BACKGROUND: Glucocorticoids have anti-inflammatory effects and have been used to treat many types of nerve injury- associated chronic pain conditions. A randomized double-blind study was performed to determine if methylprednisolone could prevent the development of neuropathic pain after a peripheral nerve injury in rats. METHODS: Two groups of rats, one group (n = 50) injected intraperitoneally with methylprednisolone (100 mg/kg/day, for 7 days starting from 3 days prior to the nerve injury) and the other (n = 58) treated with saline with same manner, were compared in terms of the incidence and intensity of allodynia after a superior caudal trunk transection at the level between the 3rd and 4th sacral spinal nerves. The tail-flick responses to normally innocuous mechanical and thermal stimuli applied to the tail were observed as the behavioral signs of neuropathic pain. RESULTS: The proportions of rats exhibiting tail-flick responses to the mechanical (but not thermal) stimuli 7, 14 and 21 days after the nerve injury were significantly smaller in the methylprednisolone-treated group (2, 3 and 4 of 50 rats, respectively) than in the saline-treated, control group (11, 14 and 15 of 58 rats, respectively) (P = 0.009). However, the pain intensity was similar in mechanical allodynia developed rats of the two groups (P > 0.05), which was estimated based on the frequency and latency of the tail-flick responses after applying mechanical and thermal stimuli, respectively. CONCLUSIONS: These results suggest that a pre-emptive treatment with high methylprednisolone doses may be used to prevent the development of mechanical allodynia following peripheral nerve injuries.
Animals ; Axotomy ; Chronic Pain ; Double-Blind Method ; Glucocorticoids ; Hyperalgesia* ; Incidence ; Methylprednisolone* ; Neuralgia ; Peripheral Nerve Injuries* ; Peripheral Nerves* ; Rats* ; Spinal Nerves

Traumatic injury or inflammatory irritation of the peripheral nervous system often leads to persistent pathophysiological pain states. It has been well-documented that, after peripheral nerve injury or inflammation, functional and anatomical alterations sweep over the entire peripheral nervous system including the peripheral nerve endings, the injured or inflamed afferent fibers, the dorsal root ganglion (DRG), and the central afferent terminals in the spinal cord. Among all the changes, ectopic discharge or spontaneous activity of primary sensory neurons is of great clinical interest, as such discharges doubtless contribute to the development of pathological pain states such as neuropathic pain. Two key sources of abnormal spontaneous activity have been identified following peripheral nerve injury: the injured afferent fibers (neuroma) leading to the DRG, and the DRG somata. The purpose of this review is to provide a global account of the abnormal spontaneous activity in various animal models of pain. Particular attention is focused on the consequence of peripheral nerve injury and localized inflammation. Further, mechanisms involved in the generation of spontaneous activity are also reviewed; evidence of spontaneous activity in contributing to abnormal sympathetic sprouting in the axotomized DRG and to the initiation of neuropathic pain based on new findings from our research group are discussed. An improved understanding of the causes of spontaneous activity and the origins of neuropathic pain should facilitate the development of novel strategies for effective treatment of pathological pain.
Animals ; Axotomy ; Ganglia, Spinal ; cytology ; Humans ; Neuralgia ; physiopathology ; Neurons, Afferent ; cytology ; Peripheral Nerve Injuries ; physiopathology ; Spinal Cord ; cytology

In the retina, dopaminergic cells express the receptor for brain-derived neurotrophic factor (BDNF), which is known to be retrogradely transported from higher center to the retina. This study was conducted to identify the effect of optic nerve transaction on the dopaminergic cells in the rat retina by immunocytochemistry using antityrosine hydroxylase (TH) antiserum. In the control retina, we found two types of TH-immunoreactive amacrine cells, type I and type II, in the inner nuclear layer (INL) adjacent to the inner plexiform layer (IPL). The type I amacrine cell varicosities formed ring-like structures in contact with AII amacrine cell somata in stratum 1 of the IPL. In the axotomized retinas, TH-labeled processes formed loose networks of fibers, unlike the dense networks in the control retina, and the ring-like structures were disrupted. Our data suggest that retrogradely transported neurotrophic factor affects the expression of TH immunoreactivity in the axotomized rat retina and may therefore influence the retinal dopaminergic system.
Amacrine Cells ; Animals ; Axotomy* ; Brain-Derived Neurotrophic Factor ; Immunohistochemistry ; Optic Nerve ; Rats* ; Retina* ; Retinaldehyde ; Tyrosine 3-Monooxygenase

Changes in morphology, immunophenotypes and proliferative activity of neuroglia are key features in most forms of CNS pathology. We compared proliferative activity of neuroglial cells in response to two different types of brain injury induced by medial forebrain bundle (MFB) axotomy. In the cannula track where acute necrosis occurs due to mechanical lesion caused by cannula inserted to incise the MFB, many BrdU-immunoreactive (ir) cells appeared around the cannula track already at 1 day post-lesion (1 dpl). Their number significantly increased by 7 dpl and then decreased, but considerable number of BrdU-ir cells was still found at 14 dpl. Some of the BrdU-ir cells were double-labeled with either OX-42 or GFAP. This finding suggests that both microglia and astrocytes are activated and proliferate immediately after the mechanical damage, and the proliferative activity is maintained in a considerable number of these cells by 14 dpl. In general, the main cell type showing BrdU immunoreactivity was amoeboid microglia within the necrotic zone immediately surrounding the cannula track, and was astrocytes in the periphery of the necrotic zone more or less apart from the cannula track. Previously, we reported that MFB axotomy induces apoptosis of dopaminergic (DA) neurons in the substantia nigra (SN). In the SN where axotomized DA neurons undergo apoptosis, only a few BrdU-ir cells were found at 1 dpl. Their number increased gradually from 3 dpl and peaked at 7 dpl, then significantly reduced at 14 dpl. Most of them were double-labeled with OX -42-positive ramified microglia but not with GFAP. This data indicates that microglia but not astrocyte are the cell type that proliferate in response to apoptotic neuronal cell death, and their morphology and proliferative activity are different from those observed in the cannula track. Meanwhile, in the both cannula track and SN, some BrdU-ir cells were thought to be neither GFAP-positive nor OX-42-positive, and thus they were presumed to be infiltrated peripheral immune cells. These results demonstrate that different types of neuronal cell death are accompanied with different neurogilal proliferative activities.
Apoptosis ; Astrocytes ; Axotomy* ; Brain Injuries ; Bromodeoxyuridine ; Catheters ; Cell Death ; Medial Forebrain Bundle* ; Microglia ; Necrosis ; Neuroglia ; Neurons ; Pathology ; Substantia Nigra

Activating transcription factor 3 (ATF3) and c-Jun play key roles in either cell death or cell survival, depending on the cellular background. To evaluate the functional significance of ATF3/c-Jun in the peripheral nervous system, we examined neuronal cell death, activation of ATF3/c-Jun, and microglial responses in facial motor nuclei up to 24 weeks after an extracranial facial nerve axotomy in adult rats. Following the axotomy, neuronal survival rate was progressively but significantly reduced to 79.1% at 16 weeks post-lesion (wpl) and to 65.2% at 24 wpl. ATF3 and phosphorylated c-Jun (pc-Jun) were detected in the majority of ipsilateral facial motoneurons with normal size and morphology during the early stage of degeneration (1-2 wpl). Thereafter, the number of facial motoneurons decreased gradually, and both ATF3 and pc-Jun were identified in degenerating neurons only. ATF3 and pc-Jun were co-localized in most cases. Additionally, a large number of activated microglia, recognized by OX6 (rat MHC II marker) and ED1 (phagocytic marker), gathered in the ipsilateral facial motor nuclei. Importantly, numerous OX6- and ED1-positive, phagocytic microglia closely surrounded and ingested pc-Jun-positive, degenerating neurons. Taken together, our results indicate that long-lasting co-localization of ATF3 and pc-Jun in axotomized facial motoneurons may be related to degenerative cascades provoked by an extracranial facial nerve axotomy.
Activating Transcription Factor 3 ; Adult ; Animals ; Axotomy ; Cell Death ; Cell Survival ; Facial Nerve ; Humans ; Microglia ; Neurons ; Peripheral Nervous System ; Rats ; Survival Rate

The aim of this study was to propose new more reliable peripheral nerve transection model to overcome the defect of the traditional sciatic axotomy model by specifically transecting L5 spinal nerve just after emerging from the intervertebral foramen and confining analysis area to the L5 spinal segment. The adult male Sprague-Dawley rats, weighing 300~350 g at the time of surgery, were used for the experiments. Four different experimental groups were used. 1. Sciatic nerve transection (Sc-Tx) group: transect the sciatic nerve in the popliteal fossa where it divided into the common peroneal nerve and tibial nerve. 2. L5 spinal nerve transection (L5-Tx) group: L5 spinal nerve was specifically transected. 3. Suture (Su) group: L5 spinal nerve was transected and immediately sutured. 4. Control group: the same surgical procedure with L5 spinal nerve transection group was performed except for the excision of L5 spinal nerve. To distinguish L5 motoneurons from the other level ones, the animals were received the retrograde tracer, FluoroGold into the axotomized proximal nerve stump. Serial coronal frozen sections at 40 microm thick through the L4 to L6 spinal segment was performed and the resultant total number of sections was about 180. Approximate serial 50 sections (approximately 2 mm) could be considered as the L5 segment based on the number of the fluorescent signals (above 20). L5 spinal segment could be differentiated from L4 and L6 segment based on their morphological characteristics under Cresyl violet stain. In L5-Tx group, at 2 and 4 weeks post-transection, the number of L5 spinal motoneurons was reduced by 8%. Meanwhile, Sc-Tx and Su groups showed no statistically notable changes. In this study, the authors could propose more reliable peripheral nerve axotomy model than the conventional sciatic nerve axotomy model by specifically transecting L5 spinal nerve and confining the investigating area within the L5 spinal segment.
Adult ; Animals ; Axotomy ; Benzoxazines ; Frozen Sections ; Humans ; Male ; Peripheral Nerve Injuries ; Peripheral Nerves ; Peroneal Nerve ; Rats ; Rats, Sprague-Dawley ; Sciatic Nerve ; Spinal Nerves ; Sutures ; Tibial Nerve ; Viola

The aim of this study was to propose new more reliable peripheral nerve transection model to overcome the defect of the traditional sciatic axotomy model by specifically transecting L5 spinal nerve just after emerging from the intervertebral foramen and confining analysis area to the L5 spinal segment. The adult male Sprague-Dawley rats, weighing 300~350 g at the time of surgery, were used for the experiments. Four different experimental groups were used. 1. Sciatic nerve transection (Sc-Tx) group: transect the sciatic nerve in the popliteal fossa where it divided into the common peroneal nerve and tibial nerve. 2. L5 spinal nerve transection (L5-Tx) group: L5 spinal nerve was specifically transected. 3. Suture (Su) group: L5 spinal nerve was transected and immediately sutured. 4. Control group: the same surgical procedure with L5 spinal nerve transection group was performed except for the excision of L5 spinal nerve. To distinguish L5 motoneurons from the other level ones, the animals were received the retrograde tracer, FluoroGold into the axotomized proximal nerve stump. Serial coronal frozen sections at 40 microm thick through the L4 to L6 spinal segment was performed and the resultant total number of sections was about 180. Approximate serial 50 sections (approximately 2 mm) could be considered as the L5 segment based on the number of the fluorescent signals (above 20). L5 spinal segment could be differentiated from L4 and L6 segment based on their morphological characteristics under Cresyl violet stain. In L5-Tx group, at 2 and 4 weeks post-transection, the number of L5 spinal motoneurons was reduced by 8%. Meanwhile, Sc-Tx and Su groups showed no statistically notable changes. In this study, the authors could propose more reliable peripheral nerve axotomy model than the conventional sciatic nerve axotomy model by specifically transecting L5 spinal nerve and confining the investigating area within the L5 spinal segment.
Adult ; Animals ; Axotomy ; Benzoxazines ; Frozen Sections ; Humans ; Male ; Peripheral Nerve Injuries ; Peripheral Nerves ; Peroneal Nerve ; Rats ; Rats, Sprague-Dawley ; Sciatic Nerve ; Spinal Nerves ; Sutures ; Tibial Nerve ; Viola

This investigation were performed in order to study the mechanism of neuronal cell degeneration and mediated gene of retinal cell death in optic nerve transection model. The optic nerve was transected at 5mm posterior from eye ball without affecting the retinal blood supply. Expression of cell deth gene were observed by p53p and Bax immunohistochemistry. The density of ganglion cels was determined by flat-mounted retinas after axotomy. Dying cells contained the pycnotic nuclei at 1 and 2 weeks and approximately 50% of ganglion cells were degenerated within 14 days after axotomy in ganglion cell layer. Bax and p53 were found to be expressed in the ganglion cell layer and inner unclear layer at 7 and 14 days after optic nerve transection. At 14 days, expression of death genes were observed strogly in ganglion cell layer and partially in inner unclear layer. this study suggest that p53p and Bax proteins may mediate ganglion cell degeneration after axotomy.
Axotomy* ; bcl-2-Associated X Protein* ; Cell Death* ; Ganglion Cysts ; Immunohistochemistry ; Neurons ; Optic Nerve Injuries ; Optic Nerve* ; Retina ; Retinal Neurons* ; Retinaldehyde*

Objective A calcium-activated chloride current (IClCa) has been observed in medium-sized sensory neurons of the dorsal root ganglion (DRG). Axotomy of the sciatic nerve induces a similar current in the majority of medium and large diameter neurons. Our aim is to identify the molecule(s) underlying this current. Methods Using conventional and quantitative RT-PCR, we examined the expression in DRG of members of three families of genes, which have been shown to have IClCa current inducing properties. Results We showed the detection of transcripts representing several members of these families, i.e. chloride channel calcium-activated (CLCA), Bestrophin and Tweety gene families in adult DRG, in the normal state and 3 d after sciatic nerve section, a model for peripheral nerve injury. Conclusion Our analysis revealed that that mBest1 and Tweety2 appear as the best candidates to play a role in the injury-induced IClCa in DRG neurons.
Animals ; Axotomy ; Chloride Channels ; biosynthesis ; genetics ; DNA Primers ; Ganglia, Spinal ; metabolism ; Gene Expression ; Mice ; Neurons, Afferent ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Sciatic Nerve ; physiology

Medial forebrain bundle (MFB) transmits the nigrostriatal dopaminergic (DA) axons, and previously we reported that transection of the MFB causes apotosis-like neurodegeneration of nigral DA neurons. On the other hand, it is likely to occur necrosis at the lesioned site where MFB is cut, due to direct mechanical transection of the brain tissue. To clarify the pathological dynamics of microglia reacting to the two different types of neuronal cell death, immunophenotypic and morphological features of microglia were compared and analyzed in the substantia nigra (SN) and lesioned site of the MFB axotomized rat brain. OX42 (mouse anti-rat CD 11b; pan-microglia marker), ED1 (mouse anti-rat lysosomal enzyme; phagocytic marker), and OX6 (mouse anti-rat MHC II) were used as primary antibodies for immunohistochemical localization of microglia, ED2 (mouse anti-rat macrophage) for macrophages, and anti-tyrosine hydro-xylase (TH) antibody for DA neurons. Quite numerous activated microglia with strong OX42 immunoreactivity were found in the SN at 1 day post-lesion (dpl), but most of them were ED1-and OX6-negative except only a few which were ED1-positive. This phenomenon was thought to be related with the stage of alert, the first step of microglial activation. It could be presumed that microglial phagocytosis may precede MHC II expression, because ED1-positive microglia appeared from 1 dpl while OX6-positive ones from 3 dpl. Number of activated microglia showing strong ED1, OX6 and OX42 immunoreactivity increased significantly by 7 ~14 dpl, and they specifically stick to various parts of dendrites and somas of TH-immunoreactive neurons of the SN. The phagocytic microglia of the SN maintained ramified form although they retained enlarged soma and shortened, thickened processes. The lesioned site was surrounded by numerous microglia showing strong OX42 and ED1 immunoreactivity as early as 1 dpl, indicating that microglial phagocytosis starts earlier in the lesioned site than in the SN. OX42-positive microglia of the lesioned site were ED2-negative, and showed amoeboid morphology already from 1 dpl. The amoeboid microglia became to be enlarged in their soma size by 3 dpl, and fused each other to form clumps within the necrotic zone by 5 ~7 dpl. The entire necrotic zone was completely filled with microglia of obscure outline with strong OX42 and ED1 immuno-reactivity. However, the majority of amoeboid microglia of the lesioned site were OX6-negative except a few. These results clearly demonstrate that activated microglia reacting to apoptotic neurodegeneration show different pathodynamic characteristics in terms of immunological phenotypes and morphology from those reacting to necrotic, mechanical lesion.
Animals ; Antibodies ; Apoptosis ; Axons ; Axotomy ; Brain ; Carisoprodol ; Cell Death ; Dendrites ; Hand ; Macrophages ; Medial Forebrain Bundle* ; Microglia* ; Necrosis ; Neurons ; Phagocytosis ; Phenotype ; Rats ; Substantia Nigra*