AT2 receptor antagonist PD123319 for 30 h, we found that Ang II and Ang II+PD123319 significantly decreased TNF-α expression, whereas CPG421140 and Ang II+losartan increased TNF-α expression. In conclusion, we demonstrate that TNF-α inhibits AT1 receptor expression at the transcription level via TNFR1 in rat DRG neurons by increasing the soluble β-catenin level through the p38 MAPK/GSK-3β pathway. In addition, Ang II appears to inhibit and induce TNF-α expression via the AT1 receptor and the AT2 receptor in DRG neurons, respectively. This is the first evidence of crosstalk between TNF-α and the Ang II/AT receptor axis in DRG neurons.
Sculptoreanu, A; Kullmann, F A; Artim, D E; Bazley, F A; Schopfer, F; Woodcock, S; Freeman, B A; de Groat, W C
Nitro-oleic acid (OA-NO(2)), an electrophilic fatty acid by-product of nitric oxide and nitrite reactions, is present in normal and inflamed mammalian tissues at up to micromolar concentrations and exhibits anti-inflammatory signaling actions. The effects of OA-NO(2) on cultured dorsal root ganglion (DRG) neurons were examined using fura-2 Ca(2+) imaging and patch clamping. OA-NO(2) (3.5-35 microM) elicited Ca(2+) transients in 20 to 40% of DRG neurons, the majority (60-80%) of which also responded to allyl isothiocyanate (AITC; 1-50 microM), a TRPA1 agonist, and to capsaicin (CAPS; 0.5 microM), a TRPV1 agonist. The OA-NO(2)-evoked Ca(2+) transients were reduced by the TRPA1 antagonist 2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(4-isopropylphenyl) acetamide (HC-030031; 5-50 microM) and the TRPV1 antagonist capsazepine (10 microM). Patch-clamp recording revealed that OA-NO(2) depolarized and induced inward currents in 62% of neurons. The effects of OA-NO(2) were elicited by concentrations >or=5 nM and were blocked by 10 mM dithiothreitol. Concentrations of OA-NO(2) >or=5 nM reduced action potential (AP) overshoot, increased AP duration, inhibited firing induced by depolarizing current pulses, and inhibited Na(+) currents. The effects of OA-NO(2) were not prevented or reversed by the NO-scavenger carboxy-2-phenyl-4,4,5,5-tetramethylimidazolineoxyl-1-oxyl-3-oxide. A large percentage (46-57%) of OA-NO(2)-responsive neurons also responded to CAPS (0.5 microM) or AITC (0.5 microM). OA-NO(2) currents were reduced by TRPV1 (diarylpiperazine; 5 microM) or TRPA1 (HC-030031; 5 microM) antagonists. These data reveal that endogenous OA-NO(2) generated at sites of inflammation may initially activate transient receptor potential channels on nociceptive afferent nerves, contributing to the initiation of afferent nerve activity, and later suppresses afferent firing.
Shanthanna, Harsha; Chan, Philip; McChesney, James; Thabane, Lehana; Paul, James
Background No proof of efficacy, in the form of a randomized controlled trial (RCT), exists to support pulsed radiofrequency (PRF) treatment of the dorsal root ganglion (DRG) for chronic lumbar radicular (CLR) pain. We determined the feasibility of a larger trial (primary objective), and also explored the efficacy of PRF in decreasing pain on a visual analog scale (VAS) and improving the Oswestry Disability Index. Methods This was a single-center, placebo-controlled, triple-blinded RCT. Patients were randomized to a placebo group (needle placement) or a treatment group (PRF at 42°C for 120 seconds to the DRG). Patients were followed up for 3 months post procedure. Outcomes with regard to pain, Oswestry Disability Index score, and side effects were analyzed on an intention-to-treat basis. Results Over 15 months, 350 potential patients were identified and 56 were assessed for eligibility. Fifteen of them did not meet the selection criteria. Of the 41 eligible patients, 32 (78%) were recruited. One patient opted out before intervention. Three patients were lost to follow-up at 3 months. Mean VAS differences were not significantly different at 4 weeks (−0.36, 95% confidence interval [CI], −2.29, 1.57) or at 3 months (−0.76, 95% CI, −3.14, 1.61). The difference in mean Oswestry Disability Index score was also not significantly different at 4 weeks (−2%, 95% CI, −14%, 10%) or 3 months (−7%, 95% CI, −21%, 6%). There were no major side effects. Six of 16 patients in the PRF group and three of 15 in the placebo group showed a >50% decrease in VAS score. Conclusion The recruitment rate was partially successful. At 3 months, the relative success of PRF-DRG was small. A large-scale trial to establish efficacy is not practically feasible considering the small effect size, which would necessitate recruitment of a challengingly large number of participants over a number of years. Until clear parameters for application of PRF are established, clinicians will need
Pei, L; Zhang, J; Zhao, F; Su, T; Wei, H; Tian, J; Li, M; Shi, J
Annexin 1 (ANXA1) has analgesic effects in inflammatory pain. We aimed to investigate the anti-nociceptive role of ANXA1, at the dorsal root ganglion (DRG) level, through an interaction with formyl-peptide-receptor-like 1 (FPR2/ALX). Inflammatory pain was evoked by injecting complete Freund's adjuvant (CFA, 50 μl) into the hindpaw of male Sprague-Dawley rats. The distribution of ANXA1 and FPR2/ALX in L4/5 DRGs was evaluated by immunofluorescence. The expression of ANXA1 was measured by western blot. The involvement of FPR2/ALX in the anti-nociception of ANXA1 was investigated by thermal (irradiant heat) and mechanical (von Frey filament) pain tests with intrathecal (i.t.) ANXA1-derived peptide (Anxa1(2-26)), FPR2/ALX agonist 5(S)-6(R)-7-trihydroxyheptanoic-acid-methyl-ester (BML-111), and antagonist N-t-Boc-Phe-Leu-Phe-Leu-Phe (Boc1). ANXA1 and FPR2/ALX localized in the satellite glial cells and neurones in L4/5 DRGs. CFA treatment (n=20) increased ANXA1 expression in L4/5 DRGs within 7 days (P<0.01). I.T. Anxa1(2-26) (20 and 100 µg µl(-1)) and BML-111 (10 and 100 nmol) reduced CFA-induced thermal and mechanical nociception within 48 h (n=40) (P<0.05). However, i.t. Boc1 10 µg intensified inflammatory pain (P<0.05) and reversed the anti-nociceptive effect of Anxa1(2-26) (n=25) (P<0.05). Moreover, ANXA1 expression increased in L4/5 DRGs after i.t. Anxa1(2-26) (20 µg µl(-1)) (P<0.05) and BML-111 (10 nmol) (P<0.01) but decreased after i.t. Boc1 (10 and 100 µg) alone (P<0.01) or Boc1 (10 µg) co-injection with Anxa1(2-26) (20 µg µl(-1)) (P<0.05). Endogenous ANXA1 expression at the DRG level is involved in CFA-induced inflammatory pain, and i.t. ANXA1 20 µg µl(-1) produces its anti-nociceptive effect through FPR2/ALX.
Smith, T; Al Otaibi, M; Sathish, J; Djouhri, L
A hallmark of peripheral neuropathic pain (PNP) is chronic spontaneous pain and/or hypersensitivity to normally painful stimuli (hyperalgesia) or normally nonpainful stimuli (allodynia).This pain results partly from abnormal hyperexcitability of dorsal root ganglion (DRG) neurons. We have previously shown, using a modified version of the lumbar 5 (L5)-spinal nerve ligation model of PNP (mSNA model involving L5-spinal nerve axotomy plus loose ligation of the lumbar 4 (L4)-spinal nerve with neuroinflammation-inducing chromic-gut), that L4 DRG neurons exhibit increased spontaneous activity, the key characteristic of neuronal hyperexcitability. The underlying ionic and molecular mechanisms of the hyperexcitability of L4 DRG neurons are incompletely understood, but could result from changes in expression and/or function of ion channels including hyperpolarization-activated cyclic nucleotide-gated (HCN) channels, which are active near the neuron's resting membrane potential, and which produce an excitatory inward current that depolarizes the membrane potential toward the threshold of action potential generation. Therefore, in the present study we used the mSNA model to investigate whether: (a) expression of HCN1-HCN3 channels is altered in L4 DRG neurons which, in the mSNA model, are essential for transmission of the evoked pain, and which contribute to chronic spontaneous pain, and (b) local (intraplantar) blockade of these HCN channels, with a specific blocker, ZD7288, attenuates chronic spontaneous pain and/or evoked pain in mSNA rats. We found 7days after mSNA: (1) a significant increase in HCN2-immunoreactivity in small (<30μm) DRG neurons (predominantly IB4-negative neurons), and in the proportion of small neurons expressing HCN2 (putative nociceptors); (2) no significant change in HCN1- or HCN3-immunoreactivity in all cell types; and (3) attenuation, with ZD7288 (100μM intraplantar), of chronic spontaneous pain behavior (spontaneous foot lifting) and mechanical
St-Jacques, Bruno; Ma, Weiya
Prostaglandin E2 (PGE2), a well-known pain mediator enriched in inflamed tissues, plays a pivotal role in the genesis of chronic pain conditions such as inflammatory and neuropathic pain. PGE2-prolonged sensitization of nociceptive dorsal root ganglion (DRG) neurons (nociceptors) may contribute to the transition from acute to chronic pain. However, the underlying cellular mechanisms are poorly understood. In this study, we tested the hypothesis that facilitating synthesis and anterograde axonal trafficking of EP receptors contribute to PGE2-prolonged nociceptor sensitization. Intraplantar (i.pl.) injection of a stabilized PGE2 analog, 16,16 dimethyl PGE2 (dmPGE2), in a dose- and time-dependent manner, not only elicited primary tactile allodynia which lasted for 1d, but also prolonged tactile allodynia evoked by a subsequent i.pl. injection of dmPGE2 from 1d to 4d. Moreover, the duration of tactile allodynia was progressively prolonged following multiple sequential i.pl. injections of dmPGE2. Co-injection of the selective EP1 or EP4 receptor antagonist, the inhibitors of cAMP, PKA, PKC, PKCε or PLC as well as an interleukin-6 (IL-6) neutralizing antiserum differentially blocked primary tactile allodynia elicited by the 1st dmPGE2 and the prolonged tactile allodynia evoked by the 2nd dmPGE2, suggesting the involvement of these signaling events in dmPGE2-induced nociceptor activation and sensitization. Co-injection of a selective COX2 inhibitor or two EP4 antagonists prevented or shortened inflammagen-prolonged nociceptor sensitization. I.pl. injection of dmPGE2 or carrageenan time-dependently increased EP4 levels in L4-6 DRG neurons and peripheral nerves. EP4 was expressed in almost half of IB4-binding nociceptors of L4-6 DRG. Taken together, our data suggest that stimulating the synthesis and anterograde axonal trafficking to increase EP4 availability at the axonal terminals of nociceptors is likely a novel mechanism underlying PGE2-prolonged nociceptor
Cheng, Xiaoyang; Dib-Hajj, Sulayman D; Tyrrell, Lynda; Te Morsche, Rene H; Drenth, Joost P H; Waxman, Stephen G
Gain-of-function missense mutations of voltage-gated sodium channel Na(V)1.7 have been linked to the painful disorder inherited erythromelalgia. These mutations hyperpolarize activation, slow deactivation and enhance currents evoked by slow ramp stimuli (ramp currents). A correlation has recently been suggested between the age of onset of inherited erythromelalgia and the extent of hyperpolarizing shifts in mutant Na(V)1.7 channel activation; mutations causing large activation shifts have been linked to early age of onset inherited erythromelalgia, while mutations causing small activation shifts have been linked to age of onset within the second decade of life. Here, we report a family with inherited erythromelalgia with an in-frame deletion of a single residue--leucine 955 (Del-L955) in DII/S6. The proband did not show symptoms until the age of 15 years, and her affected mother only experienced mild symptoms during adolescence, which disappeared at the age of 38 years. Del-L955 shows no effect on Na(V)1.7 current density and fast inactivation, but causes an approximately -24 mV shift in activation, together with increases in amplitude of persistent currents and ramp currents. The mutation also produces an approximately -40 mV shift in slow inactivation, which reduces channel availability. Comparison of the effects of the Del-L955 mutation on dorsal root ganglion neuron hyperexcitability with those produced by another inherited erythromelalgia mutation (L858F) that does not enhance slow inactivation suggests that a delayed age of onset and milder symptoms in association with a large shift of channel activation, enhanced persistent and enhanced ramp currents may be related to the approximately -40 mV shift in slow inactivation for Del-L955, the largest shift thus far demonstrated in mutant Na(V)1.7 channels. Our results suggest that despite the pivotal role of activation shift in inherited erythromelalgia development, slow inactivation may regulate clinical
Burnham, Patricia; Raiborn, Charles; Varon, Silvio
Nerve-growth factor is known to cause a considerable increase in the number of neurons putting out processes and surviving in cell cultures of dissociated dorsal-root and sympathetic ganglia from embryonic chicks. Similar effects of nerve-growth factor have now been noted with cultures of dissociated dorsal-root ganglia from newborn mice or rats. In all three sensory ganglionic systems, the effects of the nerve-growth factor on fiber production and neuronal survival could be mimicked, in the absence of the factor, by adequate increase of the non-neuronal cells in the cultures. The results suggest a hypothesis that views the role of the nerve-growth factor as subordinate to that of the non-neuronal cells. PMID:4509314
Bernardini, Nadia; Tomassy, Giulio Srubek; Tata, Ada Maria; Augusti-Tocco, Gabriella; Biagioni, Stefano
Spontaneous and potassium-induced acetylcholine release from embryonic (E12 and E18) chick dorsal root ganglia explants at 3 and 7 days in culture was investigated using a chemiluminescent procedure. A basal release ranging from 2.4 to 13.8 pm/ganglion/5 min was detected. Potassium application always induced a significant increase over the basal release. The acetylcholine levels measured in E12 explants were 6.3 and 38.4 pm/ganglion/5 min at 3 and 7 days in culture, respectively, while in E18 explant cultures they were 10.7 and 15.5 pm/ganglion/5 min. In experiments performed in the absence of extracellular Ca2+ ions, acetylcholine release, both basal and potassium-induced, was abolished and it was reduced by cholinergic antagonists. A morphometric analysis of explant fibre length suggested that acetylcholine release was directly correlated to neurite extension. Moreover, treatment of E12 dorsal root ganglion-dissociated cell cultures with carbachol as cholinergic receptor agonist was shown to induce a higher neurite outgrowth compared with untreated cultures. The concomitant treatment with carbachol and the antagonists at muscarinic receptors atropine and at nicotinic receptors mecamylamine counteracted the increase in fibre outgrowth. Although the present data have not established whether acetylcholine is released by neurones or glial cells, these observations provide the first evidence of a regulated release of acetylcholine in dorsal root ganglia.
Davis, J P; Epstein, M L
The plasticity of neural crest cells for the expression of adrenergic and cholinergic transmitter phenotypes has been well studied. The object of this study was to determine if cells of a sensory ganglion are capable of neuropeptide transmitter plasticity. We studied whether cells of the trigeminal ganglion, which do not express the neuropeptide vasoactive intestinal peptide (VIP) in vivo, would express this peptide when grown with a tissue the gut, that contains large numbers of VIP neurons. Embryonic aneural chick rectum was explanted with the embryonic quail trigeminal ganglion on the chorioallantoic membrane of chick hosts for 7-8 days. The explants were fixed, sectioned, and stained for VIP immunoreactivity (IR), for neurofilament protein immunoreactivity, and for the quail nucleolar marker. In sections of the explants we observed two populations of quail neurons: small (10-13 microns) VIP-IR cells and large (25-32 microns) cells lacking VIP-IR and resembling native trigeminal neurons. Trigeminal ganglia explanted with embryonic heart or trigeminal ganglia explanted alone lacked small VIP-IR cells but contained large VIP-negative neurons. These results show that cells of the trigeminal ganglion grown with the gut can express a neuropeptide they do not express in the absence of the gut or in vivo. Thus the embryonic trigeminal ganglion contains cells that are plastic with respect to neuropeptide expression.
Ahn, Hye-Sook; Vasylyev, Dmytro V; Estacion, Mark; Macala, Lawrence J; Shah, Palak; Faber, Catharina G; Merkies, Ingemar S J; Dib-Hajj, Sulayman D; Waxman, Stephen G
Sodium channel NaV1.7 is preferentially expressed in dorsal root ganglion (DRG) and sympathetic ganglion neurons. Gain-of-function NaV1.7 mutations/variants have been identified in the painful disorders inherited erythromelalgia and small-fiber neuropathy (SFN). DRG neurons transfected with these channel variants display depolarized resting potential, reduced current-threshold, increased firing-frequency and spontaneous firing. Whether the depolarizing shift in resting potential and enhanced spontaneous firing are due to persistent activity of variant channels, or to compensatory changes in other conductance(s) in response to expression of the variant channel, as shown in model systems, has not been studied. We examined the effect of wild-type NaV1.7 and a NaV1.7 mutant channel, D623N, associated with SFN, on resting potential and membrane potential during interspike intervals in DRG neurons. Resting potential in DRG neurons expressing D623N was depolarized compared to neurons expressing WT-NaV1.7. Exposure to TTX hyperpolarized resting potential by 7mV, increased current-threshold, decreased firing-frequency, and reduced NMDG-induced-hyperpolarization in DRG neurons expressing D623N. To assess the contribution of depolarized resting potential to DRG neuron excitability, we mimicked the mutant channel's depolarizing effect by current injection to produce equivalent depolarization; the depolarization decreased current threshold and increased firing-frequency. Voltage-clamp using ramp or repetitive action potentials as commands showed that D623N channels enhance the TTX-sensitive inward current, persistent at subthreshold membrane voltages, as predicted by a Hodgkin-Huxley model. Our results demonstrate that a variant of NaV1.7 associated with painful neuropathy depolarizes resting membrane potential and produces an enhanced inward current during interspike intervals, thereby contributing to DRG neuron hyperexcitability.
Bae, Jin Young; Kim, Jae Hyun; Cho, Yi Sul; Mah, Won; Bae, Yong Chul
Substance P (SP), calcitonin gene-related peptide (CGRP), and isolectin B4 (IB4) are widely used as markers for peripheral neurons with unmyelinated fibers, whereas neurofilament 200 (NF200), and Peripherin are used as markers for neurons with myelinated fibers, and with unmyelinated or small-caliber fibers, respectively. To study the selectivity of these markers for specific neuronal types, we analyzed their expression in neurons in the rat trigeminal ganglion by light- and electron-microscopic immunocytochemistry. Most SP-immunopositive (+), CGRP+, and IB4+ fibers were unmyelinated, but a small fraction (∼5%) were small myelinated fibers (<20 µm(2) in cross-sectional area, equivalent to <5 µm in diameter, Aδ fiber). Similarly, whereas the majority of NF200+ fibers were myelinated, a large fraction (23.9%) were unmyelinated, and whereas the majority of Peripherin+ fibers were unmyelinated and small myelinated, a significant fraction (15.5%) were large myelinated (>20 µm(2) in cross-sectional area, equivalent to >5 µm in diameter, Aβ fiber). Our findings confirm that SP, CGRP, and IB4 can be used as reliable markers for neurons with unmyelinated fibers, and question the suitability of NF200 as a marker for neurons with myelinated fibers, and of Peripherin as a marker for neurons with unmyelinated, or fine-caliber fibers.
Landmesser, Lynn; Pilar, G.
1. The development of transmission was studied in chick ciliary ganglia that had been deprived of their periphery during early embryonic development. 2. Peripherally deprived neurones in the ganglion differentiate in normal numbers and send functional axons into the post-ganglionic nerve. 3. Ganglion cells lacking a periphery follow the normal developmental sequence sending out transient dendrites at the time ganglion cell synapses are formed, and later retracting them when calyces appear. 4. Synapses, which appear functionally and ultrastructurally normal, form on all ganglion cells at the normal time and transmission is normal until Stage 34. Therefore information from the periphery is not required for ganglion cell synapse formation per se. 5. From Stages 35 to 38 most cells die, so that only 8% of the original number of cells remain in the operated ganglion. Transmission fails in many cells during this same time, but precedes cell loss by only a short time, so that deafferentation probably does not contribute substantially to cell death. 6. Both ciliary and choroid cells achieve full cytologic differentiation and are distinct from each other, indicating that the periphery is not required for the elaboration of the distinctive characteristics of these cells. Presynaptic fibres also differentiate into typical bouton as well as calyciform endings. Therefore, the type of preganglionic ending does not depend on ganglion cells establishing proper peripheral contacts. 7. It has not been possible to ascertain whether ganglion cell specificity is affected by the periphery. 8. Peripheral removal affects ganglion cell migration, so that two ganglia are formed. Approximately half of the cells migrate into the remnant optic cup forming a second misplaced ganglion. Ciliary and choroid cells occur in both ganglia and these cells go through the typical sequence of events described above. ImagesPlate 1Plate 2Plate 3 PMID:4373567
Charles, K J; Evans, M L; Robbins, M J; Calver, A R; Leslie, R A; Pangalos, M N
GABA(B) receptors are G-protein-coupled receptors mediating the slow onset and prolonged synaptic actions of GABA in the CNS. The recent cloning of two genes, GABA(B1) and GABA(B2), has revealed a novel requirement for GABA(B) receptor signalling. Studies have demonstrated that the two receptor subunits associate as a GABA(B1)/GABA(B2) heterodimer to form a functional GABA(B) receptor. In this study we have developed polyclonal antisera specific to two splice variants of the GABA(B1) subunit, GABA(B1a) and GABA(B1b), as well as an antiserum to the GABA(B2) subunit. Using affinity-purified antibodies derived from these antisera we have mapped out the distribution profile of each subunit in rat brain, spinal cord and dorsal root ganglion. In brain the highest areas of GABA(B1a), GABA(B1b) and GABA(B2) subunit expression were found in neocortex, hippocampus, thalamus, cerebellum and habenula. In spinal cord, GABA(B1) and GABA(B2) subunits were expressed in the superficial layers of the dorsal horn, as well as in motor neurones in the deeper layers of the ventral horn. GABA(B) receptor subunit immunoreactivity in dorsal root ganglion suggested that expression of GABA(B1b) was restricted to the large diameter neurones, in contrast to GABA(B1a) and GABA(B2) subunits which were expressed in both large and small diameter neurones. Although expression levels of GABA(B1) and GABA(B2) subunits varied we found no areas in which GABA(B1) was expressed in the absence of GABA(B2). This suggests that most, if not all, GABA(B1) immunoreactivity may represent functional GABA(B) receptors. Although our data are in general agreement with functional studies, some discrepancies in GABA(B1) subunit expression occurred with respect to other immunohistochemical studies. Overall our data suggest that GABA(B) receptors are widely expressed throughout the brain and spinal cord, and that GABA(B1a) and GABA(B1b) subunits can associate with GABA(B2) to form both pre- and post-synaptic receptors.
Lee, Sang Pyo; Lee, Kang Nyeong; Lee, Hang Lak; Jun, Dae Won; Yoon, Byung Chul; Choi, Ho Soon; Hwang, Se Jin; Lee, Seo Eun
Background/Aims DA-9701, a standardized extract of Pharbitis Semen and Corydalis Tuber, is a new prokinetic agent that exhibits an analgesic effect on the abdomen. We investigated whether DA-9701 affects visceral pain induced by colorectal distension (CRD) in rats. Methods A total of 21 rats were divided into three groups: group A (no CRD+no drug), group B (CRD+no drug), and group C (CRD+DA-9701). Expression of pain-related factors, substance P (SP), c-fos, and phosphorylated extracellular signal-regulated kinase (p-ERK) in the dorsal root ganglion (DRG) and spinal cord was determined by immunohistochemical staining and Western blotting. Results The proportions of neurons in the DRG and spinal cord expressing SP, c-fos, and p-ERK were higher in group B than in group A. In the group C, the proportion of neurons in the DRG and spinal cord expressing p-ERK was lower than that in group B. Western blot results for p-ERK in the spinal cord indicated a higher level of expression in group B than in group A and a lower level of expression in group C than in group B. Conclusions DA-9701 may decrease visceral pain via the downregulation of p-ERK in the DRG and spinal cord. PMID:24672654
Nakanishi, Masako; Hata, Kenji; Nagayama, Tomotaka; Sakurai, Teruhisa; Nishisho, Toshihiko; Wakabayashi, Hiroki; Hiraga, Toru; Ebisu, Shigeyuki; Yoneda, Toshiyuki
Increased production of calcitonin gene-related peptide (CGRP) in sensory neurons is implicated in inflammatory pain. The inflammatory site is acidic due to proton release from infiltrating inflammatory cells. Acid activation of peripheral nociceptors relays pain signals to the CNS. Here, we examined whether acid activated the transient receptor potential vanilloid subtype 1 (Trpv1), a widely recognized acid-sensing nociceptor and subsequently increased CGRP expression. Chemically induced inflammation was associated with thermal hyperalgesia and increased CGRP expression in dorsal root ganglion (DRG) in rats. In organ cultures of DRG, acid (pH 5.5) elevated CGRP expression and the selective Trpv1 antagonist 5'-Iodoresiniferatoxin decreased it. Trpv1-deficient DRG showed reduced CGRP increase by acid. Of note, many of CGRP/Trpv1-positive DRG neurons exhibited the phosphorylation of cAMP response element-binding protein (CREB), a nociceptive transcription factor. Knockdown of CREB by small interfering RNA or a dominant-negative form of CREB diminished acid-elevated CGRP expression. Acid elevated the transcriptional activity of CREB, which in turn stimulated CGRP gene promoter activity. These effects were inhibited by a Ca(2+)/calmodulin-dependent protein kinase (CaMK) inhibitor KN-93. In conclusion, our results suggest that inflammatory acidic environments activate Trpv1, leading to an up-regulation of CGRP expression via CaMK-CREB cascade, a series of events that may be associated with inflammatory pain.
Yu, Jianfeng; Fu, Peng; Zhang, Yan; Liu, Shuzhen; Cui, Donghong
P2X3 receptors are present in the spinal dorsal horn (SDH) and play an essential role in the regulation of nociception and pain. Pregabalin (PGB) has been used as a new antiepileptic drug in the treatment of neuropathic pain. However, it is unclear whether PGB-induced analgesia was associated with the P2X3 receptor in SDH. Here, rats were randomly divided into four groups (n = 12 per group), including 2 sham operation groups, which was treated by normal saline (Sham + NS group) or PGB (Sham + PGB group), other 2 groups with chronic compression of the dorsal root ganglion, a normal saline-treated CCD group (CCD+NS group), and a PGB-treated CCD group (CCD + PGB group). A rat model of neuropathic pain was used by compressing the right L4 and L5 dorsal root ganglia. Each group was evaluated using the mechanical withdrawal threshold (MWT). The mRNA and protein levels of the P2X3 receptor in the ipsilateral SDH were measured by RT-PCR, western blot, and immunofluorescence on 14 day after CCD operation. CCD rats showed the highest mechanical hyperalgesia and the lowest pain threshold in the four groups. Simultaneously, CCD rats showed higher P2X3 mRNA and protein expression in ipsilateral side of the SDH than the sham operation rats. However, the MWT was increased and expression of P2X3 mRNA and protein in the ipsilateral SDH in CCD rats was decreased 3 days after PGB treatment. Thus, PGB may partially reverse mechanical hyperalgesia in CCD rats by inhibiting P2X3 receptor expression in the ipsilateral SDH.
Kazanci, Burak; Tehli, Ozkan; Guclu, Bulent
Ganglion cysts usually arise from the tissues around the facet joints. It is usually associated with degenerative cahanges in facet joints. Bilateral thoracic ganglion cysts are very rare and there is no previous case that located in bilateral intervertebral foramen compressing the L1 nerve root associated with severe radiculopathy. We report a 53 years old woman who presented with bilateral groin pain and severe numbness. Magnetic resonance imaging revealed bilateral cystic mass in the intervertebral foramen between 12th thoracal and 1st lumbar vertebrae. The cystic lesions were removed after bilateral exposure of Th12-L1 foramens. The result of hystopathology confirmed the diagnosis as ganglion cyst. The ganglion cyst may compromise lumbar dorsal ganglion when it located in the intervertebral foramen. The surgeon should keep this rare entity in their mind for differential diagnosis. PMID:23908708
Tomotsuka, Naoto; Kaku, Ryuji; Obata, Norihiko; Matsuoka, Yoshikazu; Kanzaki, Hirotaka; Taniguchi, Arata; Muto, Noriko; Omiya, Hiroki; Itano, Yoshitaro; Sato, Tadasu; Ichikawa, Hiroyuki; Mizobuchi, Satoshi; Morimatsu, Hiroshi
Metastatic bone cancer causes severe pain, but current treatments often provide insufficient pain relief. One of the reasons is that mechanisms underlying bone cancer pain are not solved completely. Our previous studies have shown that brain-derived neurotrophic factor (BDNF), known as a member of the neurotrophic family, is an important molecule in the pathological pain state in some pain models. We hypothesized that expression changes of BDNF may be one of the factors related to bone cancer pain; in this study, we investigated changes of BDNF expression in dorsal root ganglia in a rat bone cancer pain model. As we expected, BDNF mRNA (messenger ribonucleic acid) and protein were significantly increased in L3 dorsal root ganglia after intra-tibial inoculation of MRMT-1 rat breast cancer cells. Among the eleven splice-variants of BDNF mRNA, exon 1–9 variant increased predominantly. Interestingly, the up-regulation of BDNF is localized in small neurons (mostly nociceptive neurons) but not in medium or large neurons (non-nociceptive neurons). Further, expression of nerve growth factor (NGF), which is known as a specific promoter of BDNF exon 1–9 variant, was significantly increased in tibial bone marrow. Our findings suggest that BDNF is a key molecule in bone cancer pain, and NGF-BDNF cascade possibly develops bone cancer pain. PMID:25050075
Hoeber, Jan; Trolle, Carl; Konig, Niclas; Du, Zhongwei; Gallo, Alessandro; Hermans, Emmanuel; Aldskogius, Hakan; Shortland, Peter; Zhang, Su-Chun; Deumens, Ronald; Kozlova, Elena N.
Dorsal root avulsion results in permanent impairment of sensory functions due to disconnection between the peripheral and central nervous system. Improved strategies are therefore needed to reconnect injured sensory neurons with their spinal cord targets in order to achieve functional repair after brachial and lumbosacral plexus avulsion injuries. Here, we show that sensory functions can be restored in the adult mouse if avulsed sensory fibers are bridged with the spinal cord by human neural progenitor (hNP) transplants. Responses to peripheral mechanical sensory stimulation were significantly improved in transplanted animals. Transganglionic tracing showed host sensory axons only in the spinal cord dorsal horn of treated animals. Immunohistochemical analysis confirmed that sensory fibers had grown through the bridge and showed robust survival and differentiation of the transplants. Section of the repaired dorsal roots distal to the transplant completely abolished the behavioral improvement. This demonstrates that hNP transplants promote recovery of sensorimotor functions after dorsal root avulsion, and that these effects are mediated by spinal ingrowth of host sensory axons. These results provide a rationale for the development of novel stem cell-based strategies for functionally useful bridging of the peripheral and central nervous system. PMID:26053681
Scamps, Frédérique; Sangari, Sina; Bowerman, Melissa; Rousset, Mathieu; Bellis, Michel; Cens, Thierry; Charnet, Pierre
Small RGK GTPases, Rad, Gem, Rem1, and Rem2, are potent inhibitors of high-voltage-activated (HVA) Ca(2+) channels expressed in heterologous expression systems. However, the role of this regulation has never been clearly demonstrated in the nervous system. Using transcriptional analysis, we show that peripheral nerve injury specifically upregulates Gem in mice dorsal root ganglia. Following nerve injury, protein expression was increased in ganglia and peripheral nerve, mostly under its phosphorylated form. This was confirmed in situ and in vitro in dorsal root ganglia sensory neurons. Knockdown of endogenous Gem, using specific small-interfering RNA (siRNA), increased the HVA Ca(2+) current only in the large-somatic-sized neurons. Combining pharmacological analysis of the HVA Ca(2+) currents together with Gem siRNA-transfection of larger sensory neurons, we demonstrate that only the P/Q-type Ca(2+) channels were enhanced. In vitro analysis of Gem affinity to various CaVβx-CaV2.x complexes and immunocytochemical studies of Gem and CaVβ expression in sensory neurons suggest that the specific inhibition of the P/Q channels relies on both the regionalized upregulation of Gem and the higher sensitivity of the endogenous CaV2.1-CaVβ4 pair in a subset of sensory neurons including the proprioceptors. Finally, pharmacological inhibition of P/Q-type Ca(2+) current reduces neurite branching of regenerating axotomized neurons. Taken together, the present results indicate that a Gem-dependent P/Q-type Ca(2+) current inhibition may contribute to general homeostatic mechanisms following a peripheral nerve injury.
Walwyn, W M; Keith, D E; Wei, W; Tan, A M; Xie, C W; Evans, C J; Kieffer, B L; Maidment, N T
Although mu opioid receptors desensitize in various cell lines in vitro, the relationship of this change in signaling efficacy to the development of tolerance in vivo remains uncertain. It is clear that a system is needed in which functional mu opioid receptor expression is obtained in appropriate neurons so that desensitization can be measured, manipulated, and mutated receptors expressed in this environment. We have developed a recombinant system in which expression of a flag-tagged mu opioid receptor is returned to dorsal root ganglia neurons from mu opioid receptor knockout mice in vitro. Flow cytometry analysis showed that adenoviral-mediated expression of the amino-terminal flag-tagged mu opioid receptor in neurons resulted in approximately 1.3x10(6) receptors/cell. Many mu opioid receptor cell lines express a similar density of receptors but this is approximately 7x greater than the number of endogenous receptors expressed by matched wild-type neurons. Inhibition of the high voltage-activated calcium currents in dorsal root ganglia neurons by the mu agonist, D-Ala(2), N-MePhe(4), Gly(5)-ol-enkephalin (DAMGO), was not different between the endogenous and flag-tagged receptor at several concentrations of DAMGO used. Both receptors desensitized equally over the first 6 h of DAMGO pre-incubation, but after 24 h the response of the endogenous receptor to DAMGO had desensitized further than the flag- tagged receptor (71+/-3 vs 29+/-7% respectively; P<0.002), indicating less desensitization in neurons expressing a higher density of receptor. Using flow cytometry to quantify the percentage of receptors remaining on the neuronal cell surface, the flag-tagged receptor internalized by 17+/-1% after 20 min and 55+/-2% after 24 h of DAMGO. These data indicate that this return of function model in neurons recapitulates many of the characteristics of endogenous mu opioid receptor function previously identified in non-neuronal cell lines.
Rigosa, J.; Weber, D. J.; Prochazka, A.; Stein, R. B.; Micera, S.
Functional electrical stimulation (FES) is used to improve motor function after injury to the central nervous system. Some FES systems use artificial sensors to switch between finite control states. To optimize FES control of the complex behavior of the musculo-skeletal system in activities of daily life, it is highly desirable to implement feedback control. In theory, sensory neural signals could provide the required control signals. Recent studies have demonstrated the feasibility of deriving limb-state estimates from the firing rates of primary afferent neurons recorded in dorsal root ganglia (DRG). These studies used multiple linear regression (MLR) methods to generate estimates of limb position and velocity based on a weighted sum of firing rates in an ensemble of simultaneously recorded DRG neurons. The aim of this study was to test whether the use of a neuro-fuzzy (NF) algorithm (the generalized dynamic fuzzy neural networks (GD-FNN)) could improve the performance, robustness and ability to generalize from training to test sets compared to the MLR technique. NF and MLR decoding methods were applied to ensemble DRG recordings obtained during passive and active limb movements in anesthetized and freely moving cats. The GD-FNN model provided more accurate estimates of limb state and generalized better to novel movement patterns. Future efforts will focus on implementing these neural recording and decoding methods in real time to provide closed-loop control of FES using the information extracted from sensory neurons.
Unger, J W; Klitzsch, T; Pera, S; Reiter, R
A number of functions for nerve growth factor (NGF) have been described over the past years, including its role for neuronal function and regeneration during toxic or metabolic neuropathies. In order to further assess the effects of NGF on the somatosensory system in diabetic neuropathy, the sural nerve, dorsal root ganglia (DRG), and dorsal horn of the spinal cord were investigated by morphological and quantitative methods in rats after 12 weeks of uncontrolled streptozotocin-induced diabetes mellitus. The results from our study suggest a twofold effect of NGF: (1) In sural nerve treatment with NGF (0.1 or 0.5 mg/kg) for 12 weeks was able to reverse distinct diabetes-related alterations in myelinated nerve fiber morphology, such as myelin thickness. These changes occurred in the entire myelinated population of sensory nerves and were not restricted to nociceptive nerve fibers. (2) The NGF effect on neurotransmitters of the sensory, nociceptive system was reflected by increased CGRP and substance P content in the DRG and in the dorsal horn of the spinal cord. No change of trkA receptor immunostaining was seen in DRGs of diabetic rats; however, a reduction of trkA immunoreactivity of DRG neurons was noted after long-term NGF treatment of healthy controls. The data demonstrate that NGF regulates a number of neuronal parameters along peripheral and central parts of the somatosensory pathway in the adult. This neurotrophic support may be essential for inducing functionally significant regenerative mechanisms in diabetic neuropathy.
Chu, Li-Wen; Chen, Jun-Yih; Wu, Pao-Chu; Wu, Bin-Nan
Atorvastatin, traditionally used to treat hyperlipidemia, belongs to a class of 3-hydroxy-3-methyl-glutaryl coenzyme A (HMG-CoA) reductase inhibitors. This study investigated the antineuroinflammatory and antihyperalgesic effects of atorvastatin in dorsal root ganglia (DRG) and spinal cord for chronic constriction injury (CCI) neuropathic pain in rats. Fifty-four Sprague-Dawley rats were divided into three groups including sham, CCI, and CCI+atorvastatin. Rats were orally administered atorvastatin (10 mg/kg/day) once daily for 2 weeks after surgery and sacrificed at days 3, 7, and 14. All animals were assessed for mechanical allodynia and thermal hyperalgesia in both hindpaws. Western blotting, immunofluorescence, and enzyme-linked immunosorbent assay (ELISA) were used to detect inflammatory proteins and proinflammatory cytokines at day 7 after surgery. Pain behaviors were significantly reduced in the CCI+atorvastatin group compared to the CCI group. Atorvastatin attenuated CCI-induced inflammatory mediators (pAkt/Akt, COX-2, iNOS, EP1, and EP4) and reduced proinflammatory cytokines TNF-α and IL-1β levels in DRG and spinal cord. Atorvastatin also inhibited nuclear pNFκB activation. Double immunofluorescent staining further demonstrated that pNFκB proteins were decreased by atorvastatin in DRG satellite cells and spinal microglia. Atorvastatin may primarily inhibit the nuclear translocation of pNFκB to prevent CCI-induced peripheral neuropathic pain. Atorvastatin exhibits antineuroinflammatory and antinociceptive properties in the central and peripheral nerve systems.
Stemkowski, Patrick L; Noh, Myung-chul; Chen, Yishen; Smith, Peter A
Chronic constriction injury of rat sciatic nerve promotes signs of neuropathic pain. This is associated with an increase in the level of interleukin 1β (IL-1β) in primary afferents that peaks at 7 days. This initial cytokine exposure has been proposed to trigger an enduring alteration in neuronal phenotype that underlies chronic hyper-excitability in sensory nerves, which initiates and maintains chronic neuropathic pain. We have shown previously that 5–6 days of exposure of rat dorsal root ganglia (DRGs) to 100 pm IL-1β increases the excitability of medium-sized neurons. We have now found using whole-cell recording that this increased excitability reverts to control levels within 3–4 days of cytokine removal. The effects of IL-1β were dominated by changes in K+ currents. Thus, the amplitudes of A-current, delayed rectifier and Ca2+-sensitive K+ currents were reduced by ∼68%, ∼64% and ∼36%, respectively. Effects of IL-1β on other cation currents were modest by comparison. There was thus a slight decrease in availability of high voltage-activated Ca2+ channel current, a small increase in rates of activation of hyperpolarization-activated cyclic nucleotide-gated channel current (IH), and a shift in the voltage dependence of activation of tetrodotoxin-sensitive sodium current (TTX-S INa) to more negative potentials. It is unlikely, therefore, that direct interaction of IL-1β with DRG neurons initiates an enduring phenotypic shift in their electrophysiological properties following sciatic nerve injury. Persistent increases in primary afferent excitability following nerve injury may instead depend on altered K+ channel function and on the continued presence of slightly elevated levels IL-1β and other cytokines. PMID:26110238
Engle, Mitchell P; Merrill, Michelle A; Marquez De Prado, Blanca; Hammond, Donna L
This study examined the distribution of γ-aminobutyric acid (GABA)(B) receptors on immunohistochemically identified neurons, and levels of GABA(B(1)) and GABA(B(2)) mRNA, in the L4 and L5 dorsal root ganglia (DRG) of the rat in the absence of injury and 2 weeks after L5 spinal nerve ligation. In uninjured DRG, GABA(B(1)) immunoreactivity colocalized exclusively with the neuronal marker (NeuN) and did not colocalize with the satellite cell marker S-100. The GABA(B(1)) subunit colocalized to >97% of DRG neurons immunoreactive (IR) for neurofilament 200 (N52) or calcitonin gene-related peptide (CGRP), or labeled by isolectin B4 (IB4). Immunoreactivity for GABA(B(2)) was not detectable. L5 spinal nerve ligation did not alter the number of GABA(B(1)) -IR neurons or its colocalization pattern in the L4 DRG. However, ligation reduced the number of GABA(B(1)) -IR neurons in the L5 DRG by ≈38% compared with sham-operated and naïve rats. Specifically, ligation decreased the number of CGRP-IR neurons in the L5 DRG by 75%, but did not decrease the percent colocalization of GABA(B(1)) in those that remained. In the few IB4-positive neurons that remained in the L5 DRG, colocalization of GABA(B(1)) -IR decreased to 75%. Ligation also decreased levels of GABA(B(1)) and GABA(B(2)) mRNA in the L5, but not the L4 DRG compared with sham-operated or naïve rats. These findings indicate that the GABA(B) receptor is positioned to presynaptically modulate afferent transmission by myelinated, unmyelinated, and peptidergic afferents in the dorsal horn. Loss of GABA(B) receptors on primary afferent neurons may contribute to the development of mechanical allodynia after L5 spinal nerve ligation.
Fukuoka, Tetsuo; Kobayashi, Kimiko; Yamanaka, Hiroki; Obata, Koichi; Dai, Yi; Noguchi, Koichi
We compared the distribution of the alpha-subunit mRNAs of voltage-gated sodium channels Nav1.1-1.3 and Nav1.6-1.9 and a related channel, Nax, in histochemically identified neuronal subpopulations of the rat dorsal root ganglia (DRG). In the naïve DRG, the expression of Nav1.1 and Nav1.6 was restricted to A-fiber neurons, and they were preferentially expressed by TrkC neurons, suggesting that proprioceptive neurons possess these channels. Nav1.7, -1.8, and -1.9 mRNAs were more abundant in C-fiber neurons compared with A-fiber ones. Nax was evenly expressed in both populations. Although Nav1.8 and -1.9 were preferentially expressed by TrkA neurons, other alpha-subunits were expressed independently of TrkA expression. Actually, all IB4(+) neurons expressed both Nav1.8 and -1.9, and relatively limited subpopulations of IB4(+) neurons (3% and 12%, respectively) expressed Nav1.1 and/or Nav1.6. These findings provide useful information in interpreting the electrophysiological characteristics of some neuronal subpopulations of naïve DRG. After L5 spinal nerve ligation, Nav1.3 mRNA was up-regulated mainly in A-fiber neurons in the ipsilateral L5 DRG. Although previous studies demonstrated that nerve growth factor (NGF) and glial cell-derived neurotrophic factor (GDNF) reversed this up-regulation, the Nav1.3 induction was independent of either TrkA or GFRalpha1 expression, suggesting that the induction of Nav1.3 may be one of the common responses of axotomized DRG neurons without a direct relationship to NGF/GDNF supply. (c) 2008 Wiley-Liss, Inc.
Schlösser, Lukas; Barthel, Franziska; Brandenburger, Timo; Neumann, Elena; Bauer, Inge; Eulenburg, Volker; Werdehausen, Robert; Hermanns, Henning
Glycinergic inhibitory neurotransmission plays a pivotal role in the development of neuropathic pain. The glycine concentration in the synaptic cleft is controlled by the glycine transporters GlyT1 and GlyT2. GlyT1 is expressed throughout the central nervous system, while GlyT2 is exclusively located in glycinergic neurons. Aim of the present study was to investigate whether GlyTs are also expressed in the peripheral sensory nervous system and whether their expression is modulated in experimental neuropathic pain. Neuropathic pain was induced in male Wistar rats by Chronic Constriction Injury (CCI) and verified by assessment of mechanical allodynia (von Frey method). Expression patterns of GlyTs and the glycine binding subunit NR1 of the N-methyl-d-aspartate (NMDA) receptor in the spinal cord and dorsal root ganglia (DRG) were analyzed by Western blot analysis, PCR and immunohistochemistry. While both GlyT1 and GlyT2 were detected in the spinal cord, only GlyT1, but not GlyT2, was detected in DRG. Immunofluorescence revealed a strictly neuronal localization of GlyT1 and a co-localization of GlyT1 and NR1 in DRG. Compared to sham procedure, spinal cord and DRG expression of GlyT1 was not altered and NR1 was unchanged in DRG 12 days after CCI. GlyT1, but not GlyT2, is expressed in the peripheral sensory nervous system. The co-expression of GlyT1 and NMDA receptors in DRG suggests that GlyT1 regulates glycine concentration at the glycine binding site of the NMDA receptor. Differential regulation of GlyT1 expression in the spinal cord or DRG, however, does not seem to be associated with the development of neuropathic pain.
Zheng, Ji-Hong; Walters, Edgar T; Song, Xue-Jun
Injury or inflammation affecting sensory neurons in dorsal root ganglia (DRG) causes hyperexcitability of DRG neurons that can lead to spontaneous firing and neuropathic pain. Recent results indicate that after chronic compression of DRG (CCD treatment), both hyperexcitability of neurons in intact DRG and behaviorally expressed hyperalgesia are maintained by concurrent activity in cAMP-protein kinase A (PKA) and cGMP-protein kinase G (PKG) signaling pathways. We report here that when tested under identical conditions, dissociation produces a pattern of hyperexcitability in small DRG neurons similar to that produced by CCD treatment, manifest as decreased action potential (AP) current threshold, increased AP duration, increased repetitive firing to depolarizing pulses, increased spontaneous firing and resting depolarization. A novel feature of this hyperexcitability is its early expression-as soon as testing can be conducted after dissociation (approximately 2 h). Both forms of injury increase the electrophysiological responsiveness of the neurons to activation of cAMP-PKA and cGMP-PKG pathways as indicated by enhancement of hyperexcitability by agonists of these pathways in dissociated or CCD-treated neurons but not in control neurons. Although inflammatory signals are known to activate cAMP-PKA pathways, dissociation-induced hyperexcitability is unlikely to be triggered by signals released from inflammatory cells recruited to the DRG because of insufficient time for recruitment during the dissociation procedure. Inhibition by specific antagonists indicates that continuing activation of cAMP-PKA and cGMP-PKG pathways is required to maintain hyperexcitability after dissociation. The reduction of hyperexcitability by blockers of adenylyl cyclase and soluble guanylyl cyclase after dissociation suggests a continuing release of autocrine and/or paracrine factors from dissociated neurons and/or satellite cells, which activate both cyclases and help to maintain acute
Zhu, Gui-Qin; Liu, Su; He, Duan-Duan; Liu, Yue-Peng; Song, Xue-Jun
The objective of this study was to explore the role of cyclic adenosine monophosphate-protein kinase A (cAMP-PKA) signaling in the development of bone cancer pain in rats. Female Sprague-Dawley rats (N=48) were divided randomly into four groups: sham (n=8), tumor cell implantation (TCI) (n=16), TCI+saline (n=8), and TCI+PKA inhibitor (n=16). Bone cancer-induced pain behaviors - thermal hyperalgesia and mechanical allodynia - were tested at postoperative days -3, -1, 1, 3, 5, 7, 10, and 14. A PKA inhibitor, Rp-cAMPS (1 mmol/l/20 μl), was injected intrathecally on postoperative days 3, 4, and 5 (early phase) or 7, 8, and 9 postoperative days (late phase). The expression of PKA mRNA in dorsal root ganglia (DRG) was detected by reverse transcription-PCR. The concentration of cAMP and activity of PKA in DRG and spinal cord were measured by enzyme-linked immunosorbent assay. TCI treatment induced significant pain behaviors, manifested as thermal hyperalgesia and mechanical allodynia. Spinal administration of the PKA inhibitor Rp-cAMPS during the early phase and late phase significantly delayed or reversed, respectively, TCI-induced thermal hyperalgesia and mechanical allodynia. TCI treatment also led to obvious tumor growth and bone destruction. The level of PKA mRNA in the DRG, as well as the concentration of cAMP and the activity of PKA, in both the DRG and spinal cord were significantly increased after TCI treatment (P<0.01). We conclude that the inhibition of the cAMP-PKA signaling pathway may reduce bone cancer pain.
Effects of sciatic nerve transection on ultrastructure, NADPH-diaphorase reaction and serotonin-, tyrosine hydroxylase-, c-Fos-, glucose transporter 1- and 3-like immunoreactivities in frog dorsal root ganglion
Rigon, F.; Rossato, D.; Auler, V.B.; Dal Bosco, L.; Faccioni-Heuser, M.C.; Partata, W.A.
Frogs have been used as an alternative model to study pain mechanisms. Since we did not find any reports on the effects of sciatic nerve transection (SNT) on the ultrastructure and pattern of metabolic substances in frog dorsal root ganglion (DRG) cells, in the present study, 18 adult male frogs (Rana catesbeiana) were divided into three experimental groups: naive (frogs not subjected to surgical manipulation), sham (frogs in which all surgical procedures to expose the sciatic nerve were used except transection of the nerve), and SNT (frogs in which the sciatic nerve was exposed and transected). After 3 days, the bilateral DRG of the sciatic nerve was collected and used for transmission electron microscopy. Immunohistochemistry was used to detect reactivity for glucose transporter (Glut) types 1 and 3, tyrosine hydroxylase, serotonin and c-Fos, as well as nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-diaphorase). SNT induced more mitochondria with vacuolation in neurons, satellite glial cells (SGCs) with more cytoplasmic extensions emerging from cell bodies, as well as more ribosomes, rough endoplasmic reticulum, intermediate filaments and mitochondria. c-Fos immunoreactivity was found in neuronal nuclei. More neurons and SGCs surrounded by tyrosine hydroxylase-like immunoreactivity were found. No change occurred in serotonin- and Glut1- and Glut3-like immunoreactivity. NADPH-diaphorase occurred in more neurons and SGCs. No sign of SGC proliferation was observed. Since the changes of frog DRG in response to nerve injury are similar to those of mammals, frogs should be a valid experimental model for the study of the effects of SNT, a condition that still has many unanswered questions. PMID:23739744
Brain-derived neurotrophic factor promotes vesicular glutamate transporter 3 expression and neurite outgrowth of dorsal root ganglion neurons through the activation of the transcription factors Etv4 and Etv5.
Liu, Dong; Liu, Zhen; Liu, Huaxiang; Li, Hao; Pan, Xinliang; Li, Zhenzhong
Brain-derived neurotrophic factor (BDNF) is critical for sensory neuron survival and is necessary for vesicular glutamate transporter 3 (VGLUT3) expression. Whether the transcription factors Etv4 and Etv5 are involved in these BDNF-induced effects remains unclear. In the present study, primary cultured dorsal root ganglion (DRG) neurons were used to test the link between BDNF and transcription factors Etv4 and Etv5 on VGLUT3 expression and neurite outgrowth. BDNF promoted the mRNA and protein expression of Etv4 and Etv5 in DRG neurons. These effects were blocked by extracellular signal-regulated protein kinase 1/2 (ERK1/2) inhibitor PD98059 but not phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 or phospholipase C-γ (PLC-γ) inhibitor U73122. Etv4 siRNA and Etv5 siRNA effectively blocked the VGLUT3 expression and neurite elongation induced by BNDF. The overexpression of Etv4 or Etv5 potentiated the effects of BNDF-induced neurite elongation and growth-associated protein 43 (GAP-43), medium neurofilament (NF-M), and light neurofilament (NF-L) expression while these effects could be inhibited by Etv4 and Etv5 siRNA. These data imply that Etv4 and Etv5 are essential transcription factors in modulating BDNF/TrkB signaling-mediated VGLUT3 expression and neurite outgrowth. BDNF, through the ERK1/2 signaling pathway, activates Etv4 and Etv5 to initiate GAP-43 expression, promote neurofilament (NF) protein expression, induce neurite outgrowth, and mediate VGLUT3 expression for neuronal function improvement. The biological effects initiated by BDNF/TrkB signaling linked to E26 transformation-specific (ETS) transcription factors are important to elucidate neuronal differentiation, axonal regeneration, and repair in various pathological states.
Rose, Kirstin E.; Lunardi, Nadia; Boscolo, Annalisa; Dong, Xinzhong; Erisir, Alev; Jevtovic-Todorovic, Vesna; Todorovic, Slobodan M.
Previous behavioural studies have revealed that CaV3.2 T-type calcium channels support peripheral nociceptive transmission and electrophysiological studies have established the presence of T-currents in putative nociceptive sensory neurons of dorsal root ganglion (DRG). To date, however, the localization pattern of this key nociceptive channel in the soma and peripheral axons of these cells has not been demonstrated due to lack of isoform-selective anti-CaV3.2 antibodies. In the present study a new polyclonal CaV3.2 antibody is used to localize CaV3.2 expression in rodent DRG neurons using different staining techniques including confocal and electron microscopy. Confocal microscopy of both acutely dissociated cells and short-term cultures demonstrated strong immunofluorescence of anti-CaV3.2 antibody that was largely confined to smaller diameter DRG neurons where it co-localized with established immuno-markers of unmyelinated nociceptors, such as, CGRP, IB4 and peripherin. In contrast, a smaller proportion of these CaV3.2-labeled DRG cells also co-expressed NF-200, a marker of myelinated sensory neurons. In the rat sciatic nerve preparation, confocal microscopy demonstrated anti-CaV3.2 immunofluorescence which was co-localized with both peripherin and NF-200. Further, electron microscopy revealed immuno-gold labelling of CaV3.2 preferentially in association with un-myelinated sensory fibres from mouse sciatic nerve. Finally, we demonstrated the expression of CaV3.2 channels in peripheral nerve endings of mouse hindpaw skin as shown by co-localisation with Mrgpd-GFP-positive fibres. The CaV3.2 expression within the soma and peripheral axons of nociceptive sensory neurons further demonstrates the importance of this channel in peripheral pain transmission. PMID:23867767
Inhibition of the TRPM2 and TRPV1 Channels through Hypericum perforatum in Sciatic Nerve Injury-induced Rats Demonstrates their Key Role in Apoptosis and Mitochondrial Oxidative Stress of Sciatic Nerve and Dorsal Root Ganglion
Uslusoy, Fuat; Nazıroğlu, Mustafa; Çiğ, Bilal
Sciatic nerve injury (SNI) results in neuropathic pain, which is characterized by the excessive Ca2+ entry, reactive oxygen species (ROS) and apoptosis processes although involvement of antioxidant Hypericum perforatum (HP) through TRPM2 and TRPV1 activation has not been clarified on the processes in SNI-induced rat, yet. We investigated the protective property of HP on the processes in the sciatic nerve and dorsal root ganglion neuron (DRGN) of SNI-induced rats. The rats were divided into five groups as control, sham, sham+HP, SNI, and SNI+HP. The HP groups received 30 mg/kg HP for 4 weeks after SNI induction. TRPM2 and TRPV1 channels were activated in the neurons by ADP-ribose or cumene peroxide and capsaicin, respectively. The SNI-induced TRPM2 and TRPV1 currents and intracellular free Ca2+ and ROS concentrations were reduced by HP, N-(p-amylcinnamoyl) anthranilic acid (ACA), and capsazepine (CapZ). SNI-induced increase in apoptosis and mitochondrial depolarization in sciatic nerve and DRGN of SNI group were decreased by HP, ACA, and CapZ treatments. PARP-1, caspase 3 and 9 expressions in the sciatic nerve, DRGN, skin, and musculus piriformis of SNI group were also attenuated by HP treatment. In conclusion, increase of mitochondrial ROS, apoptosis, and Ca2+ entry through inhibition of TRPM2 and TRPV1 in the sciatic nerve and DRGN neurons were decreased by HP treatment. The results may be relevant to the etiology and treatment of SNI by HP. PMID:28620309
Shi, Yue; Liang, Xiao-Chun; Zhang, Hong; Sun, Qing; Wu, Qun-Li; Qu, Ling
To examine the effects of the combination of quercetin (Q), cinnamaldehyde (C) and hirudin (H), a Chinese medicine formula on high glucose (HG)-induced apoptosis of cultured dorsal root ganglion (DRG) neurons. DRG neurons exposed to HG (45 mmol/L) for 24 h were employed as an in vitro model of diabetic neuropathy. Cell viability, reactive oxygen species (ROS) level and apoptosis were determined. The expression of nuclear factor of Kappa B (NF-κB), inhibitory kappa Bα(IκBα), phosphorylated IκBα and Nf-E2 related factor 2 (Nrf2) were examined using reverse transcription-polymerase chain reaction (RT-PCR) and Western blot assay. The expression of hemeoxygenase-1 (HO-1), interleukin-6 (IL-6), tumor necrosis factor (TNF-α) and caspase-3 were also examined by RT-PCR and Western blot assay. HG treatment markedly increased DRG neuron apoptosis via increasing intracellular ROS level and activating the NF-κB signaling pathway (P<0.05). Co-treatment with Q, C, H and their combination decreased HG-induced caspase-3 activation and apoptosis (P<0.05 or P<0.01). The expressions of NF-κB, IL-6 and TNF-α were down-regulated, and Nrf2/HO-1 expression was up-regulated (P<0.05 or P<0.01). QCH has better effect in scavenging ROS, activating Nrf-2/HO-1, and down-regulating the NF-κB pathway than other treatment group. DRG neurons' apoptosis was increased in diabetic conditions, which was reduced by QCH formula treatment. The possible reason could be activating Nrf-2/HO-1 pathway, scavenging ROS, and inhibition of NF-κB activation. The effect of QCH combination was better than each monomer or the combination of the two monomers.
Interleukin-6-mediated functional upregulation of TRPV1 receptors in dorsal root ganglion neurons through the activation of JAK/PI3K signaling pathway: roles in the development of bone cancer pain in a rat model.
Fang, Dong; Kong, Ling-Yu; Cai, Jie; Li, Song; Liu, Xiao-Dan; Han, Ji-Sheng; Xing, Guo-Gang
Primary and metastatic cancers that affect bone are frequently associated with severe and intractable pain. The mechanisms underlying the pathogenesis of bone cancer pain still remain largely unknown. Previously, we have reported that sensitization of primary sensory dorsal root ganglion (DRG) neurons contributes to the pathogenesis of bone cancer pain in rats. In addition, numerous preclinical and clinical studies have revealed the pathological roles of interleukin-6 (IL-6) in inflammatory and neuropathic hyperalgesia. In this study, we investigated the role and the underlying mechanisms of IL-6 in the development of bone cancer pain using in vitro and in vivo approaches. We first demonstrated that elevated IL-6 in DRG neurons plays a vital role in the development of nociceptor sensitization and bone cancer-induced pain in a rat model through IL-6/soluble IL-6 receptor (sIL-6R) trans-signaling. Moreover, we revealed that functional upregulation of transient receptor potential vanilloid channel type 1 (TRPV1) in DRG neurons through the activation of Janus kinase (JAK)/phosphatidylinositol 3-kinase (PI3K) signaling pathway contributes to the effects of IL-6 on the pathogenesis of bone cancer pain. Therefore, suppression of functional upregulation of TRPV1 in DRG neurons by the inhibition of JAK/PI3K pathway, either before surgery or after surgery, reduces the hyperexcitability of DRG neurons and pain hyperalgesia in bone cancer rats. We here disclose a novel intracellular pathway, the IL-6/JAK/PI3K/TRPV1 signaling cascade, which may underlie the development of peripheral sensitization and bone cancer-induced pain.
Bay11-7082 attenuates neuropathic pain via inhibition of nuclear factor-kappa B and nucleotide-binding domain-like receptor protein 3 inflammasome activation in dorsal root ganglions in a rat model of lumbar disc herniation
Zhang, Ailiang; Wang, Kun; Ding, Lianghua; Bao, Xinnan; Wang, Xuan; Qiu, Xubin; Liu, Jinbo
Lumbar disc herniation (LDH) is an important cause of radiculopathy, but the underlying mechanisms are incompletely understood. Many studies suggested that local inflammation, rather than mechanical compression, results in radiculopathy induced by LDH. On the molecular and cellular level, nuclear factor-kappa B (NF-κB) and nucleotide-binding domain-like receptor protein 3 (NLRP3) inflammasome have been implicated in the regulation of neuroinflammation formation and progression. In this study, the autologous nucleus pulposus (NP) was implanted in the left L5 dorsal root ganglion (DRG) to mimic LDH in rats. We investigated the expression of NF-κB and the components of NLRP3 inflammasome in the DRG neurons in rats. Western blotting and immunofluorescence for the related molecules, including NLRP3, apoptosis-associated speck-like protein containing caspase-1 activator domain (ASC), caspase-1, interleukin (IL)-1β, IL-18, IκBα, p-IκBα, p65, p-p65, and calcitonin gene-related peptide (CGRP) were examined. In the NP-treated group, the activations of NLRP3, ASC, caspase-1, IL-1β, IL-18, p-IκBα, and p-p65 in DRG neurons in rats were elevated at 1 day after surgery, and the peak occurred at 7 days. Treatment with Bay11-7082, an inhibitor of the actions of IKK-β, was able to inhibit expression and activation of the molecules (NLRP3, ASC, caspase-1, IL-1β, IL-18, p-IκBα, and p-p65) and relieve the pain in rats. Our study shows that NF-κB and NLRP3 inflammasome are involved in the maintenance of NP-induced pain, and that Bay11-7082 could alleviate mechanical allodynia and thermal hyperalgesia by inhibiting NF-κB and NLRP3 inflammasome activation. PMID:28243141
Inhibition of the TRPM2 and TRPV1 Channels through Hypericum perforatum in Sciatic Nerve Injury-induced Rats Demonstrates their Key Role in Apoptosis and Mitochondrial Oxidative Stress of Sciatic Nerve and Dorsal Root Ganglion.
Uslusoy, Fuat; Nazıroğlu, Mustafa; Çiğ, Bilal
Sciatic nerve injury (SNI) results in neuropathic pain, which is characterized by the excessive Ca(2+) entry, reactive oxygen species (ROS) and apoptosis processes although involvement of antioxidant Hypericum perforatum (HP) through TRPM2 and TRPV1 activation has not been clarified on the processes in SNI-induced rat, yet. We investigated the protective property of HP on the processes in the sciatic nerve and dorsal root ganglion neuron (DRGN) of SNI-induced rats. The rats were divided into five groups as control, sham, sham+HP, SNI, and SNI+HP. The HP groups received 30 mg/kg HP for 4 weeks after SNI induction. TRPM2 and TRPV1 channels were activated in the neurons by ADP-ribose or cumene peroxide and capsaicin, respectively. The SNI-induced TRPM2 and TRPV1 currents and intracellular free Ca(2+) and ROS concentrations were reduced by HP, N-(p-amylcinnamoyl) anthranilic acid (ACA), and capsazepine (CapZ). SNI-induced increase in apoptosis and mitochondrial depolarization in sciatic nerve and DRGN of SNI group were decreased by HP, ACA, and CapZ treatments. PARP-1, caspase 3 and 9 expressions in the sciatic nerve, DRGN, skin, and musculus piriformis of SNI group were also attenuated by HP treatment. In conclusion, increase of mitochondrial ROS, apoptosis, and Ca(2+) entry through inhibition of TRPM2 and TRPV1 in the sciatic nerve and DRGN neurons were decreased by HP treatment. The results may be relevant to the etiology and treatment of SNI by HP.
Qiu, Fang; Li, Yang; Fu, Qiang; Fan, Yong-Yan; Zhu, Chao; Liu, Yan-Hong; Mi, Wei-Dong
Stromal cell-derived factor 1 (SDF-1)/chemokine CXC motif ligand 12 (CXCL12), a chemokine that is upregulated in dorsal root ganglion (DRG) during chronic pain models, has recently been found to play a central role in pain hypersensitivity. The purpose of present study is to investigate the functional impact of SDF-1 and its receptor, chemokine CXC motif receptor 4 (CXCR4), on two TTXR sodium channels in rat DRG using electrophysiological techniques. Preincubation with SDF-1 caused a concentration-dependent increase of Nav1.8 and Nav1.9 currents amplitudes in acutely isolated small diameter DRG neurons in short-term culture. As to Nav1.9, changes in current density and kinetic properties of Nav1.9 current evoked by SDF-1(50 ng/ml) was eliminated by CXCR4 antagonist AMD3100 and phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002. The increase in Nav1.9 current was also blocked by pertussis toxin (PTX) but not cholera toxin (CTX), showing involvement of Gi/o but not Gs subunits. As to Nav1.8, inhibitors (AMD3100, PTX, CTX, LY294002) used in present study didn't inhibit the increased amplitude of Nav1.8 current and shifted activation curve of Nav1.8 in a hyperpolarizing direction in the presence of SDF-1 (50 ng/ml). In conclusion, our data demonstrated that SDF-1 may excite primary nociceptive sensory neurons by acting on the biophysical properties of Nav1.8 and Nav1.9 currents but via different mechanisms.
Ma, Cheng; Li, Cui-xian; Yi, Jian-liang; Yan, Li-ping
To observe the effect of electroacupuncture (EA) on pain threshold and contents of excitatory amino acids (EAA) in dorsal root ganglia (DRG) and spinal cord in rats with neuropathic pain. Fifty SD rats were randomly divided into control (C),model (M), sham-model (SM), EA, and sham-EA groups, with 10 cases in each. Neuropathic pain (spared nerve injury, SNI) model was established by cutting off the right common peroneal nerve and proso-tibial nerve (with the sural nerve reserved intact). Before and after surgery, the mechanical pain threshold (MPT) and thermal pain threshold (TPT) were measured respectively on the injured side under consciousness state. EA (2 Hz, 1-3 mA, adding 1 mA/10 min) was applied to "Huantiao" (GB 30) and "Weizhong" (BL 40) on the affected side for 30 min. For rats of sham-EA group, filiform needles were inserted into GB30 and BL40 simply without manipulation or electrical stimulation. The treatment was given once daily for 7 days. On the 15th day, the rats were sacrificed for sampling right L4-L6 DRG and spinal cord. The contents of neurotransmitters, glutamate (Glu) and aspartic acid (Asp) in DRG and spinal cord were detected with high performance liquid chromatography (HPLC). Micro-dialysis technique was used to collect the dialysate from the spinal cord, homogenated for measuring EAA by HPLC. In comparison with control group, after SNI, MPT decreased significantly from the 1st day on in model group. Compared with model group, on the 15th day, MPT increased significantly in both EA and sham-EA groups (P < 0.05, 0.01),but MPT of sham-EA was significantly lower than that of EA group (P < 0.05). No significant changes were found in TPT in 5 groups (P > 0.05). It suggested that sham EA still had an analgesic effect in spite of being lower than that of true EA. Compared with control group, the contents of Glu and Asp in the spinal cord tissue and micro-dialysate in model group increased significantly after SNI (P < 0.01). In comparison with
Cho, Sung Min; Rhee, Woo Tack; Choi, Soo Jung; Eom, Dae Woon
The lumbar intraspinal epidural ganglion cyst has been a rare cause of the low back pain or leg pain. Ganglion cysts and synovial cysts compose the juxtafacet cysts. Extensive studies have been performed about the synovial cysts, however, very little has been known about the ganglion cyst. Current report is about two ganglion cysts associated with implicative findings in young male patients. We discuss about the underlying pathology of the ganglion cyst based on intraoperative evidences, associated disc herniation at the same location or severe degeneration of the ligament flavum that the cyst originated from in young patients.
Scroggs, R S
The effects of forskolin on low-threshold tetrodotoxin-resistant (TTX-r) Na(+) currents was studied in small diameter (average ≈ 25 μm) dorsal root ganglion (DRG) cells. All DRG cells included in the study were categorized as type-2 or non-type-2 based on the expression of a low-threshold A-current. In all type-2 and some non-type-2 DRG cells held at -80 mV, the adenylyl cyclase (AC) activator forskolin (10 μM) up-regulated TTX-r Na(+) currents evoked with steps to -55 mV through -35 mV (low-threshold current). Up-regulation of low-threshold current by forskolin was mimicked by the protein kinase A (PKA) agonist Sp-cAMPs and the inflammatory mediator serotonin, and blocked by the PKA antagonist Rp-cAMPs. Forskolin-induced up-regulation of low-threshold current evoked from a holding potential of -60 mV was blocked by 40 ms steps to 0 mV, which presumably induced a long lasting inactivation of the low-threshold channels. Reducing to 3 ms the duration of steps to 0 mV, significantly increased the number of DRG cells where low-threshold current was up-regulated by forskolin, presumably by reducing the long-lasting inactivation of the low-threshold channels. In the same cells, high-threshold current, evoked by 40 ms or 3 ms steps to 0 mV, was consistently up-regulated by forskolin. The selective Na(V)1.8 channel blocker A-803467 markedly blocked high-threshold current but not low-threshold current. The different voltage protocols observed to activate and inactivate the low- and high-threshold currents, and the observation that A-803467 blocked high- but not low-threshold current suggests that the two currents were mediated by different channels, possibly Na(V)1.8 and Na(V)1.9, respectively. Inflammatory mediators may simultaneously up-regulate Na(V)1.8 and Na(V)1.9 channels in the same nociceptor via a AC/PKA signaling pathway, increasing nociceptor signaling strength, and lowering nociceptor threshold, respectively.
Suen, Matthew; Fung, B; Lung, C P
Ganglion cysts are soft tissue swellings occurring most commonly in the hand or wrist. Apart from swelling, most cysts are asymptomatic. Other symptoms include pain, weakness, or paraesthesia. The two main concerns patients have are the cosmetic appearance of the cysts and the fear of future malignant growth. It has been shown that 58% of cysts will resolve spontaneously over time. Treatment can be either conservative or through surgical excision. This review concluded that nonsurgical treatment is largely ineffective in treating ganglion cysts. However, it advised to patients who do not surgical treatment but would like symptomatic relief. Compared to surgery, which has a lower recurrence rate but have a higher complication rate with longer recovery period. It has been shown that surgical interventions do not provide better symptomatic relief compared to conservative treatment. If symptomatic relief is the patient's primary concern, a conservative approach is preferred, whilst surgical intervention will decrease the likelihood of recurrence.
Ovariectomy-Induced Mitochondrial Oxidative Stress, Apoptosis, and Calcium Ion Influx Through TRPA1, TRPM2, and TRPV1 Are Prevented by 17β-Estradiol, Tamoxifen, and Raloxifene in the Hippocampus and Dorsal Root Ganglion of Rats.
Yazğan, Yener; Nazıroğlu, Mustafa
Relative 17β-estradiol (E2) deprivation and excessive production of mitochondrial oxygen free radicals (OFRs) with a high amount of Ca(2+) influx TRPA1, TRPM2, and TRPV1 activity is one of the main causes of neurodegenerative disease in postmenopausal women. In addition to the roles of tamoxifen (TMX) and raloxifene (RLX) in cancer and bone loss treatments, regulator roles in Ca(2+) influx and mitochondrial oxidative stress in neurons have not been reported. The aim of this study was to evaluate whether TMX and RLX interactions with TRPA1, TRPM2, and TRPV1 in primary hippocampal (HPC) and dorsal root ganglion (DRG) neuron cultures of ovariectomized (OVX) rats. Forty female rats were divided into five groups: a control group, an OVX group, an OVX+E2 group, an OVX+TMX group, and an OVX+RLX group. The OVX+E2, OVX+TMX, and OVX+RLX groups received E2, TMX, and RLX, respectively, for 14 days after the ovariectomy. E2, ovariectomy-induced TRPA1, TRPM2, and TRPV1 current densities, as well as accumulation of cytosolic free Ca(2+) in the neurons, were returned to the control levels by E2, TMX, and RLX treatments. In addition, E2, TMX, and RLX via modulation of TRPM2 and TRPV1 activity reduced ovariectomy-induced mitochondrial membrane depolarization, apoptosis, and cytosolic OFR production. TRPM2, TRPV1, PARP, and caspase-3 and caspase-9 expressions were also decreased in the neurons by the E2, TMX, and RLX treatments. In conclusion, we first reported the molecular effects of E2, TMX, and RLX on TRPA1, TRPM2, and TRPV1 channel activation in the OVX rats. In addition, we observed neuroprotective effects of E2, RLX, and TMX on oxidative and apoptotic injuries of the hippocampus and peripheral pain sensory neurons (DRGs) in the OVX rats. Graphical Abstract Possible molecular pathways of involvement of DEX in cerebral ischemia-induced apoptosis, oxidative stress, and calcium accumulation through TRPA1, TRPM2 and TRPV1 in the hippocampus and DRG neurons of rats. The N domain
Kahya, Mehmet Cemal; Nazıroğlu, Mustafa; Övey, İshak Suat
Neuropathic pain and hippocampal injury can arise from the overload of diabetes-induced calcium ion (Ca(2+)) entry and oxidative stress. The transient receptor potential (TRP) melastatin 2 (TRPM2) and TRP vanilloid type 1 (TRPV1) are expressed in sensory neurons and hippocampus. Moreover, activations of TRPM2 and TRPV1 during oxidative stress have been linked to neuronal death. Melatonin (MEL) and selenium (Se) have been considered potent antioxidants that detoxify a variety of reactive oxygen species (ROS) in neurological diseases. In order to better characterize the actions of MEL and Se in diabetes-induced peripheral pain and hippocampal injury through modulation of TRPM2 and TRPV1, we tested the effects of MEL and Se on apoptosis and oxidative stress in the hippocampal and dorsal root ganglion (DRG) neurons of streptozotocin (STZ)-induced diabetic rats. Fifty-eight rats were divided into six groups. The first group was used as control. The second group was used as the diabetic group. The third and fourth groups received Se and MEL, respectively. Intraperitoneal Se and MEL were given to diabetic rats in the fifth and sixth groups. On the 14th day, hippocampal and DRG neuron samples were freshly taken from all animals. The neurons were stimulated with a TRPV1 channel agonist (capsaicin) and a TRPM2 channel agonist (cumene hydroperoxide). We observed a modulator role of MEL and Se on intracellular free Ca(2+) concentrations, current densities of TRPM2 and TRPV1 channels, apoptosis, caspase 3, caspase 9, mitochondrial depolarization, reduced glutathione, glutathione peroxidase, lipid peroxidation, and intracellular ROS production values in the neurons. In addition, procaspase 3 and 9 activities in western blot analyses of the brain cortex were also decreased by MEL and Se treatments. In conclusion, in our diabetes experimental model, TRPM2 and TRPV1 channels are involved in the Ca(2+) entry-induced neuronal death and modulation of this channel activity by MEL and
Gonsalvez, David G; Cane, Kylie N; Landman, Kerry A; Enomoto, Hideki; Young, Heather M; Anderson, Colin R
Cell proliferation during nervous system development is poorly understood outside the mouse neocortex. We measured cell cycle dynamics in the embryonic mouse sympathetic stellate ganglion, where neuroblasts continue to proliferate following neuronal differentiation. At embryonic day (E) 9.5, when neural crest-derived cells were migrating and coalescing into the ganglion primordium, all cells were cycling, cell cycle length was only 10.6 h, and S-phase comprised over 65% of the cell cycle; these values are similar to those previously reported for embryonic stem cells. At E10.5, Sox10(+) cells lengthened their cell cycle to 38 h and reduced the length of S-phase. As cells started to express the neuronal markers Tuj1 and tyrosine hydroxylase (TH) at E10.5, they exited the cell cycle. At E11.5, when >80% of cells in the ganglion were Tuj1(+)/TH(+) neuroblasts, all cells were again cycling. Neuroblast cell cycle length did not change significantly after E11.5, and 98% of Sox10(-)/TH(+) cells had exited the cell cycle by E18.5. The cell cycle length of Sox10(+)/TH(-) cells increased during late embryonic development, and ∼25% were still cycling at E18.5. Loss of Ret increased neuroblast cell cycle length at E16.5 and decreased the number of neuroblasts at E18.5. A mathematical model generated from our data successfully predicted the relative change in proportions of neuroblasts and non-neuroblasts in wild-type mice. Our results show that, like other neurons, sympathetic neuron differentiation is associated with exit from the cell cycle; sympathetic neurons are unusual in that they then re-enter the cell cycle before later permanently exiting.
Ozdinler, P Hande; Ulupinar, Emel; Erzurumlu, Reha S
Nerve growth factor (NGF) and related neurotrophins are target-derived survival factors for sensory neurons. In addition, these peptides modulate neuronal differentiation, axon guidance, and synaptic plasticity. We tested axonal behavior of embryonic trigeminal neurons towards localized sources of NGF in collagen gel assays. Trigeminal axons preferentially grow towards lower doses of localized NGF and grow away from higher concentrations at earlier stages of development, but do not show this response later. Dorsal root ganglion axons also show similar responses to NGF, but NGF-dependent superior cervical ganglion axons do not. Such axonal responses to localized NGF sources were also observed in Bax-/- mice, suggesting that the axonal effects are largely independent of cell survival. Immunocytochemical studies indicated that axons, which grow towards or away from localized NGF are TrkA-positive, and TrkA-/- TG axons do not respond to any dose of NGF. We further show that axonal responses to NGF are absent in TG derived from mice that lack the p75 neurotrophin receptor (p75NTR). Collectively, our results suggest that localized sources of NGF can direct axon outgrowth from trigeminal ganglion in a dose- and age-dependent fashion, mediated by p75NTR signaling through TrkA expressing axons.
van Lindert, Erik; Bornemann, Antje; Hey, Otto; Perneczky, Axel; Müller-Forell, Wibke
An amyloidoma is a local deposition of amyloid that becomes a space-occupying lesion. Amyloidomas of the central nervous system are very uncommon lesions and only four amyloidomas of the gasserian ganglion have been reported so far. We present the neuroradiologic and surgical characteristics of three more amyloidomas of the gasserian ganglion seen at one neurosurgical department in 11 years. ImagesFigure 1Figure 2p215-bFigure 3 PMID:17170961
[Changes in the expression of large-conductance calcium-activated potassium channels in dorsal root ganglion neurons after electrical injury in rats' sciatic nerves and its influence on sensory conduction function].
Wang, Guangning; Li, Xueyong; Xu, Xiaoli; Ren, Pan
To study the changes in the expression of large-conductance calcium-activated potassium (BKCa) channels in dorsal root ganglion (DRG) neurons after electrical injury in rats' sciatic nerves and its influence on sensory conduction function. One-hundred and thirty-six adult SD rats were divided into normal control group, sham electrical injury group, and 75, 100, 125 V electrical injury groups according to the random number table, with 8 rats in normal control group and 32 rats in each of the rest 4 groups. Rats in normal control group were routinely fed without any treatment. Blunt dissection of the sciatic nerves of left hind leg of rats was performed in sham electrical injury group, while sciatic nerves of left hind leg of rats in electrical injury groups were electrically injured with corresponding voltage. Eight rats of normal control group fed for one week, and 8 rats from each of the rest four groups on post injury day (PID) 3 and in post injury week (PIW) 1, 2, 3 respectively were collected to detect the paw withdrawal mechanical threshold (PWMT). In addition, rats of 100 V electrical injury group in PIW 1 were collected and intrathecally injected with NS1619 after former PWMT detection, and PWMT was detected per 30 minutes within three hours post injection. The rats in each group at each time point were sacrificed after PWMT detection. The DRG of L4 to L6 segments of spinal cord was sampled to observe the BKCa channels distribution with immunohistochemical staining and to detect the protein and mRNA expressions of BKCa channels with Western blotting and reverse transcription-polymerase chain reaction respectively. Data were processed with one-way analysis of variance, analysis of variance of factorial design, and SNK test. (1) The PWMT values of rats in 75 and 100 V electrical injury groups on PID 3 and in PIW 1, 2, 3 were (5.8±0.6), (5.0±0.8), (4.2±0.3), (5.9±1.1) g; (5.3±1.3), (5.9±2.0), (4.5±2.7), (4.3±1.3) g, respectively, which were
Brenneis, Georg; Scholtz, Gerhard
Early neurogenesis in arthropods has been in the focus of numerous studies, its cellular basis, spatio-temporal dynamics and underlying genetic network being by now comparably well characterized for representatives of chelicerates, myriapods, hexapods and crustaceans. By contrast, neurogenesis during late embryonic and/or post-embryonic development has received less attention, especially in myriapods and chelicerates. Here, we apply (i) immunolabeling, (ii) histology and (iii) scanning electron microscopy to study post-embryonic ventral nerve cord development in Pseudopallene sp., a representative of the sea spiders (Pycnogonida), the presumable sister group of the remaining chelicerates. During early post-embryonic development, large neural stem cells give rise to additional ganglion cell material in segmentally paired invaginations in the ventral ectoderm. These ectodermal cell regions - traditionally designated as 'ventral organs' - detach from the surface into the interior and persist as apical cell clusters on the ventral ganglion side. Each cluster is a post-embryonic neurogenic niche that features a tiny central cavity and initially still houses larger neural stem cells. The cluster stays connected to the underlying ganglionic somata cortex via an anterior and a posterior cell stream. Cell proliferation remains restricted to the cluster and streams, and migration of newly produced cells along the streams seems to account for increasing ganglion cell numbers in the cortex. The pycnogonid cluster-stream-systems show striking similarities to the life-long neurogenic system of decapod crustaceans, and due to their close vicinity to glomerulus-like neuropils, we consider their possible involvement in post-embryonic (perhaps even adult) replenishment of olfactory neurons - as in decapods. An instance of a potentially similar post-embryonic/adult neurogenic system in the arthropod outgroup Onychophora is discussed. Additionally, we document two transient posterior
Maffei, Lamberto; Galli-Resta, Lucia
The spontaneous discharges of neighboring retinal ganglion cells were recorded simultaneously in anesthetized prenatal rats between embryonic days 18 and 21. We report here that in the majority of cases the firings of neighboring retinal ganglion cells are strongly correlated during prenatal life. Correlation in the discharges of neighboring cells during development has long been suggested as a way to consolidate synaptic connections with a target cell onto which they converge, a model first proposed by Hebb. Correlation in the activities of neighboring neurons in the retina could be the basis of developmental processes such as refinement of retinotopic maps in the brain and segregation of the inputs from the two eyes.
Magliulo, Giuseppe; Iannella, Giannicola; Valente, Michele; Greco, Antonio; Appiani, Mario Ciniglio
Objectives Discussion of a rare case of angioleiomyoma involving the geniculate ganglion and the intratemporal facial nerve segment and its surgical treatment. Design Case report. Setting Presence of an expansive lesion englobing the geniculate ganglion without any lesion to the cerebellopontine angle. Participants A 45-year-old man with a grade III facial paralysis according to the House-Brackmann scale of evaluation. Main Outcomes Measure Surgical pathology, radiologic appearance, histological features, and postoperative facial function. Results Removal of the entire lesion was achieved, preserving the anatomic integrity of the nerve; no nerve graft was necessary. Postoperative histology and immunohistochemical studies revealed features indicative of solid vascular leiomyoma. Conclusion Angioleiomyoma should be considered in the differential diagnosis of geniculate ganglion lesions. Optimal postoperative facial function is possible only by preserving the anatomical and functional integrity of the facial nerve. PMID:23943721
Magliulo, Giuseppe; Iannella, Giannicola; Valente, Michele; Greco, Antonio; Ciniglio Appiani, Mario
Objectives Discussion of a rare case of angioleiomyoma involving the geniculate ganglion and the intratemporal facial nerve segment and its surgical treatment. Design Case report. Setting Presence of an expansive lesion englobing the geniculate ganglion without any lesion to the cerebellopontine angle. Participants A 45-year-old man with a grade III facial paralysis according to the House-Brackmann scale of evaluation. Main Outcomes Measure Surgical pathology, radiologic appearance, histological features, and postoperative facial function. Results Removal of the entire lesion was achieved, preserving the anatomic integrity of the nerve; no nerve graft was necessary. Postoperative histology and immunohistochemical studies revealed features indicative of solid vascular leiomyoma. Conclusion Angioleiomyoma should be considered in the differential diagnosis of geniculate ganglion lesions. Optimal postoperative facial function is possible only by preserving the anatomical and functional integrity of the facial nerve.
... treatment of a ganglion cyst is not surgical. • Observation. Because the ganglion is not cancerous and may ... to go home a er a period of observation in the recovery area. There may be some ...
Bontempo, Nicholas A; Weiss, Arnold-Peter C
Arthroscopy is an advancing field in orthopedics, the applications of which have been expanding over time. Traditionally, excision of ganglion cysts has been done in an open fashion. However, more recently, studies show outcomes following arthroscopic excision to be as good as open excision. Cosmetically, the incisions are smaller and heal faster following arthroscopy. In addition, there is the suggested benefit that patients will regain function and return to work faster following arthroscopic excision. More prospective studies comparing open and arthroscopic excision of ganglion cysts need to be done in order to delineate if there is a true functional benefit.
Nieuwland, Jeroen; Stamm, Petra; Wen, Bo; Randall, Ricardo S; Murray, James A H; Bassel, George W
Seeding establishment following seed germination requires activation of the root meristem for primary root growth. We investigated the hormonal and genetic regulation of root meristem activation during Arabidopsis seed germination. In optimal conditions, radicle cell divisions occur only after the completion of germination and require de novo GA synthesis. When the completion of germination is blocked by ABA, radicle elongation and cell divisions occurred in these non-germinating seeds. Conversely under GA-limiting conditions, ABA-insensitive mutants complete germination in the absence of radicle meristem activation and growth. Radicle meristem activation and extension can therefore occur independently of completion of the developmental transition of germination. The cell cycle regulator KRP6 partially represses GA-dependent activation of the cell cycle. Germination of krp6 mutant seeds occurs more rapidly, is slightly insensitive to ABA in dose-response assays, but also hypersensitive to the GA synthesis inhibitor PAC. These conflicting phenotypes suggest the cell cycle uncouples GA and ABA responses in germinating Arabidopsis seeds, and that KRP6 acts downstream of GA to inhibit mitotic cell cycle activation during germination.
DeCastro, S; Sparks, J R; Lapey, J D; Freidberg, S R
A case report, the third in the literature, is presented of a patient whose progressive numbness in the second and third divisions of the trigeminal nerve led to the discovery of an isolated amyloidoma of the gasserian ganglion. The clinical impression of tumor was confirmed by surgical and pathologic findings.
Lui, Tun Hing
The tarsal tunnel ganglion is a cause of posterior tarsal tunnel syndrome. Open resection of the ganglion calls for release of the flexor retinaculum and dissection around the tibial neurovascular bundle. This can induce fibrosis around the tibial nerve. We report the technique of endoscopic resection of the tarsal tunnel ganglion. It is indicated for tarsal tunnel ganglia arising from the adjacent joints or tendon sheaths and compressing the tibial nerve from its deep side. It is contraindicated if there is other pathology of the tarsal tunnel that demands open surgery; if the ganglion compresses the tibial nerve from its superficial side, which calls for a different endoscopic approach using the ganglion portal; or if an intraneural ganglion of the tibial nerve is present. The purpose of this technical note is to describe a minimally invasive approach for endoscopic resection of the tarsal tunnel ganglion.
Fateev, M M; Zaikina, M G
The retrograde transport of horseradish peroxidase (HRP) has been used to study the localization and the number of neurons innervating the heart in the right stellate ganglion and accessory cervical ganglion, spinal cord and dorsal root ganglia of the cat. HRP was applied to the central cuts of anastomose of the stellate ganglion with the vagal nerve, of the vagosympathetic trunk caudal to anastomose and of the inferior cardiac nerve. HRP-labelled neurons were detected in the stellate ganglion in the regions which give off nerves, whereas in the accessory cervical ganglion labelled neurons were distributed throughout the whole ganglion. HRP-stained cells were found in the anastomose. In the spinal cord labelled neurons were detected in the lateral horn of T1-T5 segments. In the dorsal root ganglion the greatest number of neurons was observed in T2-T4 segments.
Niemi, Jon P; DeFrancesco-Lisowitz, Alicia; Cregg, Jared M; Howarth, Madeline; Zigmond, Richard E
Neuroinflammation plays a critical role in the regeneration of peripheral nerves following axotomy. An injury to the sciatic nerve leads to significant macrophage accumulation in the L5 DRG, an effect not seen when the dorsal root is injured. We recently demonstrated that this accumulation around axotomized cell bodies is necessary for a peripheral conditioning lesion response to occur. Here we asked whether overexpression of the monocyte chemokine CCL2 specifically in DRG neurons of uninjured mice is sufficient to cause macrophage accumulation and to enhance regeneration or whether other injury-derived signals are required. AAV5-EF1α-CCL2 was injected intrathecally, and this injection led to a time-dependent increase in CCL2 mRNA expression and macrophage accumulation in L5 DRG, with a maximal response at 3 weeks post-injection. These changes led to a conditioning-like increase in neurite outgrowth in DRG explant and dissociated cell cultures. This increase in regeneration was dependent upon CCL2 acting through its primary receptor CCR2. When CCL2 was overexpressed in CCR2-/- mice, macrophage accumulation and enhanced regeneration were not observed. To address the mechanism by which CCL2 overexpression enhances regeneration, we tested for elevated expression of regeneration-associated genes in these animals. Surprisingly, we found that CCL2 overexpression led to a selective increase in LIF mRNA and neuronal phosphorylated STAT3 (pSTAT3) in L5 DRGs, with no change in expression seen in other RAGs such as GAP-43. Blockade of STAT3 phosphorylation by each of two different inhibitors prevented the increase in neurite outgrowth. Thus, CCL2 overexpression is sufficient to induce macrophage accumulation in uninjured L5 DRGs and increase the regenerative capacity of DRG neurons via a STAT3-dependent mechanism. Copyright © 2015 Elsevier Inc. All rights reserved.
Niemi, Jon P.; DeFrancesco-Lisowitz, Alicia; Cregg, Jared; Howarth, Madeline; Zigmond, Richard E.
Neuroinflammation plays a critical role in the regeneration of peripheral nerves following axotomy. An injury to the sciatic nerve leads to significant macrophage accumulation in the L5 DRG, an effect not seen when the dorsal root is injured. We recently demonstrated that this accumulation around axotomized cell bodies is necessary for a peripheral conditioning lesion response to occur. Here we asked whether overexpression of the monocyte chemokine CCL2 specifically in DRG neurons of uninjured mice is sufficient to cause macrophage accumulation and to enhance regeneration or whether other injury-derived signals are required. AAV5-EF1α-CCL2 was injected intrathecally, and this injection led to a time-dependent increase in CCL2 mRNA expression and macrophage accumulation in L5 DRG, with a maximal response at 3 wk post-injection. These changes led to a conditioning-like increase in neurite outgrowth in DRG explant and dissociated cell cultures. This increase in regeneration was dependent upon CCL2 acting through its primary receptor CCR2. When CCL2 was overexpressed in CCR2 −/− mice, macrophage accumulation and enhanced regeneration were not observed. To address the mechanism by which CCL2 overexpression enhances regeneration, we tested for elevated expression of regeneration-associated genes in these animals. Surprisingly, we found that CCL2 overexpression led to a selective increase in LIF mRNA and neuronal phosphorylated STAT3 (pSTAT3) in L5 DRGs, with no change in expression seen in other RAGs such as GAP-43. Blockade of STAT3 phosphorylation by each of two different inhibitors prevented the increase in neurite outgrowth. Thus, CCL2 overexpression is sufficient to induce macrophage accumulation in uninjured L5 DRGs and increase the regenerative capacity of DRG neurons via a STAT3-dependent mechanism. PMID:26431741
Effect of electroacupuncture on thermal pain threshold and expression of calcitonin-gene related peptide, substance P and γ-aminobutyric acid in the cervical dorsal root ganglion of rats with incisional neck pain.
Qiao, Li-Na; Liu, Jun-Ling; Tan, Lian-Hong; Yang, Hai-Long; Zhai, Xu; Yang, Yong-Sheng
Acupuncture therapy effectively reduces post-surgical pain, but its mechanism of action remains unclear. The aim of this study was to investigate whether expression of γ-aminobutyric acid (GABA) and the neuropeptides substance P (SP) and calcitonin gene-related peptide (CGRP) in the primary sensory neurons of cervical dorsal root ganglia (DRG) are involved in electroacupuncture (EA)-induced analgesia in a rat model of incisional neck pain. The pain model was established by making a longitudinal midline neck incision in 60 rats. Another 15 rats underwent sham surgery (normal group). Post-incision, 15 rats remained untreated (model group) and 45 rats underwent EA (frequency 2/100 Hz, intensity 1 mA) at bilateral LI18, LI4-PC6 or ST36-GB34 (n=15 each) for 30 min at 4 hours, 24 hours, and 48 hours post-surgery, followed by thermal pain threshold (PT) measurement. 30 min later, the rats were euthanased and cervical (C3-6) DRGs removed for measurement of immunoreactivity and mRNA expression of SP/CGRP and the GABAergic neuronal marker glutamic acid decarboxylase 67 (GAD67). Thermal PT was significantly lower in the model group versus the normal group and increased in the LI18 and LI4-PC6 groups but not the ST36-GB34 group compared with the model group. Additionally, EA at LI18 and LI4-PC6 markedly suppressed neck incision-induced upregulation of mRNA/protein expression of SP/CGRP, and upregulated mRNA/protein expression of GAD67 in the DRGs of C3-6 segments. EA at LI18/LI4-PC6 increases PT in rats with incisional neck pain, which is likely related to downregulation of pronociceptive mediators SP/CGRP and upregulation of the inhibitory transmitter GABA in the primary sensory neurons of cervical DRGs. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.
Lo, Chen-Peng; Cheng, Ching-Li; Lin, Sing-Ru; Lee, Ming-Hui; Chang, Jean-Yun; Subhan, Dudekula; Tsai, Ming-Shu; Pugazhenthi, Subbiah; Chang, Nan-Shan; Chen, Shur-Tzu
Background Tumor suppressor WOX1 (also named WWOX or FOR) is known to participate in neuronal apoptosis in vivo. Here, we investigated the functional role of WOX1 and transcription factors in the delayed loss of axotomized neurons in dorsal root ganglia (DRG) in rats. Methodology/Principal Findings Sciatic nerve transection in rats rapidly induced JNK1 activation and upregulation of mRNA and protein expression of WOX1 in the injured DRG neurons in 30 min. Accumulation of p-WOX1, p-JNK1, p-CREB, p-c-Jun, NF-κB and ATF3 in the nuclei of injured neurons took place within hours or the first week of injury. At the second month, dramatic nuclear accumulation of WOX1 with CREB (>65% neurons) and NF-κB (40–65%) occurred essentially in small DRG neurons, followed by apoptosis at later months. WOX1 physically interacted with CREB most strongly in the nuclei as determined by FRET analysis. Immunoelectron microscopy revealed the complex formation of p-WOX1 with p-CREB and p-c-Jun in vivo. WOX1 blocked the prosurvival CREB-, CRE-, and AP-1-mediated promoter activation in vitro. In contrast, WOX1 enhanced promoter activation governed by c-Jun, Elk-1 and NF-κB. WOX1 directly activated NF-κB-regulated promoter via its WW domains. Smad4 and p53 were not involved in the delayed loss of small DRG neurons. Conclusions/Significance Rapid activation of JNK1 and WOX1 during the acute phase of injury is critical in determining neuronal survival or death, as both proteins functionally antagonize. In the chronic phase, concurrent activation of WOX1, CREB, and NF-κB occurs in small neurons just prior to apoptosis. Likely in vivo interactions are: 1) WOX1 inhibits the neuroprotective CREB, which leads to eventual neuronal death, and 2) WOX1 enhances NF-κB promoter activation (which turns to be proapoptotic). Evidently, WOX1 is the potential target for drug intervention in mitigating symptoms associated with neuronal injury. PMID:19918364
Brenneis, Georg; Scholtz, Gerhard
Early neurogenesis in arthropods has been in the focus of numerous studies, its cellular basis, spatio-temporal dynamics and underlying genetic network being by now comparably well characterized for representatives of chelicerates, myriapods, hexapods and crustaceans. By contrast, neurogenesis during late embryonic and/or post-embryonic development has received less attention, especially in myriapods and chelicerates. Here, we apply (i) immunolabeling, (ii) histology and (iii) scanning electron microscopy to study post-embryonic ventral nerve cord development in Pseudopallene sp., a representative of the sea spiders (Pycnogonida), the presumable sister group of the remaining chelicerates. During early post-embryonic development, large neural stem cells give rise to additional ganglion cell material in segmentally paired invaginations in the ventral ectoderm. These ectodermal cell regions – traditionally designated as ‘ventral organs’ – detach from the surface into the interior and persist as apical cell clusters on the ventral ganglion side. Each cluster is a post-embryonic neurogenic niche that features a tiny central cavity and initially still houses larger neural stem cells. The cluster stays connected to the underlying ganglionic somata cortex via an anterior and a posterior cell stream. Cell proliferation remains restricted to the cluster and streams, and migration of newly produced cells along the streams seems to account for increasing ganglion cell numbers in the cortex. The pycnogonid cluster-stream-systems show striking similarities to the life-long neurogenic system of decapod crustaceans, and due to their close vicinity to glomerulus-like neuropils, we consider their possible involvement in post-embryonic (perhaps even adult) replenishment of olfactory neurons – as in decapods. An instance of a potentially similar post-embryonic/adult neurogenic system in the arthropod outgroup Onychophora is discussed. Additionally, we document two transient
Quilliam, J. P.
Paraldehyde and methylpentynol blocked transmission of nerve impulses through the superior cervical ganglion of the cat when the drugs were administered intra-arterially to the ganglion or intravenously using the nictitating membrane as an indicator. Electrical studies showed that concentrations of methylpentynol and paraldehyde which blocked transmission in the isolated rat superior cervical ganglion were without action on the preganglionic nerve fibre. In amounts which blocked transmission in the isolated rat ganglion, paraldehyde had no depolarizing activity directly on the ganglion cells and did not interfere with the depolarizing activity of added acetylcholine. The results suggest that the block in transmission of the impulse could be accounted for by a decrease in the release of acetylcholine from the preganglionic nerve terminals. In both species the block was reversible. PMID:13662589
QUILLIAM, J P
Paraldehyde and methylpentynol blocked transmission of nerve impulses through the superior cervical ganglion of the cat when the drugs were administered intra-arterially to the ganglion or intravenously using the nictitating membrane as an indicator. Electrical studies showed that concentrations of methylpentynol and paraldehyde which blocked transmission in the isolated rat superior cervical ganglion were without action on the preganglionic nerve fibre. In amounts which blocked transmission in the isolated rat ganglion, paraldehyde had no depolarizing activity directly on the ganglion cells and did not interfere with the depolarizing activity of added acetylcholine. The results suggest that the block in transmission of the impulse could be accounted for by a decrease in the release of acetylcholine from the preganglionic nerve terminals. In both species the block was reversible.
Do, Michael Tri Hoang; Yau, King-Wai
Life on earth is subject to alternating cycles of day and night imposed by the rotation of the earth. Consequently, living things have evolved photodetective systems to synchronize their physiology and behavior with the external light-dark cycle. This form of photodetection is unlike the familiar "image vision," in that the basic information is light or darkness over time, independent of spatial patterns. "Nonimage" vision is probably far more ancient than image vision and is widespread in living species. For mammals, it has long been assumed that the photoreceptors for nonimage vision are also the textbook rods and cones. However, recent years have witnessed the discovery of a small population of retinal ganglion cells in the mammalian eye that express a unique visual pigment called melanopsin. These ganglion cells are intrinsically photosensitive and drive a variety of nonimage visual functions. In addition to being photoreceptors themselves, they also constitute the major conduit for rod and cone signals to the brain for nonimage visual functions such as circadian photoentrainment and the pupillary light reflex. Here we review what is known about these novel mammalian photoreceptors.
Chen, Ying-Jiun J.; Friedman, Brad A.; Ha, Connie; Durinck, Steffen; Liu, Jinfeng; Rubenstein, John L.; Seshagiri, Somasekar; Modrusan, Zora
Many subtypes of cortical interneurons (CINs) are found in adult mouse cortices, but the mechanism generating their diversity remains elusive. We performed single-cell RNA sequencing on the mouse embryonic medial ganglionic eminence (MGE), the major birthplace for CINs, and on MGE-like cells differentiated from embryonic stem cells. Two distinct cell types were identified as proliferating neural progenitors and immature neurons, both of which comprised sub-populations. Although lineage development of MGE progenitors was reconstructed and immature neurons were characterized as GABAergic, cells that might correspond to precursors of different CINs were not identified. A few non-neuronal cell types were detected, including microglia. In vitro MGE-like cells resembled bona fide MGE cells but expressed lower levels of Foxg1 and Epha4. Together, our data provide detailed understanding of the embryonic MGE developmental program and suggest how CINs are specified. PMID:28361918
Monoamine agonists and antagonists were applied to the lobster cardiac ganglion in an attempt to clarify the different actions of 5-hydroxytryptamine (5HT) and dopamine (DA) on this rhythmic pattern generator. Experiments were designed to determine whether the similar responses to 5HT and DA applied to the anterior region of the ganglion could be separated by pharmacological approaches, and whether the different responses to 5HT applied to the anterior and posterior regions of the ganglion could be attributed to mediation by different receptors. A small number of the 5HT agonists which were tested mimic the effects of 5HT, in that they increase the frequency of bursting and decrease burst duration when applied to the whole ganglion, but decrease burst frequency and increase burst duration when applied only to the posterior half. Other 5HT agonists decrease frequency and prolong bursts when applied to the whole ganglion. Of the DA agonists tested, none acts as DA itself does. Rather, they mimic the effects of 5HT applied to the posterior ganglion, by slowing bursting and prolonging bursts. The actions of agonists do not correspond in any clear way to the receptor specificities as defined in vertebrates. Most antagonists tested do not show similar specificities to their effects in vertebrates. In particular, most of the DA antagonists tested are more effective in blocking exogenous 5HT than DA. One monoamine agonist directly alters the properties of endogenous burst-organizing potentials (driver potentials) in the motorneurons of the ganglion.
Ohtani, Nobuyo; Goto, Tomohide; Waeber, Christian; Bhide, Pradeep G.
Dopamine is a neuromodulator the functions of which in the regulation of complex behaviors such as mood, motivation, and attention are well known. Dopamine appears in the brain early in the embryonic period when none of those behaviors is robust, raising the possibility that dopamine may influence brain development. The effects of dopamine on specific developmental processes such as neurogenesis are not fully characterized. The neostriatum is a dopamine-rich region of the developing and mature brain. If dopamine influenced neurogenesis, the effects would likely be pronounced in the neostriatum. Therefore, we examined whether dopamine influenced neostriatal neurogenesis by influencing the cell cycle of progenitor cells in the lateral ganglionic eminence (LGE), the neuroepithelial precursor of the neostriatum. We show that dopamine arrives in the LGE via the nigrostriatal pathway early in the embryonic period and that neostriatal neurogenesis progresses in a dopamine-rich milieu. Dopamine D1-like receptor activation reduces entry of progenitor cells from the G1-to S-phase of the cell cycle, whereas D2-like receptor activation produces the opposite effects by promoting G1- to S-phase entry. D1-like effects are prominent in the ventricular zone, and D2-like effects are prominent in the subventricular zone. The overall effects of dopamine on the cell cycle are D1-like effects, most likely because of the preponderance of D1-like binding sites in the embryonic neostriatum. These data reveal a novel developmental role for dopamine and underscore the relevance of dopaminergic signaling in brain development. PMID:12684471
Singh, B; Ramsaroop, L; Partab, P; Moodley, J; Satyapal, K S
In recent years the second thoracic ganglion has gained anatomical significance as an important conduit for sympathetic innervation of the upper extremity. Thoracoscopic excision of the second thoracic ganglion is now widely recognized as affording the most effective treatment option for palmar hyperhidrosis. This study recorded the incidence, location and associated additional neural connections of the second thoracic ganglion. Bilateral dissection of 20 adult cadavers was undertaken, and all neural connections of the second thoracic ganglion were recorded. Nineteen cadavers (95%) demonstrated additional neural connections between the first thoracic ventral ramus and second intercostal nerve. These were classified as either type A (47.5%) or type B (45%) using the intrathoracic ramus (nerve of Kuntz) between the second intercostal nerve and the ventral ramus of the first thoracic nerve as a basis on both right and left sides. The second thoracic ganglion was commonly located (92.5%) in the second intercostal space at the level of the intervertebral disc between the second and third thoracic vertebrae. Fused ganglia between the second thoracic and first thoracic (5%) and stellate (5%) ganglia were noted. These findings should assist the operating surgeon with a clear knowledge of the anatomy of the second thoracic ganglion during thoracoscopic sympathectomy with a view to improving the success rate for upper limb sympathectomy.
Cooke, I M
1. An overview of studies on the decapod crustacean cardiac ganglion is given emphasizing contributions to questions of general interest in cellular neurophysiology. 2. John Welsh, in 1951, introduced this 9-celled, semi-autonomous ganglion as a preparation offering physiologists unique experimental possibilities. 3. It exhibits remarkable reliability and stability in rhythmic pattern generation. The neurons show endogenous burst-forming capability mediated by "driver potentials". 4. These regenerative, Ca-mediated potentials are restricted to the soma, while impulse-generating membrane is segregated to the distal axon. 5. Thus, voltage-clamp analysis of the ionic currents underlying the burst-forming potentials is possible by isolating the soma with a ligature. 6. The isolated ganglion is spontaneously active, but the normal mechanism of pacemaking remains to be clarified, including the possible contribution of stretch-sensitive dendrites. 7. The activity of the ganglion is subject to modulation by neurohumors. These include the transmitter at intraganglionic synapses, transmitters of the pair of inhibitory and the two pairs of acceleratory fibers, and neurohormones released from the pericardial organs. The transmitters are not established. 8. Effects on the ganglion of substances isolated from the pericardial organs have been described. 9. These include 5-hydroxytryptamine, dopamine, octopamine, and two peptides. 10. One of these, proctolin, produces a long-lasting sequence of effects. 11. The work continues to raise new questions for which the ganglion offers excellent research material.
Saha, Susmita; Greferath, Ursula; Vessey, Kirstan A; Grayden, David B; Burkitt, Anthony N; Fletcher, Erica L
Inherited retinal degeneration such as retinitis pigmentosa (RP) is associated with photoreceptor loss and concomitant morphological and functional changes in the inner retina. It is not known whether these changes are associated with changes in the density and distribution of synaptic inputs to retinal ganglion cells (RGCs). We quantified changes in ganglion cell density in rd1 and age-matched C57BL/6J-(wildtype, WT) mice using the immunocytochemical marker, RBPMS. Our data revealed that following complete loss of photoreceptors, (∼3months of age), there was a reduction in ganglion cell density in the peripheral retina. We next examined changes in synaptic inputs to A type ganglion cells by performing double labeling experiments in mice with the ganglion cell reporter lines, rd1-Thy1 and age-matched wildtype-Thy1. Ribbon synapses were identified by co-labelling with CtBP2 (RIBEYE) and conventional synapses with the clustering molecule, gephyrin. ON RGCs showed a significant reduction in RIBEYE-immunoreactive synapse density while OFF RGCs showed a significant reduction in the gephyrin-immmunoreactive synapse density. Distribution patterns of both synaptic markers across the dendritic trees of RGCs were unchanged. The change in synaptic inputs to RGCs was associated with a reduction in the number of immunolabeled rod bipolar and ON cone bipolar cells. These results suggest that functional changes reported in ganglion cells during retinal degeneration could be attributed to loss of synaptic inputs. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.
Liu, Jiali; Chen, Dawei; Goldstein, Ronald S; Cui, Sheng
Sex steroids can influence developmental processes and support the survival of neurons in the embryonic central nervous system. Recent studies have shown that estrogen receptors are also expressed in the peripheral nervous system, in the dorsal root ganglia (DRG) of chick embryos. However, no studies have examined the effects of sex steroids on development of embryonic DRG. In the present study, 0.2 microg, 1.0 microg, 5.0 microg 10 microg, 20 microg, 25 microg, and 40 microg doses of testosterone or estradiol were delivered to chick embryos at Hamburger and Hamilton stage 18 (E3). The actions of these doses of sex steroids on the development of the C5DRG (fifth cervical ganglion, a "normal" DRG) and C2DRG (a transient ganglion known as a "Froriep's DRG") were then evaluated by quantifying ganglionic volumes, cell number, proliferation, and apoptosis after 1 day of growth to stage 23. We found that both testosterone and estradiol promoted proliferation of cells in both normal DRG and the Froriep's ganglia. By contrast, estradiol significantly increased the number of apoptotic cells, while testosterone strongly inhibited apoptosis. These actions of sex steroids on DRG development were dose-dependent, and C5DRG and C2DRG showed different sensitivities to the applied sex steroids. In addition, the present results demonstrated that specific ER and AR inhibitors (tamoxifen and flutamide) did not influence the effects of 5 microg E2 and 5 microg T on C2 and C5DRG significantly. These results demonstrate that male and female sex steroids can modulate DRG development through an epigenetic mechanism, as had been shown for the central nervous system.
Stocker, K M; Baizer, L; Ciment, G
Growth associated protein (GAP)-43 is a membrane-bound phosphoprotein expressed in neurons and is particularly abundant during periods of axonal outgrowth in development and regeneration of the nervous system. In previous work, we cloned a full-length chicken GAP-43 cDNA and described the expression of its corresponding mRNA during early development of the chicken nervous system. We report here that the GAP-43 mRNA is also expressed transiently in developing limbs of chicken embryos, which contain axons of spinal cord and dorsal root ganglion neurons, but do not contain neuronal cell bodies. GAP-43 mRNA was first detectable by RNA blot analysis in limbs from Embryonic Day 5 (E5) embryos, reached maximal levels between E6 and E8, and diminished by E10. In situ hybridization analysis showed that the GAP-43 mRNA was localized in distal regions of developing limbs and was particularly abundant in the mesenchyme surrounding the digital cartilage. In some regions of the limb, GAP-43 immunoreactivity colocalized in cells that were also immunoreactive for meromyosin, a muscle-specific marker. These data suggest that both GAP-43 mRNA and the protein are expressed in nonneuronal cells of the developing limb, some of which may be part of the muscle cell lineage.
Levine, M W
Impulse trains simulating the maintained discharges of retinal ganglion cells were generated by digital realizations of the integrate-and-fire model. If the mean rate were set by a "bias" level added to "noise," the variability of firing would be related to the mean firing rate as an inverse square root law; the maintained discharges of retinal ganglion cells deviate systematically from such a relationship. A more realistic relationship can be obtained if the integrate-and-fire mechanism is "leaky"; with this refinement, the integrate-and-fire model captures the essential features of the data. However, the model shows that the distribution of intervals is insensitive to that of the underlying variability. The leakage time constant, threshold, and distribution of the noise are confounded, rendering the model unspecifiable. Another aspect of variability is presented by the variance of responses to repeated discrete stimuli. The variance of response rate increases with the mean response amplitude; the nature of that relationship depends on the duration of the periods in which the response is sampled. These results have defied explanation. But if it is assumed that variability depends on mean rate in the way observed for maintained discharges, the variability of responses to abrupt changes in lighting can be predicted from the observed mean responses. The parameters that provide the best fits for the variability of responses also provide a reasonable fit to the variability of maintained discharges.
Kloos, Anne D; Muller, Kenneth J; Modney, Barbara K
Functional recovery following central nervous system (CNS) injury in adult animals may depend on the reestablishment of the precise pattern of connections made during development. When the nervous system is injured during embryonic development, functional recovery may involve the formation of atypical connections. Can such atypical synapses regenerate in adults, particularly in a nervous system known for its capacity for repair? When the S interneuron in one segmental ganglion of the leech Hirudo is killed during development, two neighboring S cells extend their axons into the ganglion and restore function by making electrical synapses with the usual synaptic targets of the killed S cell. Although adult S-cell axons reliably regenerated their usual synaptic connections, the novel synapses induced following embryonic injury failed to regenerate in adults. In these preparations severed S-cell axons did not reach the denervated ganglion but grew close to it, independent of the distance required to grow. Thus, the developmental changes that permit aberrant but functional connections in embryos do not lead to a similar change in the capacity for axon growth and subsequent synapse regeneration in adults. (c) 2007 Wiley-Liss, Inc.
Van den Dungen, Sophie; Marchesi, Simona; Ezzedine, Rabih; Bindou, David; Lorea, Patrick
Soft tissue ganglion cysts are the most common benign tumours of the wrist; their pathogenesis remains controversial. We prospectively screened the radiographic appearance of the wrists of 51 patients presenting to a single surgeon with dorsal wrist ganglions during a one-year period. Postero-anterior and lateral radiographs were systematically performed looking for possible associated intraosseous ganglion cysts. There were 51 dorsal soft tissue ganglion cysts in 51 patients. We detected 29 associated intraosseous ganglia in 24 patients (47%): 16 ganglia in the lunate bone (55%), 5 in the capitate bone, 7 in the scaphoid and 1 in the trapezoid. Mean size of the intraosseous ganglia was 3 mm (range, 2 to 5 mm). This high prevalence of intraosseous ganglia in association with soft tissue ganglia has to our knowledge never been reported previously. A common aetiology for these two types of ganglion cysts may explain this high association rate.
Sun, Ying-Jie; Kobayashi, Hiroto; Yoshida, Saori; Shirasawa, Nobuyuki; Naito, Akira
In vertebrates, vestibular and cochlear ganglion (VG and CG, respectively) cells are bipolar neurons with myelinated axons and perikarya. The time course of the myelination of the VG and CG cells during development of chick embryos was investigated. Chick VG and CG from embryonic day at 7-20 (E7-20) were prepared for a transmission electron microscopy, myelin basic protein immunohistochemistry, and real-time quantitative RT-PCR. In the VG cells, myelination was first observed on the peripheral axons of the ampullar nerves at E10, on the utricular and saccular nerves at E12, and on the lagenar and neglecta nerves at E13. In the VG central axons, myelination was first seen on the ampullar nerves at E11, on the utricular and saccular nerves at E13, and on the lagenar nerves at E13. In the CG cells, the myelination was first observed on the peripheral and central axons at E14. In both VG and CG, myelination was observed on the perikarya at E17. These results suggest that the onset of the axonal myelination on the VG cells occurred earlier than that on the CG cells, whereas the perikaryal myelination occurred at about the same time on the both types of ganglion cells. Moreover, the myelination on the ampullar nerves occurred earlier than that on the utricular and saccular nerves. The myelination on the peripheral axons occurred earlier than that on the central axons of the VG cells, whereas that on the central and peripheral axons of the CG cells occurred at about the same time. The regional differences in myelination in relation to the onset of functional activities in the VG and CG cells are discussed. Copyright © 2013 ISDN. Published by Elsevier Ltd. All rights reserved.
Linden, R W; Scott, B J
1. In anaesthetized cats recordings have been made in the mesencephalic nucleus of the fifth cranial nerve and the trigeminal ganglion from neurones that respond when forces are applied to the mandibular canine tooth. The site of the mechanoreceptors in the periodontal ligament and their distribution around the tooth root have been determined. 2. Receptors with their cell bodies in the mesencephalic nucleus were found to be situated in the periodontal ligament in a discrete area intermediate between the fulcrum and apex of the tooth, while those in the trigeminal ganglion were situated in the whole area of the periodontal ligament between the fulcrum and apex of the tooth. 3. All of the located mechanoreceptors responded maximally when that part of the ligament in which they lay was put under tension. 4. The directional sensitivities of the mechanoreceptors suggested that there was an uneven distribution around the tooth root of receptors with cell bodies in the mesencephalic nucleus. In contrast mechanoreceptors with cell bodies in the trigeminal ganglion were distributed more equally around the tooth root. The rationale for the differences requires further investigation. PMID:2795482
Hayashida, Yuki; Partida, Gloria J.; Ishida, Andrew T.
We describe here methods for dissociating retinal ganglion cells from adult goldfish and rat without proteolytic enzymes, and show responses of ganglion cells isolated this way to step-wise voltage changes and fluctuating current injections. Taking advantage of the laminar organization of vertebrate retinas, photoreceptors and other cells were lifted away from the distal side of freshly isolated goldfish retinas, after contact with pieces of membrane filter. Likewise, cells were sliced away from the distal side of freshly isolated rat retinas, after these adhered to a membrane filter. The remaining portions of retina were incubated in an enzyme-free, low Ca2+ solution, and triturated. After aliquots of the resulting cell suspension were plated, ganglion cells could be identified by dye retrogradely transported via the optic nerve. These cells showed no obvious morphological degeneration for several days of culture. Perforated-patch whole-cell recordings showed that the goldfish ganglion cells spike tonically in response to depolarizing constant current injections, that these spikes are temporally precise in response to fluctuating current injections, and that the largest voltage-gated Na+ currents of these cells were larger than those of ganglion cells isolated with a neutral protease. PMID:15196824
Coombs, J; van der List, D; Wang, G-Y; Chalupa, L M
The mouse retina offers an increasingly valuable model for vision research given the possibilities for genetic manipulation. Here we assess how the structural properties of mouse retinal ganglion cells relate to the stratification pattern of the dendrites of these neurons within the inner plexiform layer. For this purpose, we used 14 morphological measures to classify mouse retinal ganglion cells parametrically into different clusters. Retinal ganglion cells were labeled in one of three ways: Lucifer Yellow injection, 'DiOlistics' or transgenic expression of yellow fluorescent protein. The resulting analysis of 182 cells revealed 10 clusters of monostratified cells, with dendrites confined to either On or Off sublaminae of the inner plexiform layer, and four clusters of bistratified cells, dendrites spanning the On and Off sublaminae. We also sought to establish how these parametrically identified retinal ganglion cell clusters relate to cell types identified previously on the basis of immunocytochemical staining and the expression of yellow fluorescent protein. Cells labeled with an antibody against melanopsin were found to be located within a single cluster, while those labeled with the SMI-32 antibody were in four different clusters. Yellow fluorescent protein expressing cells were distributed within 13 of the 14 clusters identified here, which demonstrates that yellow fluorescent protein expression is a useful method for labeling virtually the entire population of mouse retinal ganglion cells. Collectively, these findings provide a valuable baseline for future studies dealing with the effects of genetic mutations on the morphological development of these neurons.
Hayashida, Yuki; Partida, Gloria J; Ishida, Andrew T
We describe here methods for dissociating retinal ganglion cells from adult goldfish and rat without proteolytic enzymes, and show responses of ganglion cells isolated this way to step-wise voltage changes and fluctuating current injections. Taking advantage of the laminar organization of vertebrate retinas, photoreceptors and other cells were lifted away from the distal side of freshly isolated goldfish retinas, after contact with pieces of membrane filter. Likewise, cells were sliced away from the distal side of freshly isolated rat retinas, after these adhered to a membrane filter. The remaining portions of retina were incubated in an enzyme-free, low Ca2+ solution, and triturated. After aliquots of the resulting cell suspension were plated, ganglion cells could be identified by dye retrogradely transported via the optic nerve. These cells showed no obvious morphological degeneration for several days of culture. Perforated-patch whole-cell recordings showed that the goldfish ganglion cells spike tonically in response to depolarizing constant current injections, that these spikes are temporally precise in response to fluctuating current injections, and that the largest voltage-gated Na+ currents of these cells were larger than those of ganglion cells isolated with a neutral protease.
Conner, Andrew K; Fung, Kar-Ming; Peterson, Jo Elle G; Glenn, Chad A; Martin, Michael D
Macroscopic ectopic or heterotopic ganglionic tissue within the cauda equina is a very rare pathological finding and is usually associated with spinal dysraphism. However, it may mimic genuine neoplasms of the cauda equina. The authors describe a 29-year-old woman with a history of back pain, right leg pain, and urinary incontinence in whom imaging demonstrated an enhancing mass located in the cauda equina at the L1-2 interspace. The patient subsequently underwent biopsy and was found to have a focus of ectopic ganglionic tissue that was 1.3 cm in greatest dimension. To the authors' knowledge, ectopic or heterotopic ganglionic tissue within the cauda equina in a patient without evidence of spinal dysraphism has never been reported. This patient presented with imaging and clinical findings suggestive of a neoplasm, and an open biopsy proved the lesion to be ectopic ganglionic tissue. The authors suggest that ectopic ganglionic tissue be added to the list of differential diagnoses of a space-occupying lesion arising from the cauda equina.
Osborne, Neville N
Retinal ganglion cell axons within the globe are functionally specialized being richly provided with many mitochondria. The mitochondria produce the high energy requirement for nerve conduction in the unmyelinated part of the ganglion cell axons. We have proposed that in the initiation of glaucoma, an alteration in the quality of blood flow dynamics in the optic nerve head causes a compromise in the retinal ganglion cell axon energy requirement, rendering the ganglion cells susceptible to additional insults. One secondary insult might be light entering the eye to further affect ganglion cell axon mitochondrial function. Other insults to the ganglion cells might be substances (e.g., glutamate, nitric oxide, TNF-alpha) released from astrocytes. These effects ultimately cause ganglion cell death because of the inability of mitochondria to maintain normal function. We therefore suggest that ganglion cell apoptosis in glaucoma is both receptor and mitochondrial mediated. Agents targeted specifically at enhancing ganglion cell mitochondrial energy production should therefore be beneficial in a disease like glaucoma. Ganglion cell death in glaucoma might therefore, in principle, not be unlike the pathophysiology of numerous neurological disorders involving energy dysregulation and oxidative stress. The trigger(s) for ganglion cell apoptosis in glaucoma is/are likely to be multifactorial, and the rationale for targeting impaired energy production as a possibility of improving a patient's quality of life is based on logic derived from laboratory studies where neuronal apoptosis is shown to occur via different mechanisms. Light-induced neuronal apoptosis is likely to be more relevant to ganglion cell death in glaucoma than, for example, neuronal apoptosis associated with Parkinson's disease. Logic suggests that enhancing mitochondrial function generally will slow down ganglion cell apoptosis and therefore benefit glaucoma patients. On the basis of our laboratory studies, we
Spinner, Robert J; Amrami, Kimberly K; Wang, Huan; Kliot, Michel; Carmichael, Stephen W
The appearances of intraneural ganglion cysts are being elucidated. We previously introduced the cross-over phenomenon to explain how a fibular (peroneal) or tibial intraneural ganglion cyst arising from the superior tibiofibular joint could give rise to multiple cysts: cyst fluid ascending up the primarily affected nerve could reach the level of the sciatic nerve, fill its common epineurial sheath and spread circumferentially (cross over), at which time pressure fluxes could result in further ascent up the sciatic or descent down the same parent nerve or the opposite, previously unaffected fibular or tibial nerves. In this study, we hypothesized that cross-over could occur in other nerves, potentially leading to the formation of more than one intraneural ganglion cyst in such situations. We analyzed the literature and identified a single case that we could review where proximal extension of an intraneural ganglion cyst involving a nerve at a different site could theoretically undergo cross-over in another major nerve large enough for available magnetic resonance images to resolve this finding. A case of a suprascapular intraneural ganglion cyst previously reported by our group that arose from the glenohumeral joint and extended to the neck was reanalyzed for the presence or absence of cross-over. An injection of dye into the outer epineurium of the suprascapular nerve in a fresh cadaveric specimen was performed to test for cross-over experimentally. Retrospective review of this case of suprascapular intraneural ganglion cyst demonstrated evidence to support previously unrecognized cross-over at the level of the upper trunk, with predominant ascent up the C5 and the C6 nerve roots and subtle descent down the anterior and posterior divisions of the upper trunk as well as the proximal portion of the suprascapular nerve. This appearance gave rise to multiple interconnected intraneural ganglion cysts arising from a single distant connection to the glenohumeral joint
Adn, M; Hamlat, A; Morandi, X; Guegan, Y
Intraneural ganglion cyst of the tibial nerve is very rare. To date, only 5 cases of this entity in the popliteal fossa have been reported. We report a new case and review the previously reported cases. A 40-year-old man experienced a mild vague pain in the medial half of his right foot for 3 years. Magnetic resonance imaging scan demonstrated a soft-tissue mass along the right tibial nerve. At surgery, an intraneural ganglion cyst was evacuated. After 12 months, the patient was pain-free with no signs of recurrence. Trauma might be a contributing factor to the development of intraneural ganglion cysts. Application of microsurgical techniques is encouraged.
Spinner, Robert J; Carmichael, Stephen W; Wang, Huan; Parisi, Thomas J; Skinner, John A; Amrami, Kimberly K
On the basis of the principles of the unifying articular theory, predictable patterns of proximal ascent have been described for fibular (peroneal) and tibial intraneural ganglion cysts in the knee region. The mechanism underlying distal descent into the terminal branches of the fibular and tibial nerves has not been previously elucidated. The purpose of this study was to demonstrate if and when cyst descent distal to the articular branch-joint connection occurs in intraneural ganglion cysts to understand directionality of intraneural cyst propagation. In Part I, the clinical records and MRIs of 20 consecutive patients treated at our institution for intraneural ganglion cysts (18 fibular and two tibial) arising from the superior tibiofibular joint were retrospectively analyzed. These patients underwent cyst decompression and disconnection of the articular branch. Five of these patients developed symptomatic cyst recurrence after cyst decompression without articular branch disconnection which was done elsewhere prior to our intervention. In Part II, five additional patients with intraneural ganglion cysts (three fibular and two tibial) treated at other institutions without disconnection of the articular branch were compared. These patients in Parts I and II demonstrated ascent of intraneural cyst to differing degrees (12 had evidence of sciatic nerve cross-over). In addition, all of these patients demonstrated previously unrecognized MRI evidence of intraneural cyst extending distally below the level of the articular branch to the joint of origin: cyst within the proximal most portions of the deep fibular and superficial fibular branches in fibular intraneural ganglion cysts and descending tibial branches in tibial intraneural ganglion cysts. The patients in Part I had complete resolution of their cysts at follow-up MRI examination 1 year postoperatively. The patients in Part II had intraneural recurrences postoperatively within the articular branch, the parent
Sarig, Oren; Hass, Avraham; Oron, Amir
Ganglion cysts are considered the most common tumor of the wrist and hand. They are most common between the second and fourth decades of life. The most common anatomical location is the dorsal wrist. This article includes a general review of these cysts including symptoms, pathology and methods of diagnosis, as well as a review of these cysts in specific anatomic locations. The article also includes an updated review of the literature comparing open surgery vs. arthroscopic treatment. The authors believe that arthroscopic surgery of ganglion cysts will gain an important role in the treatment of these cysts.
Matsuda, S; Desaki, J; Aburaya, J; Sakanaka, M
The perikaryal projections of sensory ganglion neurons in chick embryos were observed by scanning electron microscopy after removal of the connective tissues and satellite cells by enzymatic digestion treatment. The perikaryal projections were seen not only on the surface of the perikarya but also on the surface of the stem processes. The projections were up to 3 microm in length, and their transverse diameters ranged between 0.12 and 0.24 microm from incubation (embryonic) day 10 to posthatching day 2. On days 6 and 8 of incubation, thicker projections with transverse diameters of 0.24-0.9 microm were observed transiently in addition to those described above, and some of them looked like vestiges of neuronal processes during development. The thin projections emerging mainly in the later developmental stages increased in number as spindle-shaped bipolar neurons differentiated into (pseudo)unipolar cells. Morphometric analysis revealed that the density of perikaryal projections correlated well with the shape and size of each neuron; thin perikaryal projections were more numerous on those of mature pseudounipolar neurons than on the surface of premature ganglion neurons, and they increased in number as the individual ganglion cell bodies grew larger. The neuronal shape- and size-dependent increase in perikaryal projections during development may support the hypothesis that perikaryal projections are structural devices for increasing neuronal surface areas and possibly the efficiency of metabolic activities.
Subramony, P; Dryer, S E
Chronic blockade of synaptic transmission in ovo using mecamylamine, a neuronal nicotinic receptor antagonist, caused a large increase in naturally occurring cell death in the embryonic chick ciliary ganglion. However, the Ca(2+)-activated K+ currents in embryonic day 13 mecamylamine-treated ciliary ganglion neurons were indistinguishable from those of saline-treated controls. Therefore, the trophic effect of preganglionic innervation on the developmental expression of Ca(2+)-activated K+ current is not dependent upon intact nicotinic cholinergic synaptic transmission and may instead be mediated by a nerve terminal-derived differentiation factor.
Crandall, James E.; Goodman, Timothy; McCarthy, Deirdre M.; Duester, Gregg; Bhide, Pradeep G.; Dräger, Ursula C.; McCaffery, Peter
The ganglionic eminence contributes cells to several forebrain structures including the cerebral cortex, for which it provides GABAergic interneurons. Migration of neuronal precursors from the retinoic-acid rich embryonic ganglionic eminence to the cerebral cortex is known to be regulated by several factors, but retinoic acid has not been previously implicated. We found retinoic acid to potently inhibit cell migration in slice preparations of embryonic mouse forebrains, which was reversed by an antagonist of the dopamine-D2 receptor, whose gene is transcriptionally regulated by retinoic acid. Histonedeacetylase inhibitors, which amplify nuclear receptor-mediated transcription, potentiated the inhibitory effect of retinoic acid. Surprisingly, when retinoic acid signalling was completely blocked with a pan-retinoic acid receptor antagonist, this also decreased cell migration into the cortex, implying that a minimal level of endogenous retinoic acid is necessary for tangential migration. Given these opposing effects of retinoic acid in vitro, the in vivo contribution of retinoic acid to migration was tested by counting GABAergic interneurons in cortices of adult mice with experimental reductions in retinoic acid signalling: a range of perturbations resulted in significant reductions in the numerical density of some GABAergic interneuron subpopulations. These observations suggest functions of retinoic acid in interneuron diversity and organization of cortical excitatory–inhibitory balance. PMID:21895658
Dourado, M M; Brumwell, C; Wisgirda, M E; Jacob, M H; Dryer, S E
The expression of appropriate ensembles of ionic channels is necessary for the differentiation and normal function of vertebrate neurons. Cell-cell interactions may regulate the expression and properties of ionic channels in embryonic neurons. Previous studies have shown that the expression of A-type K+ channels (IA) and Ca2+-activated K+ channels (lK[Ca]) is abnormal in chick ciliary ganglion neurons developing in vitro in the absence of normal cell-cell interactions. Other voltage-activated currents develop normally under these conditions. The present studies were designed to establish the role of the target tissues and the preganglionic innervation in regulating the expression of these currents in embryonic chick ciliary ganglion neurons developing in situ. Surgical manipulations were used to remove the developing optic vesicle, which contains the target tissues, the mid-dorsal region of the midbrain primordium, which contains the preganglionic nucleus, or both, all prior to the formation of the ciliary ganglion. IA and IK[Ca] were then examined in acutely isolated neurons that developed in ovo in the presence (OV+) or absence (OV-) of the normal target tissues, in the presence (MB+) or absence (MB-) of preganglionic innervation, and in the absence of both preganglionic innervation and target tissues (OV-/MB-). The amplitude of IA was unaffected by the operations. However, the activation and inactivation kinetics of IA were two- to threefold faster in OV- or OV-/MB- cells compared to neurons isolated from control OV+ ganglia at embryonic days 11-14 (E11-E14). There were no changes in the voltage dependence of activation or steady-state inactivation, or in the time course of recovery from inactivation. By contrast, neurons isolated from MB- ganglia expressed an IA with amplitude, voltage dependence, and kinetics that were indistinguishable from those of control MB+ and OV+ ganglia. Therefore, interactions with target tissues in the eye play a role in determining
Richards, K S; Marder, E
The motor patterns produced by the stomatogastric ganglion (STG) are strongly influenced by descending modulatory inputs from anterior ganglia. With these inputs intact, in control saline, the motor patterns produced by the stomatogastric nervous system of embryonic and larval lobsters are slower and less regular than those of adult lobsters. We studied the effects of the hormonal modulator, crustacean cardioactive peptide (CCAP) on the discharge patterns of STG motor patterns in embryos, larvae, and adult Maine lobsters, Homarus americanus, with the anterior inputs present and absent. In adults, CCAP initiated robust pyloric rhythms from STGs isolated from their descending control and modulatory inputs. Likewise, CCAP initiated robust activity in isolated embryonic and larval STGs. Nonetheless, quantitative analyses revealed that the frequency and regularity of the STG motor neuron discharge seen in the presence of CCAP in isolated STGs from embryos were significantly lower than those seen late in larval life and in adults under the same conditions. In contrast, when the descending control and modulatory pathways to the STG were left intact, the embryonic and larval burst frequency seen in the presence of CCAP was increased by CCAP, whereas the burst frequency in adults was decreased by CCAP, so that in CCAP the frequencies at all stages were statistically indistinguishable. These data argue that immature embryonic motor patterns seen in the absence of CCAP are a function of immaturity in both the STG and in the descending and modulatory pathways.
Kathryn Robbins; Philip M. Wargo
Root rots of central hardwoods are diseases caused by fungi that infect and decay woody roots and sometimes also invade the butt portion of the tree. By killing and decaying roots, root rotting fungi reduce growth, decrease tree vigor, and cause windthrow and death. The most common root diseases of central hardwoods are Armillaria root rot, lnonotus root rot, and...
and cardiovascular function during the neostigmine pressor response have been studied during the period of this report. Area 1 is the study of pre...and post ganglionic nerve potentials. Area 2 is a study of the peripheral vascular tone during the neostigmine pressor response.
Schmerber, Sébastien; Vasdev, Ashok; Chahine, Karim; Tournaire, Romain; Bing, Fabrice
To identify petrous internal carotid bleeding aneurysm as a complication of gasserian ganglion thermocoagulation. A single case presenting with epistaxis and otorrhagia 1 month after gasserian ganglion thermocoagulation in the treatment of refractory trigeminal neuralgia. Gasserian ganglion thermocoagulation, computed tomographic scan, and angiocomputed tomographic scan revealing petrous internal carotid ruptured aneurysm and internal carotid embolization. Radiologic diagnosis of the vascular injury after gasserian ganglion thermocoagulation. Radiologic identification of ruptured internal carotid artery as the cause of simultaneous epistaxis and otorrhagia. Gasserian ganglion thermocoagulation may cause aneurysm and rupture of the petrous portion of the internal carotid artery.
Cairns, Brian E; O'Brien, Melissa; Dong, Xu-Dong; Gazerani, Parisa
It has been proposed that after nerve injury or tissue inflammation, fractalkine (CX3CL1) released from dorsal root ganglion neurons acts on satellite glial cells (SGCs) through CX3C receptor 1 (CX3CR1) to induce neuroplastic changes. The existence and importance of fractalkine/CX3CR1 signaling in the trigeminal ganglia has not yet been clarified. This study investigated (1) whether trigeminal ganglion neurons that innervate temporalis muscle and their associated SGCs contain fractalkine and/or express CX3CR1, (2) if intraganglionic injection of fractalkine increases the mechanical sensitivity of temporalis muscle afferent fibers, (3) whether complete Freund's adjuvant (CFA)-induced inflammation of the temporalis muscle alters the expression of fractalkine or its receptor in the trigeminal ganglion, and (4) if intraganglionic administration of CX3CR1 antibodies alters afferent mechanical sensitivity. Immunohistochemistry and in vivo electrophysiological recordings in male and female rats were used to address these questions. It was found that ∼50 % of temporalis ganglion neurons and ∼25 % of their associated SGCs express CX3CR1, while only neurons expressed fractalkine. Temporalis muscle inflammation increased the expression of fractalkine, but only in male rats. Intraganglionic injection of fractalkine (25 g/ml; 3 μl) induced prolonged afferent mechanical sensitization. Intraganglionic injection of CX3CR1 antibody increased afferent mechanical threshold, but this effect was greater in controls than in rats with CFA-induced muscle inflammation. These findings raise the possibility that basal fractalkine signalling within the trigeminal ganglion plays an important role in mechanical sensitivity of masticatory muscle sensory afferent fibers and that inhibition of CX3CR1 signaling within the trigeminal ganglia may induce analgesia through a peripheral mechanism.
Cervical sympathetic and stellate ganglion blocks (SGB) provide a valuable diagnostic and therapeutic benefit to sympathetically maintained pain syndromes in the head, neck, and upper extremity. With the ongoing efforts to improve the safety of the procedure, the techniques for SGB have evolved over time, from the use of the standard blind technique, to fluoroscopy, and recently to the ultrasound (US)-guided approach. Over the past few years, there has been a growing interest in the ultrasound-guided technique and the many advantages that it might offer. Fluoroscopy is a reliable method for identifying bony surfaces, which facilitates identifying the C6 and C7 transverse processes. However, this is only a surrogate marker for the cervical sympathetic trunk. The ideal placement of the needle tip should be anterolateral to the longus colli muscle, deep to the prevertebral fascia (to avoid spread along the carotid sheath) but superficial to the fascia investing the longus colli muscle (to avoid injecting into the muscle substance). Identifying the correct fascial plane can be achieved with ultrasound guidance, thus facilitating the caudal spread of the injectate to reach the stellate ganglion at C7-T1 level, even if the needle is placed at C6 level. This allows for a more effective and precise sympathetic block with the use of a small injectate volume. Ultrasound-guided SGB may also improve the safety of the procedure by direct visualization of vascular structures (inferior thyroidal, cervical, vertebral, and carotid arteries) and soft tissue structures (thyroid, esophagus, and nerve roots). Accordingly, the risk of vascular and soft tissue injury may be minimized.
Lodato, Simona; Tomassy, Giulio Srubek; De Leonibus, Elvira; Uzcategui, Yoryani G; Andolfi, Gennaro; Armentano, Maria; Touzot, Audrey; Gaztelu, Jose M; Arlotta, Paola; Menendez de la Prida, Liset; Studer, Michèle
In rodents, cortical interneurons originate from the medial ganglionic eminence (MGE) and caudal ganglionic eminence (CGE) according to precise temporal schedules. The mechanisms controlling the specification of CGE-derived interneurons and their role in cortical circuitry are still unknown. Here, we show that COUP-TFI expression becomes restricted to the dorsal MGE and CGE at embryonic day 13.5 in the basal telencephalon. Conditional loss of function of COUP-TFI in subventricular precursors and postmitotic cells leads to a decrease of late-born, CGE-derived, VIP (vasoactive intestinal peptide)- and CR (calretinin)-expressing bipolar cortical neurons, compensated by the concurrent increase of early-born MGE-derived, PV (parvalbumin)-expressing interneurons. Strikingly, COUP-TFI mutants are more resistant to pharmacologically induced seizures, a phenotype that is dependent on GABAergic signaling. Together, our data indicate that COUP-TFI controls the delicate balance between MGE- and CGE-derived cortical interneurons by regulating intermediate progenitor divisions and ultimately affecting the activity of the cortical inhibitory circuitry.
Vernino, Steven; Lindstrom, Jon; Hopkins, Steve; Wang, Zhengbei; Low, Phillip A.
In myasthenia gravis (MG), autoantibodies bind to the α1 subunit and other subunits of the muscle nicotinic acetylcholine receptor (AChR). Autoimmune autonomic ganglionopathy (AAG) is an antibody-mediated neurological disorder caused by antibodies against neuronal AChRs in autonomic ganglia. Subunits of muscle and neuronal AChR are homologous. We examined the specificity of AChR antibodies in patients with MG and AAG. Ganglionic AChR autoantibodies found in AAG patients are specific for AChRs containing the α3 subunit. Muscle and ganglionic AChR antibody specificities are distinct. Antibody crossreactivity between AChRs with different α subunits is uncommon but can occur. PMID:18485491
Rücker, M.; Berg, S.; Armstrong, R. T.; Georgiadis, A.; Ott, H.; Schwing, A.; Neiteler, R.; Brussee, N.; Makurat, A.; Leu, L.; Wolf, M.; Khan, F.; Enzmann, F.; Kersten, M.
During imbibition, initially connected oil is displaced until it is trapped as immobile clusters. While initial and final states have been well described before, here we image the dynamic transient process in a sandstone rock using fast synchrotron-based X-ray computed microtomography. Wetting film swelling and subsequent snap off, at unusually high saturation, decreases nonwetting phase connectivity, which leads to nonwetting phase fragmentation into mobile ganglia, i.e., ganglion dynamics regime. We find that in addition to pressure-driven connected pathway flow, mass transfer in the oil phase also occurs by a sequence of correlated breakup and coalescence processes. For example, meniscus oscillations caused by snap-off events trigger coalescence of adjacent clusters. The ganglion dynamics occurs at the length scale of oil clusters and thus represents an intermediate flow regime between pore and Darcy scale that is so far dismissed in most upscaling attempts.
Depace, D M
The permeability of the blood vessels in the superior cervical ganglion of the rat was tested by intravenous injection of horseradish peroxidase (HRP). By light microscopy, peroxidase activity was found in three locations: in the capsule of the ganglion, in the lumina of the blood vessels, and within macrophages. Electron microscopy revealed that virtually all ganglionic blood vessels contained HRP 5 minutes following its administration. The intensity of peroxidase activity declined over the period of 15 minutes. The enzyme was localized on the luminal surface of the endothelial cells, attaching to the glycocalyx. Endothelial microvilli, projecting into the vessel lumen, were also covered with peroxidase. Micropinocytotic vesicles on the luminal surface of the endothelium contained reaction product. Some of these vesicles were free within the cytoplasm of the endothelium but none was observed on the abluminal surface. Peroxidase activity was not detected in the extracellular space even after 15 minutes. The majority of blood vessels in the superior cervical ganglion possess a continuous endothelium with tight junctions; features associated with the blood-brain barrier of the central nervous system and peripheral nerves. It is proposed that these vessels perform a barrier function between the capillary circulation and the superior cervical ganglion.
Ahumada, Albert J.
Unsupervised learning models have been proposed based on experience (Ahumada and Mulligan, 1990;Wachtler, Doi, Lee and Sejnowski, 2007) that allow the cortex to develop units with LM specific color opponent receptive fields like the blob cells reported by Hubel and Wiesel on the basis of visual experience. These models used ganglion cells with LM indiscriminate wiring as inputs to the learning mechanism, which was presumed to occur at the cortical level.
Liao, Chenlong; Yang, Min; Liu, Pengfei; Zhong, Wenxiang; Zhang, Wenchuan
Abstract Rationale: Thalamic pain is a distressing and treatment-resistant type of central post-stroke pain. Although stellate ganglion block is an established intervention used in pain management, its use in the treatment of thalamic pain has never been reported. Patient concerns: A 66-year-old woman presented with a 3-year history of severe intermittent lancinating pain on the right side of the face and the right hand. The pain started from the ulnar side of the right forearm after a mild ischemic stroke in bilateral basal ganglia and left thalamus. Weeks later, the pain extended to the dorsum of the finger tips and the whole palmar surface, becoming more severe. Meanwhile, there was also pain with similar characteristics emerging on her right face, resembling atypical trigeminal neuralgia. Diagnoses: Thalamic pain was diagnosed. Interventions: After refusing the further invasive treatment, she was suggested to try stellate ganglion block. Outcomes: After a 3-day period of pain free (numerical rating scale: 0) postoperatively, she reported moderate to good pain relief with a numerical rating scale of about 3 to 4 lasting 1 month after the first injection. Pain as well as the quality of life was markedly improved with less dose of analgesic agents. Lessons: Stellate ganglion block may be an optional treatment for thalamic pain. PMID:28151918
Shetty, Gautam M; Nha, Kyung Wook; Patil, Sachin P; Chae, Dong Ju; Kang, Ki Hoon; Yoon, Jung Ro; Choo, Suk Kyu; Yi, Jeong Woo; Kim, Ji Hoon; Baek, Jong Ryoon
Ganglion cysts of the posterior cruciate ligament (PCL) are uncommon lesions found incidentally on MRI and arthroscopy. Twenty patients (11 males and nine females) with the mean age of 35 years presenting with a variety of knee signs and symptoms were found to have PCL cysts on MRI. Out of these, thirteen patients (65%) had isolated symptomatic PCL cysts and seven patients had associated chondral and meniscal lesions. Eight out of the 20 patients (40%) gave a history of antecedent trauma. On arthroscopy, the majority of the cysts were situated at the midsubstance of the ligament with inter-cruciate distension and no involvement of the substance of the ligament. The content of the cysts varied with the majority having yellowish viscous fluid and three containing serous and bloody fluid. All cysts were successfully treated arthroscopically through standard anterior, posteromedial and posterolateral portals with no signs of recurrence on MRI at a mean followup of 24 months. PCL cysts may clinically mimic meniscal or chondral lesions and preoperatively, MRI is essential for the diagnosis of ganglion cysts arising from the PCL. Ganglion cysts of the PCL can be successfully treated arthroscopically using standard portals.
Harahush, Blake K; Hart, Nathan S; Collin, Shaun P
The development of the visual system in anamniotic vertebrates is a continual process, allowing for ontogenetic changes in retinal topography and spatial resolving power. We examined the number and distribution of retinal ganglion cells in wholemounted retinae throughout the protracted embryonic development (∼5 months) of a chondrichthyan, i.e. the brown-banded bamboo shark Chiloscyllium punctatum, from the beginning of retinal cell differentiation (approximately halfway through embryogenesis) to adulthood. We also identified and quantified the number of apoptosed cells within the ganglion cell layer to evaluate the contribution of apoptosis to changes in retinal topography. C. punctatum undergoes rapid changes in ganglion cell distribution during embryogenesis, where high levels of apoptosis, especially around the retinal periphery, result in relative increases in ganglion cell density in the central retina which progressively extend nasally and temporally to form a meridional band at hatching. After hatching, C. punctatum forms and maintains a horizontal streak, showing only minor changes in topography during growth, with basal levels of apoptosis. The total number of retinal ganglion cells reaches 547,881 in adult sharks, but the mean (3,228 cells·mm(-2)) and peak (4,983 cells·mm(-2)) retinal ganglion cell densities are highest around the time of hatching. Calculated estimates of spatial resolving power, based on ganglion cell spacing (assuming a hexagonal mosaic) and assessment of the focal length from cryosections of the eye, increase from 1.47 cycles·degree(-1) during embryogenesis to 4.29 cycles·degree(-1) in adults. The increase in spatial resolving power across the retinal meridian would allow this species to hunt and track faster, more mobile prey as it reaches maturity.
Quilliam, J. P.; Shand, D. G.
By comparing the effects on ganglionic transmission and on the pre- and post-ganglionic nerves in the isolated superior cervical ganglion preparation of the rat, the selectivity of several drugs was assessed quantitatively. Hexamethonium, tetraethylammonium, nicotine and tubocurarine blocked transmission in concentrations which did not affect nervous conduction and were considered to be highly selective in action. Atropine, amylobarbitone and paraldehyde depressed nervous conduction appreciably in ganglion-blocking doses, but not enough to account wholly for the block in transmission and they were therefore considered as being moderately selective. The ganglion blocking actions of mephenesin, procaine, methylpentynol, methylpentynol carbamate and benactyzine were nonspecific, showing general depression of neuronal activity. Ganglion block with bretylium was nonselective in its site of depression of the postganglionic neurone in concentrations which only partly depressed the preganglionic nerve. PMID:14228129
Demircay, Emre; Ofluoglu, Demet; Ozel, Omer; Oztop, Pinar
Intra-articular ganglion cysts of the anterior cruciate ligament (ACL) are rare, and bilateral ganglion cysts are even rarer. These cysts may cause intermittent or chronic nonspecific knee discomfort. Although three cases of bilateral ganglion cysts have been reported in the literature, the knees were not simultaneously affected in those cases. Herein, we report the case of a 56-year-old woman who presented with simultaneous bilateral ganglion cysts of the ACL that were symptomatic. She was successfully treated with arthroscopic resection and debridement. We also present a brief review of the literature, highlighting the aetiology, diagnosis and management of ganglion cysts of the ACL. To the best of our knowledge, this is the first report of simultaneous bilateral intra-articular ganglion cysts of the ACL.
Kapral, S; Krafft, P; Gosch, M; Fridrich, P; Weinstabl, C
We present a patient who developed a high central neural block after stellate-ganglion-blockade. The underlying pathophysiology was assessed via sonographic imaging. Stellate ganglion block was performed in a 19-year old patient according to the standard technique. Multiple aspiration tests were negative and a test dose of 3 ml bupivacaine 0.25% was injected. After a 3 min interval another 5 ml were injected. Two minutes after the local anaesthetic administration the patient reported nausea and sensations in the upper extremity. Spontaneous respiration efforts stopped, and the patient became unconscious. Tracheal intubation was performed, and the patient was ventilated in a controlled mode for two hours. Heart rate as well as blood pressure remained within the normal range. Neurologic recovery occurred rapidly and extubation was performed about two and a half hours after the event. Our sonographic studies demonstrated a local anaesthetic depot directly at the root of C 6, with a mean diameter of 10 mm and a length of 5 to 6 cm (about a third smaller than expected). Sonographic studies and clinical symptoms of our patient are most likely to occur with a subdural extra-arachnoidal block. Ultrasonographic guided puncture enhances the patient's safety by the opportunity to directly visualise transverse process, nerval root as well as local anaesthetic depot. In case of depot formation directly at the nerval root, injection may be stopped and the needle repositioned. Furthermore, direct visualisation of the great vessels (A. vertebralis) prevents intravascular injection and haematoma formation.
Kongstaponkit, S; Pradidarcheep, W; Toutip, S; Chunhabundit, P; Somana, R
Since there is only a limited number of studies of the blood supply to the trigeminal ganglion (TG) in mammalian species, the TG from 16 common tree shrews (Tupaia glis) were investigated by light microscope, transmission electron microscope (TEM) and the corrosion cast technique in conjunction with scanning electron microscope (SEM). It was found that the TG contained clusters of neurons in the peripheral region whereas the bundles of nerve fibers were located more centrally. Each ganglionic neuron had a concentric nucleus and was ensheathed by satellite cells. It was noted that blood vessels of a continuous type were predominantly found in the area where the neurons were densely located and were much less frequently observed in the area occupied by nerve fibers. With TEM, the TG was shown to be mainly associated with large neurons containing big nuclei and prominent nucleoli. The blood supply of the TG is derived from the most rostral branch of the pontine artery, from the stapedial artery or sometimes from the supraorbital artery, and from the accessory meningeal artery which is a branch of the maxillary artery passing through the foramen ovale. These arteries give off branches and become capillary networks in the ganglion before draining blood to the peripheral region. The veins at the medial border drained into the cavernous sinus directly or through the inferior hypophyseal vein, while those at the lateral side of the ganglion carried the blood into the pterygoid plexus via an accessory meningeal vein. The veins along the trigeminal nerve root joined the posterior part of the cavernous sinus. These studies establish a unique anatomical distribution of the TG blood supply in the tree shrew and the utility of the cast/SEM technique in discerning detailed features of the blood supply in the nervous system.
Gatta, Gemma; Ferrari, Andrea; Stiller, Charles A; Pastore, Guido; Bisogno, Gianni; Trama, Annalisa; Capocaccia, Riccardo
Embryonal cancers are a heterogeneous group of rare cancers which mainly occur in children and adolescents. The aim of the present study was to estimate the burden (incidence, prevalence, survival and proportion of cured) for the principal embryonal cancers in Europe (EU27), using population-based data from cancer registries (CRs) participating in RARECARE. We identified 3322 cases diagnosed from 1995 to 2002 (latest period for which data are available): 44% neuroblastoma, 35% nephroblastoma, 13% retinoblastoma and 6% hepatoblastoma. Very few cases of pulmonary blastoma (43 cases) and pancreatoblastoma (seven cases) were diagnosed. About 2000 new embryonal cancers were estimated every year in EU27, for an annual incidence rate of 4 per million (1.8 neuroblastoma, 1.4 nephroblastoma, and 0.5 retinoblastoma); 91% of cases occurred in patients under 15 years. Five-year relative survival for all embryonal cancers was 80% (99% retinoblastoma, 90% nephroblastoma, 71% hepatoblastoma and 68% neuroblastoma). Overall survival was lower in adolescents and adults than in those under 15 years. The cure rate was estimated at 80%. Slightly less than 40,000 persons were estimated alive in EU27 with a diagnosis of embryonal cancer in 2008. Nephroblastoma was the most prevalent (18,150 cases in EU27), followed by neuroblastoma (12,100), retinoblastoma (5200), hepatoblastoma (2700) and pulmonary blastoma (614). This is the first study to delineate the embryonal cancer burden in Europe by age, sex and European region. Survival/cure rate is generally high, but there are considerable gaps in our understanding of the natural histories of these rare diseases particularly in adults.
Roehm, Pamela C; Hansen, Marlan R
Degeneration of spiral ganglion neurons following hair cell loss carries critical implications for efforts to rehabilitate severe cases of hearing loss with cochlear implants or hair cell regeneration. This review considers recently identified neurotrophic factors and therapeutic strategies which promote spiral ganglion neuron survival and neurite growth. Replacement of these factors may help preserve or regenerate the auditory nerve in patients with extensive hair cell loss. Spiral ganglion neurons depend on neurotrophic factors supplied by hair cells and other targets for their development and continued survival. Loss of this trophic support leads to spiral ganglion neuron death via apoptosis. Hair cells support spiral ganglion neuron survival by producing several peptide neurotrophic factors such as neurotrophin-3 and glial derived neurotrophic factor. In addition, neurotransmitter release from the hair cells drives membrane electrical activity in spiral ganglion neurons which also supports their survival. In animal models, replacement of peptide neurotrophic factors or electrical stimulation with an implanted electrode attenuates spiral ganglion neuron degeneration following deafferentation. Cell death inhibitors can also preserve spiral ganglion neuron populations. Preliminary studies show that transfer of stem cells or neurons from other ganglia are two potential strategies to replace lost spiral ganglion neurons. Inducing the regrowth of spiral ganglion neuron peripheral processes to approximate or contact cochlear implant electrodes may help optimize signaling from a diminished population of neurons. Recent studies of spiral ganglion neuron development and survival have identified several trophic and neuritogenic factors which protect these specialized cells from degeneration following hair cell loss. While still preliminary, such strategies show promise for future clinical applications.
Meister, Markus; Lagnado, Leon; Baylor, Denis A.
To analyze the rules that govern communication between eye and brain, visual responses were recorded from an intact salamander retina. Parallel observation of many retinal ganglion cells with a microelectrode array showed that nearby neurons often fired synchronously, with spike delays of less than 10 milliseconds. The frequency of such synchronous spikes exceeded the correlation expected from a shared visual stimulus up to 20-fold. Synchronous firing persisted under a variety of visual stimuli and accounted for the majority of action potentials recorded. Analysis of receptive fields showed that concerted spikes encoded information not carried by individual cells; they may represent symbols in a multineuronal code for vision.
Medina, Juana; Rivlin, Michael; Chan, Joanna; Beredjiklian, Pedro K
Ganglion cysts are the most common wrist tumors, and 60 -70% originate dorsally from the scapholunate interval. Ossification of these lesions is exceedingly rare, with only one such lesion located in the finger reported in the literature. We present a case of an ossified dorsal wrist ganglion in a 68-year-old woman.
Medina, Juana; Rivlin, Michael; Chan, Joanna; Beredjiklian, Pedro K.
Ganglion cysts are the most common wrist tumors, and 60 -70% originate dorsally from the scapholunate interval. Ossification of these lesions is exceedingly rare, with only one such lesion located in the finger reported in the literature. We present a case of an ossified dorsal wrist ganglion in a 68-year-old woman. PMID:27847858
Jose, Jean; O'Donnell, Kevin; Lesniak, Bryson
Ganglion cysts have been previously described throughout the body, most commonly about the wrist, hand, knee, ankle, and feet. When symptomatic, they may interfere with joint mechanics, resulting in snapping, catching, and locking. Intratendinous ganglion cysts lack a synovial epithelial lining and are thought to develop from the mucoid degeneration of connective tissue caused by chronic irritation, chronic repetitive injury, and chronic ischemia. On magnetic resonance imaging, ganglion cysts originating from tendons, ligaments, tendon sheaths, menisci, or joint capsules appear as well-defined lobulated masses that follow simple or complex fluid signal intensity on all pulse sequences, with enhancing walls and internal septations on post-contrast images. There may be appreciable degeneration and partial tearing of the structure of origin, particularly if associated with tendons. On ultrasonography, they present as hypoechoic masses, with internal septations and lobulations of varying sizes, without significant vascularity on power or color Doppler sampling. A thin fluid neck extending from the structure of origin (tail sign), when present, is a reliable sign of a ganglion cyst. This article describes a sonographically guided technique to treat symptomatic ganglion cysts within the patellar tendon. Complete evacuation of the ganglion cyst, with disappearance of the tail sign, is considered the determining factor for a successful procedure. A similar technique can be used for the treatment of other symptomatic intratendinous ganglion cysts elsewhere in the body. To our knowledge, symptomatic intratendinous ganglion cysts within the patellar tendon and their treatment have not been previously reported.
Willis-Owen, Charles A; Konyves, Arpad; Martin, David K
Symptomatic ganglion cysts of the cruciate ligaments are rare, and bilateral cases are extremely rare, with only one reported case in the literature. We report a case of bilateral cruciate ligament ganglion cysts successfully treated with arthroscopic resection, and review the literature regarding aetiology, diagnosis and management.
Prasad, Nikhil K; Spinner, Robert J; Smith, Jay; Howe, Benjamin M; Amrami, Kimberly K; Iannotti, Joseph P; Dahm, Diane L
OBJECT High-resolution magnetic resonance imaging (MRI) can distinguish between intraneural ganglion cysts and paralabral (extraneural) cysts at the glenohumeral joint. Suprascapular intraneural ganglion cysts share the same pathomechanism as their paralabral counterparts, emanating from a tear in the glenoid labrum. The authors present 2 cases to demonstrate that the identification and arthroscopic repair of labral tears form the cornerstone of treatment for intraneural ganglion cysts of the suprascapular nerve. METHODS Two patients with suprascapular intraneural ganglion cysts were identified: 1 was recognized and treated prospectively, and the other, previously reported as a paralabral cyst, was identified retrospectively through the reinter-pretation of high-resolution MR images. RESULTS Both patients achieved full functional recovery and had complete radiological involution of the intraneural ganglion cysts at the 3-month and 12-month follow-ups, respectively. CONCLUSIONS Previous reports of suprascapular intraneural ganglion cysts described treatment by an open approach to decompress the cysts and resect the articular nerve branch to the glenohumeral joint. The 2 cases in this report demonstrate that intraneural ganglion cysts, similar to paralabral cysts, can be treated with arthroscopic repair of the glenoid labrum without resection of the articular branch. This approach minimizes surgical morbidity and directly addresses the primary etiology of intraneural and extraneural ganglion cysts.
Tai, Huanhuan; Lu, Xin; Opitz, Nina; Marcon, Caroline; Paschold, Anja; Lithio, Andrew; Nettleton, Dan; Hochholdinger, Frank
Maize develops a complex root system composed of embryonic and post-embryonic roots. Spatio-temporal differences in the formation of these root types imply specific functions during maize development. A comparative transcriptomic study of embryonic primary and seminal, and post-embryonic crown roots of the maize inbred line B73 by RNA sequencing along with anatomical studies were conducted early in development. Seminal roots displayed unique anatomical features, whereas the organization of primary and crown roots was similar. For instance, seminal roots displayed fewer cortical cell files and their stele contained more meta-xylem vessels. Global expression profiling revealed diverse patterns of gene activity across all root types and highlighted the unique transcriptome of seminal roots. While functions in cell remodeling and cell wall formation were prominent in primary and crown roots, stress-related genes and transcriptional regulators were over-represented in seminal roots, suggesting functional specialization of the different root types. Dynamic expression of lignin biosynthesis genes and histochemical staining suggested diversification of cell wall lignification among the three root types. Our findings highlight a cost-efficient anatomical structure and a unique expression profile of seminal roots of the maize inbred line B73 different from primary and crown roots.
Tai, Huanhuan; Lu, Xin; Opitz, Nina; Marcon, Caroline; Paschold, Anja; Lithio, Andrew; Nettleton, Dan; Hochholdinger, Frank
Maize develops a complex root system composed of embryonic and post-embryonic roots. Spatio-temporal differences in the formation of these root types imply specific functions during maize development. A comparative transcriptomic study of embryonic primary and seminal, and post-embryonic crown roots of the maize inbred line B73 by RNA sequencing along with anatomical studies were conducted early in development. Seminal roots displayed unique anatomical features, whereas the organization of primary and crown roots was similar. For instance, seminal roots displayed fewer cortical cell files and their stele contained more meta-xylem vessels. Global expression profiling revealed diverse patterns of gene activity across all root types and highlighted the unique transcriptome of seminal roots. While functions in cell remodeling and cell wall formation were prominent in primary and crown roots, stress-related genes and transcriptional regulators were over-represented in seminal roots, suggesting functional specialization of the different root types. Dynamic expression of lignin biosynthesis genes and histochemical staining suggested diversification of cell wall lignification among the three root types. Our findings highlight a cost-efficient anatomical structure and a unique expression profile of seminal roots of the maize inbred line B73 different from primary and crown roots. PMID:26628518
Egbogah, E.O.; Wright, R.J.; Dawe, R.A.
This paper presents a simple theory of the movement of a discontinuous oil droplet (ganglion) through a model porous medium. A quantitative description of the ganglion flow in the system was obtained through a tractable solution to the balance of forces controlling ganglion stability during flow of two immiscible fluids within a well-defined geometry. Calculations were based on a constricted conical (divergent-convergent) pore model. Experimental data from a tetragonally packed sphere model were used interactively with a theoretical static analysis to synthesize the relevant features of the ganglion mechanics into a coherent theory of oil mobilization. The model analysis also permits the computation of relative ganglion velocity under various flow conditions. This is an essential parameter for enhanced oil recovery modelling which facilitates the prediction of oil bank movements in porous media. 34 refs.
Shimokobe, Hisao; Gotoh, Masafumi; Mitsui, Yasuhiro; Yoshikawa, Eiichiro; Kume, Shinichiro; Okawa, Takahiro; Higuchi, Fujio; Nagata, Kensei; Shiba, Naoto
Studies have demonstrated favorable outcomes of arthroscopic decompression for ganglion cyst in the supraspinous fossa; however, little attention has been paid to the difficulty in detecting these cysts during arthroscopy. In this report, we present 2 cases in which ganglion cysts in the supraspinous fossa were undetectable during arthroscopy. The ganglion cysts were not identified in these cases during surgery despite arthroscopic decompression being performed through the area in which the cyst was expected until the suprascapular nerve was entirely exposed. After surgery, magnetic resonance imaging (MRI) confirmed the disappearance of the ganglion cyst and external rotation strength was fully improved, without shoulder pain. We emphasize here that surgeons should be aware of this difficulty when performing arthroscopic decompression of ganglion cysts in the supraspinous fossa.
Lira-Ruan, Verónica; Ruiz-Kubli, Mariel
Rice (Oryza sativa) contains five copies of the non-symbiotic hemoglobin (hb) gene, namely hb1 to hb5. Previous analysis by RT-PCR revealed that rice hb1 expresses in roots and leaves and hb2 expresses in leaves. However, it is not known whether or not hb1 and hb2 express in rice embryonic organs. Here, we report the expression of hb1 and hb2 genes in rice embryonic organs using RT-PCR and specific oligos for Hb1 and Hb2. Our results indicate that hb1 and hb2 genes express in embryonic organs in rice growing under normal conditions. Specifically, hb1 expresses in rice embryos and seminal roots, and hb2 expresses in embryos, coleoptiles and seminal roots. These observations suggest that Hb1 and Hb2 coexist and function in rice embryonic organs. PMID:21966570
Boada, M Danilo
The trigeminal ganglia (TG) innervate a heterogeneous set of highly sensitive and exposed tissues. Weak, innocuous stimuli can evoke pain as a normal response in some areas such as the cornea. This observation implies, however, the capability of low-threshold mechanoreceptors, inducing pain in the normal condition. To clarify this matter, the present study correlates the electrical signature (both fiber conduction velocity and somatic electrical properties) with receptor field, mechanical threshold, and temperature responsiveness of sensory afferents innervating tissues with dissimilar sensitivity (skin vs. cornea) in the trigeminal domain. Intracellular recordings were obtained in vivo from 148 neurons of the left TG of 62 mice. In 111 of these neurons, the peripheral receptor field was successfully localized: 96 of them innervated the hairy skin, while the remaining 15 innervated the cornea. The electrical signature was defined and peripheral responses correlated with tissue target. No high threshold neurons were found in the cornea. Moreover, the electrical signature of corneal afferents resembles nociceptive neurons in the skin. TG skin afferents showed similar membrane electrical signature and sensory modality as skin afferents from dorsal root ganglion, although TG afferents exhibited a shorter duration of afterhyperpolarization then those previously described in dorsal root ganglion. These data suggest than new or different ways to classify and study TG sensory neurons may be required.
Kitazawa, Ayako; Shimizu, Norio
Mouse induced pluripotent stem (iPS) cells are known to have the ability to differentiate into various cell lineages including neurons in vitro. We have reported that chick dorsal root ganglion (DRG)-conditioned medium (CM) promoted the differentiation of mouse embryonic stem (ES) cells into motor neurons. We investigated the formation of undifferentiated iPS cell colonies and the differentiation of iPS cells into neurons using DRG-CM. When iPS cells were cultured in DMEM containing leukemia inhibitory factor (LIF), the iPS cells appeared to be maintained in an undifferentiated state for 19 passages. The number of iPS cell colonies (200 μm in diameter) was maximal at six days of cultivation and the colonies were maintained in an undifferentiated state, but the iPS cell colonies at ten days of cultivation had hollows inside the colonies and were differentiated. By contrast, the number of ES cell colonies (200 μm in diameter) was maximal at ten days of cultivation. The iPS cells were able to proliferate and differentiate easily into various cell lineages, compared to ES cells. When iPS cell colonies were cultured in a manner similar to ES cells with DMEM/F-12K medium supplemented with DRG-CM, the iPS cells mainly differentiated into motor and sensory neurons. These results suggested that the differentiation properties of iPS cells differ from those of ES cells.
Sakamoto, Tatsunori; Nakagawa, Takayuki; Endo, Tsuyoshi; Kim, Tae-Soo; Iguchi, Fukuichiro; Naito, Yasushi; Sasai, Yoshiki; Ito, Juichi
The potential of embryonic stem (ES) cells to differentiate into inner ear hair cells was examined in this study. Undifferentiated mouse ES cells transplanted into neomycin-damaged mouse inner ears were evaluated by immunohistochemistry 4 weeks after transplantation. Some ES cells were positive for E-cadherin or NCAM, and most transplanted cells were positive for SSEA3 and Ki67. None were positive for Myosin VIIa or MF20. These results indicate that the damaged inner ear may have some activity inducing ES cells to develop into ectoderm cells, but the effect was insufficient to induce inner ear hair cells. Next, SDIA/BMP-treated ES cells were transplanted into embryonic chicken inner ear rudiments. Embryonic chickens were expected to share the same developmental systems as mice. SDIA/BMP treatment drove ES cells to the population including neural crest cells and probably placode cells ES colonies were found next to or in the otic vesicles but were not a part of vesicle walls, indicating that transplanted ES cells could not be expected to be the same kind of cells as chicken otic vesicle cells Some ES colonies were found at the vestibulo-cochlear ganglions. To induce inner ear hair cells in this system, the competency of ES cells and otic induction signals should be defined further.
Boyan, George; Williams, Leslie; Legl, Andrea; Herbert, Zsofia
The central complex of the grasshopper Schistocerca gregaria develops to completion during embryogenesis. A major cellular contribution to the central complex is from the w, x, y, z lineages of the pars intercerebralis, each of which comprises over 100 cells, making them by far the largest in the embryonic protocerebrum. Our focus has been to find a cellular mechanism that allows such a large number of cell progeny to be generated within a restricted period of time. Immunohistochemical visualization of the chromosomes of mitotically active cells has revealed an almost identical linear array of proliferative cells present simultaneously in each w, x, y, z lineage at 50% of embryogenesis. This array is maintained relatively unchanged until almost 70% of embryogenesis, after which mitotic activity declines and then ceases. The array is absent from smaller lineages of the protocerebrum not associated with the central complex. The proliferative cells are located apically to the zone of ganglion mother cells and amongst the progeny of the neuroblast. Comparisons of cell morphology, immunoreactivity (horseradish peroxidase, repo, Prospero), location in lineages and spindle orientation have allowed us to distinguish the proliferative cells in an array from neuroblasts, ganglion mother cells, neuronal progeny and glia. Our data are consistent with the proliferative cells being secondary (amplifying) progenitors and originating from a specific subtype of ganglion mother cell. We propose a model of the way that neuroblasts, ganglion mother cells and secondary progenitors together produce the large cell numbers found in central complex lineages.
Young, R. S.
The relationship between the developing embryo (both plant and animal) and a gravitational field has long been contemplated. The difficulty in designing critical experiments on the surface of the earth because of its background of 1 g, has been an obstacle to a resolution of the problem. Biological responses to gravity (particularly in plants) are obvious in many cases; however, the influence of gravity as an environmental input to the developing embryo is not as obvious and has proven to be extremely difficult to define. In spite of this, over the years numerous attempts have been made using a variety of embryonic materials to come to grips with the role of gravity in development. Three research tools are available: the centrifuge, the clinostat, and the orbiting spacecraft. Experimental results are now available from all three sources. Some tenuous conclusions are drawn, and an attempt at a unifying theory of gravitational influence on embryonic development is made.
Young, R. S.
The relationship between the developing embryo (both plant and animal) and a gravitational field has long been contemplated. The difficulty in designing critical experiments on the surface of the earth because of its background of 1 g, has been an obstacle to a resolution of the problem. Biological responses to gravity (particularly in plants) are obvious in many cases; however, the influence of gravity as an environmental input to the developing embryo is not as obvious and has proven to be extremely difficult to define. In spite of this, over the years numerous attempts have been made using a variety of embryonic materials to come to grips with the role of gravity in development. Three research tools are available: the centrifuge, the clinostat, and the orbiting spacecraft. Experimental results are now available from all three sources. Some tenuous conclusions are drawn, and an attempt at a unifying theory of gravitational influence on embryonic development is made.
Prasad, Nikhil K; Desy, Nicholas M; Howe, B Matthew; Amrami, Kimberly K; Spinner, Robert J
Over the last decade, the mechanism of formation of intraneural ganglion cysts has been established through a meticulous review of clinical findings and correlation with patterns produced on magnetic resonance imaging (MRI). Pathognomonic imaging patterns distinguish these rare lesions from the more common extraneural variants in almost all cases. In this report, we present a new pattern of cyst occurrence in the subparaneurial compartment of the nerve and provide potential anatomic explanations for its pathogenesis. Using an anatomic framework of connective tissue compartments of the nerve, we reviewed 63 (56 fibular and seven tibial) intraneural ganglion cysts in the knee region evaluated at our institution and all reports with MRI in the world's literature for evidence of cyst occurrence in the subparaneurial compartment. We identified six cases (five in the common fibular nerve and one in the tibial nerve) at our institution that had MR evidence of cyst in the subparaneurial compartment with a new complex lobulated pattern. All cases had articular branch connections to the superior tibiofibular joint, which at operation were resected along with the joints. Follow-up revealed complete recovery in all instances and no clinical or radiological signs of recurrence. Three cases out of 80 in the literature exhibited the new complex lobulated MRI pattern. We present a new pattern of intraneural ganglion cyst occurrence in a potential space that surrounds peripheral nerves--the subparaneurial compartment. We believe that the unifying articular theory applies to the pathogenesis and management of these rare variants.
Endo, Jun; Yamaguchi, Satoshi; Sasho, Takahisa
Intratendinous ganglion cysts are rare lesions of unknown etiology that originate within a tendon. We report the case of a 34-year-old female with an intratendinous ganglion in the plantar portion of the flexor hallucis longus tendon. The intratendinous ganglion recurred after ultrasound-guided needle aspiration. Tendoscopic excision of the intratendinous ganglion cyst achieved a satisfactorily result without recurrence.
Mittal, Samarth; Singla, Amit; Nag, H L; Meena, Sanjay; Lohiya, Ramprakash; Agarwal, Abhinav
Intra-articular ganglion cysts of the knee joint are rare and most frequently are an incidental finding on MRI and arthroscopy. Most of the previous studies have reported a single ganglion cyst in the knee. There have been previous reports of more than one cyst in the same knee but not in the same structure within the knee. We are reporting a case of dual ACL (anterior cruciate ligament) ganglion cysts one of which was missed on radiological examination but later detected during arthroscopy. To the best of our knowledge, no such case has been reported in the indexed English literature till date.
Young, Allison; Petros, Timothy; Karayannis, Theofanis; McKenzie Chang, Melissa; Lavado, Alfonso; Iwano, Tomohiko; Nakajima, Miho; Taniguchi, Hiroki; Huang, Z. Josh; Heintz, Nathaniel; Oliver, Guillermo; Matsuzaki, Fumio; Machold, Robert P.
Neurogliaform (RELN+) and bipolar (VIP+) GABAergic interneurons of the mammalian cerebral cortex provide critical inhibition locally within the superficial layers. While these subtypes are known to originate from the embryonic caudal ganglionic eminence (CGE), the specific genetic programs that direct their positioning, maturation, and integration into the cortical network have not been elucidated. Here, we report that in mice expression of the transcription factor Prox1 is selectively maintained in postmitotic CGE-derived cortical interneuron precursors and that loss of Prox1 impairs the integration of these cells into superficial layers. Moreover, Prox1 differentially regulates the postnatal maturation of each specific subtype originating from the CGE (RELN, Calb2/VIP, and VIP). Interestingly, Prox1 promotes the maturation of CGE-derived interneuron subtypes through intrinsic differentiation programs that operate in tandem with extrinsically driven neuronal activity-dependent pathways. Thus Prox1 represents the first identified transcription factor specifically required for the embryonic and postnatal acquisition of CGE-derived cortical interneuron properties. SIGNIFICANCE STATEMENT Despite the recognition that 30% of GABAergic cortical interneurons originate from the caudal ganglionic eminence (CGE), to date, a specific transcriptional program that selectively regulates the development of these populations has not yet been identified. Moreover, while CGE-derived interneurons display unique patterns of tangential and radial migration and preferentially populate the superficial layers of the cortex, identification of a molecular program that controls these events is lacking. Here, we demonstrate that the homeodomain transcription factor Prox1 is expressed in postmitotic CGE-derived cortical interneuron precursors and is maintained into adulthood. We found that Prox1 function is differentially required during both embryonic and postnatal stages of development to
Maturana, Matias I; Turpin, Andrew; McKendrick, Allison M; Kameneva, Tatiana
This research takes a step towards discovering underlying ionic channel changes in the glaucomatous ganglion cells. Glaucoma is characterized by a gradual death of retinal ganglion cells. In this paper, we propose a hypothesis that the ionic channel concentrations change during the progression of glaucoma. We use computer simulation of a multi-compartment morphologically correct model of a mouse retinal ganglion cell to verify our hypothesis. Using published experimental data, we alter the morphology of healthy ganglion cells to replicate glaucomatous cells. Our results suggest that in glaucomatous cell, the sodium channel concentration decreases in the soma by 30% and by 60% in the dendrites, calcium channel concentration decreases by 10% in all compartments, and leak channel concentration increases by 40% in the soma and by 100% in the dendrites.
Dragoni, S; Giombini, A; Di Cesare, A; Ripani, M
The objective of the study is to describe two cases of proximal tibiofibular ganglion cysts in high level athletes. In May 2003 and March 2005 two athletes (one tennis player in the top eighty of the Italian national ranking and a gymnast belonging to the Italian rhythmic gymnastics national team) were referred to our institution complaining of postero-lateral knee discomfort and the presence of localized swelling over the fibular head and the antero-lateral aspect of the leg, with a clinically suspected diagnosis of ganglion cyst of the proximal tibiofibular joint. Ultrasonography clearly detected the fluid-filled structures while magnetic resonance imaging confirmed the diagnosis, also showing precisely the anatomic relationship between the ganglions and the surrounding structures. Both athletes underwent surgical excision and the histological examination was compatible with a proximal tibiofibular joint ganglion cyst; as yet they have had no recurrence.
Hameed, Shamsi Abdul; Sujir, Premjit; Naik, Monappa A; Rao, Sharath K
Ganglion cysts are more commonly associated with the anterior cruciate ligament than the posterior cruciate ligament (PCL). A literature review showed that all reported cases of ganglion cysts to date involved adults. We report a rare case of ganglion cyst in the PCL of a four-year-old boy, and discuss its aetiology, clinical presentation, imaging features and management. Ganglion cysts of the PCL may be confused with meniscal cysts arising from tears of the posterior horn of the medial meniscus on magnetic resonance (MR) imaging. Hence, the posterior horn of the medial meniscus has to be carefully evaluated to rule out a tear. MR imaging is the method of choice to confirm diagnosis, and arthroscopic resection is a safe treatment modality even in children.
Petros, Timothy J; Rebsam, Alexandra; Mason, Carol A
The retina and its sole output neuron, the retinal ganglion cell (RGC), comprise an excellent model in which to examine biological questions such as cell differentiation, axon guidance, retinotopic organization and synapse formation. One drawback is the inability to efficiently and reliably manipulate gene expression in RGCs in vivo, especially in the otherwise accessible murine visual pathways. Transgenic mice can be used to manipulate gene expression, but this approach is often expensive, time consuming, and can produce unwanted side effects. In chick, in ovo electroporation is used to manipulate gene expression in RGCs for examining retina and RGC development. Although similar electroporation techniques have been developed in neonatal mouse pups, adult rats, and embryonic murine retinae in vitro, none of these strategies allow full characterization of RGC development and axon projections in vivo. To this end, we have developed two applications of electroporation, one in utero and the other ex vivo, to specifically target embryonic murine RGCs[5, 6]. With in utero retinal electroporation, we can misexpress or downregulate specific genes in RGCs and follow their axon projections through the visual pathways in vivo, allowing examination of guidance decisions at intermediate targets, such as the optic chiasm, or at target regions, such as the lateral geniculate nucleus. Perturbing gene expression in a subset of RGCs in an otherwise wild-type background facilitates an understanding of gene function throughout the retinal pathway. Additionally, we have developed a companion technique for analyzing RGC axon growth in vitro. We electroporate embryonic heads ex vivo, collect and incubate the whole retina, then prepare explants from these retinae several days later. Retinal explants can be used in a variety of in vitro assays in order to examine the response of electroporated RGC axons to guidance cues or other factors. In sum, this set of techniques enhances
Takahashi, Masataka; Zhang, Zhen-Du; Macdonald, R Loch
Sphenopalatine ganglion stimulation activates perivascular vasodilatory nerves in the ipsilateral anterior circle of Willis. This experiment tested whether stimulation of the ganglion could reverse vasospasm and improve cerebral perfusion after subarachnoid hemorrhage (SAH) in monkeys. Thirteen cynomolgus monkeys underwent baseline angiography followed by creation of SAH by placement of autologous blood against the right intradural internal carotid artery, the middle cerebral artery (MCA), and the anterior cerebral artery. Seven days later, angiography was repeated, and the right sphenopalatine ganglion was exposed microsurgically. Angiography was repeated 15 minutes after exposure of the ganglion. The ganglion was stimulated electrically 3 times, and angiography was repeated during and 15 and 30 minutes after stimulation. Cerebral blood flow (CBF) was monitored using laser Doppler flowmetry, and intracranial pressure (ICP) was measured throughout. The protocol was repeated again. Evans blue was injected and the animals were killed. The brains were removed for analysis of water and Evans blue content and histology. Subarachnoid hemorrhage was associated with significant vasospasm of the ipsilateral major cerebral arteries (23% ± 10% to 39% ± 4%; p < 0.05, paired t-tests). Exposure of the ganglion and sham stimulation had no significant effects on arterial diameters, ICP, or CBF (4 monkeys, ANOVA and paired t-tests). Sphenopalatine ganglion stimulation dilated the ipsilateral extracranial and intracranial internal carotid artery, MCA, and anterior cerebral artery compared with the contralateral arteries (9 monkeys, 7% ± 9% to 15% ± 19%; p < 0.05, ANOVA). There was a significant increase in ipsilateral CBF. Stimulation had no effect on ICP or brain histology. Brain water content did not increase but Evans blue content was significantly elevated in the MCA territory of the stimulated hemisphere. Sphenopalatine ganglion stimulation decreased vasospasm and increased
Grierson, Claire; Nielsen, Erik; Ketelaarc, Tijs; Schiefelbein, John
Roots hairs are cylindrical extensions of root epidermal cells that are important for acquisition of nutrients, microbe interactions, and plant anchorage. The molecular mechanisms involved in the specification, differentiation, and physiology of root hairs in Arabidopsis are reviewed here. Root hair specification in Arabidopsis is determined by position-dependent signaling and molecular feedback loops causing differential accumulation of a WD-bHLH-Myb transcriptional complex. The initiation of root hairs is dependent on the RHD6 bHLH gene family and auxin to define the site of outgrowth. Root hair elongation relies on polarized cell expansion at the growing tip, which involves multiple integrated processes including cell secretion, endomembrane trafficking, cytoskeletal organization, and cell wall modifications. The study of root hair biology in Arabidopsis has provided a model cell type for insights into many aspects of plant development and cell biology. PMID:24982600
Gruhl, Alexander; Bartolomaeus, Thomas
In contrast to other Bryozoa, members of the subtaxon Phylactolaemata bear a subepithelial cerebral ganglion that resembles a hollow vesicle rather than being compact. In older studies this ganglion was said to originate by an invagination of the pharyngeal epithelium. Unfortunately, documentation for this is fragmentary. In chordates the central nervous system also arises by an invagination-like process, but this mode is uncommon among invertebrate phyla. As a first attempt to gather more data about this phenomenon, cerebral ganglia in two phylactolaemate species, Fredericella sultana and Plumatella emarginata, were examined at the ultrastructural level. In both species the ganglion bears a small central lumen. The ganglionic cells are organized in the form of a neuroepithelium. They are polarized and interconnected by adherens junctions on their apical sides and reside on a basal lamina. The nerve cell somata are directed towards the central lumen, whereas the majority of nervous processes are distributed basally. Orientation of the neuroepithelial cells can be best explained by the possibility that they develop by invagination. A comparison with potential outgroups reveals that a neuroepithelial ganglion is at least derived. Since, however, a reliable phylogenetic system of the Bryozoa is missing, a decision on whether such a ganglion is apomorphic for Bryozoa or evolved within this taxon can hardly be made. (c) 2007 Wiley-Liss, Inc.
Jiang, Zheng; Bulley, Simon; Guzzone, Joseph; Ripps, Harris; Shen, Wen
Taurine (2-aminoethylsuphonic acid) is present in nearly all animal tissues, and is the most abundant free amino acid in muscle, heart, CNS and retina. Although it is known to be a major cytoprotectant and essential for normal retinal development, its role in retinal neurotransmission and modulation is not well understood. We investigated the response of taurine in retinal ganglion cells, and its effect on synaptic transmission between ganglion cells and their pre-synaptic neurons. We find that taurine-elicited currents in ganglion cells could be fully blocked by both strychnine and SR95531, glycine and GABAA receptor antagonists, respectively. This suggests that taurine-activated receptors might share the antagonists with GABA and glycine receptors. The effect of taurine at micromolar concentrations can effectively suppress spontaneous vesicle release from the pre-synaptic neurons, but had limited effects on light-evoked synaptic signals in ganglion cells. We also describe a metabotropic effect of taurine in the suppression of light-evoked response in ganglion cells. Clearly, taurine acts in multiple ways to modulate synaptic signals in retinal output neurons, ganglion cells. PMID:23392924
Lee, Sherwin C.; Ishida, Andrew T.
Antisera directed against hyperpolarization-activated mixed-cation (“Ih”) and K+ (“Kir”) channels bind to some somata in the ganglion cell layer of rat and rabbit retina. Additionally, the termination of hyperpolarizing current injections can trigger spikes in some cat retinal ganglion cells, suggesting a rebound depolarization due to activation of Ih. However, patch-clamp studies have reported that rat ganglion cells lack inward rectification, or present an inwardly rectifying K+ current. We therefore tested whether hyperpolarization activates Ih in dissociated, adult rat retinal ganglion cell somata. We report here that while we found no inward rectification in some cells, and a Kir-like current in a few cells, hyperpolarization activated Ih in roughly 75% of the cells we recorded from in voltage clamp. We show that this current is blocked by Cs+ or ZD7288 and only slightly reduced by Ba2+, that the current amplitude and reversal potential are sensitive to extracellular Na+ and K+, and that we found no evidence of Kir in cells presenting Ih. In current clamp, injecting hyperpolarizing current induced a slowly relaxing membrane hyperpolarization that rebounded to a few action potentials when the hyperpolarizing current was stopped; both the membrane potential relaxation and rebound spikes were blocked by ZD7288. These results provide the first measurement of Ih in mammalian retinal ganglion cells, and indicate that the ion channels of rat retinal ganglion cells may vary in ways not expected from previous voltage and current recordings. PMID:17488978
Segev, Ronen; Puchalla, Jason; Berry, Michael J
Recently, we reported a novel technique for recording all of the ganglion cells in a retinal patch and showed that their receptive fields cover visual space roughly 60 times over in the tiger salamander. Here, we carry this analysis further and divide the population of ganglion cells into functional classes using quantitative clustering algorithms that combine several response characteristics. Using only the receptive field to classify ganglion cells revealed six cell types, in agreement with anatomical studies. Adding other response measures served to blur the distinctions between these cell types rather than resolve further classes. Only the biphasic off type had receptive fields that tiled the retina. Even when we attempted to split these classes more finely, ganglion cells with almost identical functional properties were found to have strongly overlapping spatial receptive fields. A territorial spatial organization, where ganglion cell receptive fields tend to avoid those of other cells of the same type, was only found for the biphasic off cell. We further studied the functional segregation of the ganglion cell population by computing the amount of visual information shared between pairs of cells under natural movie stimulation. This analysis revealed an extensive mixing of visual information among cells of different functional type. Together, our results indicate that the salamander retina uses a population code in which every point in visual space is represented by multiple neurons with subtly different visual sensitivities.
Abbas, Syed Y; Hamade, Khaldoun C; Yang, Ellen J; Nawy, Scott; Smith, Robert G; Pettit, Diana L
Retinal ganglion cells receive inputs from multiple bipolar cells which must be integrated before a decision to fire is made. Theoretical studies have provided clues about how this integration is accomplished but have not directly determined the rules regulating summation of closely timed inputs along single or multiple dendrites. Here we have examined dendritic summation of multiple inputs along On ganglion cell dendrites in whole mount rat retina. We activated inputs at targeted locations by uncaging glutamate sequentially to generate apparent motion along On ganglion cell dendrites in whole mount retina. Summation was directional and dependent13 on input sequence. Input moving away from the soma (centrifugal) resulted in supralinear summation, while activation sequences moving toward the soma (centripetal) were linear. Enhanced summation for centrifugal activation was robust as it was also observed in cultured retinal ganglion cells. This directional summation was dependent on hyperpolarization activated cyclic nucleotide-gated (HCN) channels as blockade with ZD7288 eliminated directionality. A computational model confirms that activation of HCN channels can override a preference for centripetal summation expected from cell anatomy. This type of direction selectivity could play a role in coding movement similar to the axial selectivity seen in locust ganglion cells which detect looming stimuli. More generally, these results suggest that non-directional retinal ganglion cells can discriminate between input sequences independent of the retina network.
Abbas, Syed Y.; Hamade, Khaldoun C.; Yang, Ellen J.; Nawy, Scott; Smith, Robert G.; Pettit, Diana L.
Retinal ganglion cells receive inputs from multiple bipolar cells which must be integrated before a decision to fire is made. Theoretical studies have provided clues about how this integration is accomplished but have not directly determined the rules regulating summation of closely timed inputs along single or multiple dendrites. Here we have examined dendritic summation of multiple inputs along On ganglion cell dendrites in whole mount rat retina. We activated inputs at targeted locations by uncaging glutamate sequentially to generate apparent motion along On ganglion cell dendrites in whole mount retina. Summation was directional and dependent13 on input sequence. Input moving away from the soma (centrifugal) resulted in supralinear summation, while activation sequences moving toward the soma (centripetal) were linear. Enhanced summation for centrifugal activation was robust as it was also observed in cultured retinal ganglion cells. This directional summation was dependent on hyperpolarization activated cyclic nucleotide-gated (HCN) channels as blockade with ZD7288 eliminated directionality. A computational model confirms that activation of HCN channels can override a preference for centripetal summation expected from cell anatomy. This type of direction selectivity could play a role in coding movement similar to the axial selectivity seen in locust ganglion cells which detect looming stimuli. More generally, these results suggest that non-directional retinal ganglion cells can discriminate between input sequences independent of the retina network. PMID:23516351
Haller, Justin M.; Potter, Michael Q.; Sinclair, Micah; Hutchinson, Douglas T.
Background Intraneural ganglions in peripheral nerves of the upper extremity are extremely rare and poorly understood. Case Description We report a patient with symptoms consistent with de Quervain tenosynovitis who was found to have an intraneural ganglion in the superficial radial nerve. The ganglion did not communicate with the wrist joint. We removed the intraneural ganglion, and the patient's symptoms resolved. At her 6-month postoperative follow-up, she remained asymptomatic. Literature Review: There is only one case report of intraneural ganglion in the superficial radial nerve. In that case, the patient had symptoms consistent with nerve irritation, including radiating pain and paresthesias. In contrast to that previous report, the patient in the current case had only localized pain, no paresthesias, and a physical exam consistent with de Quervain tenosynovitis. Clinical Relevance This case demonstrates that an intraneural ganglion cyst can mimic the symptoms of de Quervain tenosynovitis without the more usual presentation of painful paresthesias. PMID:25364639
Walsh, C.; Polley, E.H.
The ganglion cells of the cat's retina form several classes distinguishable in terms of soma size, axon diameter, dendritic morphology, physiological properties, and central connections. Labeling with (/sup 3/H)thymidine shows that the ganglion cells which survive in the adult are produced as several temporally shifted, overlapping waves: medium-sized cells are produced before large cells, whereas the smallest ganglion cells are produced throughout the period of ganglion cell generation. Large cells and medium-sized cells show the same distinctive pattern of production, forming rough spirals around the area centralis. The oldest cells tend to lie superior and nasal to the area centralis, whereas cells in the inferior nasal retina and inferior temporal retina are, in general, progressively younger. Within each retinal quadrant, cells nearer the area centralis tend to be older than cells in the periphery, but there is substantial overlap. The retinal raphe divides the superior temporal quadrant into two zones with different patterns of cell addition. Superior temporal retina near the vertical meridian adds cells only slightly later than superior nasal retina, whereas superior temporal retina near the horizontal meridian adds cells very late, contemporaneously with inferior temporal retina. The broader wave of production of smaller ganglion cells seems to follow this same spiral pattern at its beginning and end. The presence of the area centralis as a nodal point about which ganglion cell production in the retinal quadrants pivots suggests that the area centralis is already an important retinal landmark even at the earliest stages of retinal development.
Paudel, Sharada; Kim, Yeoun-Hee; Huh, Man-Il; Kim, Song-Ja; Chang, Yongmin; Park, Young Jeung; Lee, Kyoo Won; Jung, Jae-Chang
Here, we examined the role of ADAM10 during retinal cell differentiation in retinal sections and in vitro cultures of developing chick retinal cells from embryonic day 6 (ED6). Immunohistochemistry showed that ADAM10 is abundantly expressed in the inner zone of neuroblastic layer at ED5, and it becomes more highly expressed in the ganglion cell layer at ED7 and ED9. Western blotting confirmed that ADAM10 was expressed as an inactive pro-form that was processed to a shorter, active form in control cultured cells, but in cultures treated with an ADAM10 inhibitor (GI254023X) and ADAM10-specific siRNA, the level of mature ADAM10 decreased. Phase-contrast microscopy showed that long neurite extensions were present in untreated cultures 24 h after plating, whereas cultures treated with GI254023X showed significant decreases in neurite extension. Immunofluorescence staining revealed that there were far fewer differentiated ganglion cells in ADAM10 siRNA and GI254023X-treated cultures compared to controls, whereas the photoreceptor cells were unaltered. The Pax6 protein was more strongly detected in the differentiated ganglion cells of control cultures compared to ADAM10 siRNA and GI254023X-treated cultures. N-cadherin ectodomain shedding was apparent in control cultures after 24 h, when ganglion cell differentiation was observed, but ADAM10 siRNA and GI254023X treatment inhibited these processes. In contrast, N-cadherin staining was strongly detected in photoreceptor cells regardless of ADAM10 siRNA and GI254023X treatment. Taken together, these data indicate that the inhibition of ADAM10 can inhibit Pax6 expression and N-cadherin ectodomain shedding in retinal cells, possibly affecting neurite outgrowth and ganglion cell differentiation.
Pfaff, F; Schlottau, K; Scholes, S; Courtenay, A; Hoffmann, B; Höper, D; Beer, M
In June 2013, a 4-year-old Welsh Mountain ewe and in March 2014 a 10-day-old lamb of the same breed and the same flock presented progressive neurological signs including depressed sensorium, tremor, and unusual behaviour. Neuropathological examination of the brain and spinal cord detected non-suppurative polioencephalomyelitis and dorsal root ganglionitis, characteristic of a neurotropic viral agent in both sheep. Metagenomic analysis of different tissue samples from both animals identified a novel Ovine Astrovirus (OvAstV). The presence of viral genome in the central nervous system was confirmed by RT-qPCR. Although the cases presented nine months apart, the identified OvAstV shared nearly identical sequences, differing in only three nucleotide positions across the complete genome. Phylogenetic analysis revealed a close relation of OvAstV to neurotropic bovine astroviruses and an enteric OvAstV. In conclusion, these are the first reported cases of astrovirus infection in domestic sheep that were associated with encephalitis and ganglionitis.
Mead, Ben; Tomarev, Stanislav
Retinal ganglion cells (RGC) bear the sole responsibility of propagating visual stimuli to the brain. Their axons, which make up the optic nerve, project from the retina to the brain through the lamina cribrosa and in rodents, decussate almost entirely at the optic chiasm before synapsing at the superior colliculus. For many traumatic and degenerative ocular conditions, the dysfunction and/or loss of RGC is the primary determinant of visual loss and are the measurable endpoints in current research into experimental therapies. To actually measure these endpoints in rodent models, techniques must ascertain both the quantity of surviving RGC and their functional capacity. Quantification techniques include phenotypic markers of RGC, retrogradely transported fluorophores and morphological measurements of retinal thickness whereas functional assessments include electroretinography (flash and pattern) and visual evoked potential. The importance of the accuracy and reliability of these techniques cannot be understated, nor can the relationship between RGC death and dysfunction. The existence of up to 30 types of RGC complicates the measuring process, particularly as these may respond differently to disease and treatment. Since the above techniques may selectively identify and ignore particular subpopulations, their appropriateness as measures of RGC survival and function may be further limited. This review discusses the above techniques in the context of their subtype specificity.
The function of retinal ganglion cells (RGCs) can be non-invasively assessed in experimental and genetic models of glaucoma by means of variants of the ERG technique that emphasize the activity of inner retina neurons. The best understood technique is the Pattern Electroretinogram (PERG) in response to contrast-reversing gratings or checkerboards, which selectively depends on the presence of functional RGCs. In glaucoma models, the PERG can be altered before histological loss of RGCs; PERG alterations may be either reversed with moderate IOP lowering or exacerbated with moderate IOP elevation. Under particular luminance-stimulus conditions, the Flash-ERG displays components that may reflect electrical activity originating in the proximal retina and be altered in some experimental glaucoma models (positive Scotopic Threshold response, pSTR; negative Scotopic Threshold Response, nSTR; Photopic Negative Response, PhNR; Oscillatory Potentials, OPs; multifocal ERG, mfERG). It is not yet known which of these components is most sensitive to glaucomatous damage. Electrophysiological assessment of RGC function appears to be a necessary outcome measure in experimental glaucoma models, which complements structural assessment and may even predict it. Neuroprotective strategies could be tested based on enhancement of baseline electrophysiological function that results in improved RGC survival. The use of electrophysiology in glaucoma models may be facilitated by specifically designed instruments that allow high throughput, robust assessment of electrophysiological function. PMID:25998495