Normal Rat Spinal Cord Research Articles

Adhesion and proliferation of bone marrow stromal cells on acellular spinal cord scaffolds

Objectives This study aimed to produce an acellular spinal cord scaffold-bone marrow stromal cell (ASCS-BMSC) complex in which the growth of BMSCs transplanted into the spinal cord of rats could be simulated in vitro, facilitating the observation and evaluation of the growth of BMSCs on the ASCS for the first time. Methods Freeze-thaw, chemical extraction and mechanical shaking approaches were used to remove the cellular components and prepare a rat ASCS containing only the extracellular matrix (ECM) structure from the rat spinal cord. BMSCs were embedded into ASCSs and freeze-dried agarose scaffolds (FASs), and cell migration and proliferation were observed via fluorescence microscopy and the MTT assay. Results Compared with the normal rat spinal cord, the ASCS had no cell structure and retained ECM components such as type IV collagen, fibronectin and laminin, showing a three-dimensional network structure with good voids. The growth and proliferation of BMSCs on the ASCS was good, as shown by the MTT assay. Scanning electron microscopy showed that BMSCs covered 65% of the ASCS surface, and the mitochondria of BMSCs were developed and adhered to collagen fibres, as demonstrated by transmission electron microscopy. HE staining showed that BMSCs could grow inside the ASCS, and immunohistochemical staining showed that BMSCs still expressed CD44 and CD90 on the ASCS and had stem cell characteristics. Conclusions The results of the experiment indicate that the ASCS has the ability to improve cell adhesion and proliferation. Thus, the ASCS-BMSC combination may be used to treat spinal cord injury.

3D Digital Anatomic Angioarchitecture of the Rat Spinal Cord: A Synchrotron Radiation Micro-CT Study.

Comprehensive analysis of 3D angioarchitecture within the intact rat spinal cord remains technically challenging due to its sophisticated anatomical properties. In this study, we aim to present a framework for ultrahigh-resolution digitalized mapping of the normal rat spinal cord angioarchitecture and to determine the physiological parameters using synchrotron radiation micro-CT (SRμCT). Male SD rats were used in this ex vivo study. After a proportional mixture of contrast agents perfusion, the intact spinal cord covered the cervical spinal from the upper of the 1st cervical vertebra to the 5th lumbar vertebra was harvested and cut into proper lengths within three distinct regions: Cervical 3–5 levels, Thoracic 10–12 levels, Lumbar 3–5 levels spinal cord and examined using SRμCT. This method enabled the replication of the complicated microvasculature network of the normal rat spinal cord at the ultrahigh-resolution level, allowing for the precise quantitative analysis of the vascular morphological difference among cervical, thoracic and lumbar spinal cord in a 3D manner. Apart from a series of delicate 3D digital anatomical maps of the rat spinal cord angioarchitecture ranging from the cervical and thoracic to the lumbar spinal cord were presented, the 3D reconstruction data of SRμCT made the 3D printing of the spinal cord targeted selected microvasculature reality, that possibly provided deep insight into the nature and role of spinal cord intricate angioarchitecture. Our data proposed a new approach to outline systematic visual and quantitative evaluations on the 3D arrangement of the entire hierarchical microvasculature of the normal rat spinal cord at ultrahigh resolution. The technique may have great potential and become useful for future research on the poorly understood nature and function of the neurovascular interaction, particularly to investigate their pathology changes in various models of neurovascular disease.

Expression and Cellular Localization of IFITM1 in Normal and Injured Rat Spinal Cords.

Interferon-induced transmembrane protein 1 (IFITM1) is a member of the IFITM family that is associated with some acute-phase cytokine-stimulated response. Recently, we demonstrated that IFITM1 was significantly upregulated in the injured spinal cords at the mRNA level. However, its expression and cellular localization at the protein level is still unclear. Here, a rat model of spinal cord injury (SCI) was performed to investigate the spatio-temporal expression of IFITM1 after SCI. IFITM1 mRNA and protein were assessed by quantitative reverse transcription-PCR and western blot, respectively. IHC was used to identify its cellular localization. We revealed that IFITM1 could be found in sham-opened spinal cords and gradually increased after SCI. It reached peak at 7 and 14 days postinjury (dpi) and still maintained at a relatively higher level at 28 dpi. IHC showed that IFITM1 expressed in GFAP+ and APC+ cells in sham-opened spinal cords. After SCI, in addition to the above-mentioned cells, it could also be found in CD45+ and CD68+ cells, and its expression in CD45+, CD68+, and GFAP+ cells was increased significantly. These results demonstrate that IFITM1 is mainly expressed in astrocytes and oligodendroglia in normal spinal cords, and could rapidly increase in infiltrated leukocytes, activated microglia, and astrocytes after SCI.

Immunohistochemical study of Krüppel-like factor 4 in the spinal cords of rats with experimental autoimmune encephalomyelitis

The expression and localization of Krüppel-like factor (KLF) 4, a class of zinc-finger transcription factors, was investigated in the spinal cords of rats with experimental autoimmune encephalomyelitis (EAE) using western blotting and immunohistochemistry. KLF4 expression was increased significantly in EAE-affected spinal cords compared with normal rat spinal cords. The elevated levels of KLF4 in the spinal cords of rats with EAE remained significant, even during the recovery stage of EAE. The cellular phenotype of KLF4 in EAE lesions consisted of some T cells, macrophages, and reactive astrocytes, whereas it was expressed constitutively in resting astrocytes and neurons, but not in ramified microglial cells in normal spinal cords. Collectively, we postulate that autoimmune T cells and macrophages activate KLF4 and subsequently do not proliferate or exhibit phenotypic switching from M1 to M2 macrophages, respectively. In addition, we hypothesize that the increased and sustained expression of KLF4 in reactive astrocytes during EAE was associated with suppressed CNS inflammation, as well as reduced numbers of pro-inflammatory T cells and M1 macrophages.

The antihyperalgesic effects of intrathecal bupropion, a dopamine and noradrenaline reuptake inhibitor, in a rat model of neuropathic pain.

Antidepressants are often used for the treatment of neuropathic pain, and their analgesic effects rely on increased noradrenaline and serotonin levels in the spinal cord. Clinical studies have also shown that bupropion, a dopamine and noradrenaline reuptake inhibitor, has strong efficacy in neuropathic pain; however, the role of spinal cord dopamine in neuropathic pain is unknown. We hypothesized that bupropion inhibits neuropathic pain by increasing noradrenaline and dopamine in the spinal cord. In the present study, we determined the efficacy and underlying mechanisms of intrathecal administration of bupropion in a rat model of neuropathic pain. Male Sprague-Dawley rats were anesthetized, and right L5 spinal nerve ligation (SNL) was performed to produce mechanical hyperalgesia of the hindpaw. Withdrawal threshold to a paw pressure test was measured before and after intrathecal administration of bupropion, without or with intrathecal antagonists for α2-adrenoceptors and dopamine D2 receptors. In vivo microdialysis was performed in the dorsal horn of the lumbar spinal cord to measure noradrenaline and dopamine concentrations after intrathecal injection of bupropion. We also measured the noradrenaline and dopamine contents in the ipsilateral dorsal lumbar spinal cord in normal rats and in rats 2, 3, and 4 weeks after SNL. Intrathecal injection of bupropion produced a dose-dependent antihyperalgesic effect (3, 10, 30, and 100 μg, P < 0.001). The effect (30 μg) was dose-dependently reversed by intrathecal pretreatment (15 minutes before bupropion injection) with the α2-adrenoceptor antagonist idazoxan (3, 10, and 30 μg, P < 0.001) and D2 receptor antagonist sulpiride (3, 10, and 30 μg, P < 0.001). Microdialysis revealed that noradrenaline and dopamine concentrations in the spinal dorsal horn were increased after intrathecal injection of bupropion (30 μg, P < 0.001 and P = 0.001, respectively). Furthermore, the noradrenaline and dopamine contents in the spinal dorsal horn were increased 2 weeks after SNL (P < 0.001 and P = 0.044, respectively) and then decreased gradually. These findings suggest that plasticity of descending inhibitory pathways such as the noradrenaline and dopamine systems contributes to the maintenance of neuropathic pain and that spinal cord noradrenaline and dopamine both play an inhibitory role in neuropathic pain.

Supraspinal Projection of Serotonergic and Noradrenergic Pathways Modulates Nociceptive Transmission in the Lower Urinary Tract of Rats

To investigate the effect of descending serotonergic and noradrenergic pathways on nociception in the lower urinary tract (LUT). Female Sprague-Dawley rats were used. Following intraperitoneal administration of Vehicle or Milnacipran (30 mg/kg), which is one of serotonin-noradrenaline reuptake inhibitors (SNRI), 0.1% AA was infused into the bladder in normal (n = 4, each) and spinal cord injury (SCI) (n = 4, each) rats for 2 h on consciousness, and c-Fos, 5-HT and DβH were stained using immunohistochemistry at the L6 spinal cord as spinal areas associated with LUT. In SCI rats, 5-HT or DβH-positive fibers were not observed at the L6 spinal cord, while there were many 5-HT and DβH-positive fibers in normal rats. The total number of c-Fos-positive cells was significantly increased in SCI rats compared to Normal rats (209.4 ± 7.1 in Normal, 336.4 ± 28.9 in SCI, P < 0.05), which indicated that interruption of supraspinal modulation enhances nocieptive transmission in the LUT. Regarding the effect of Milnacipran administration, the number of c-Fos-positive cells was significantly decreased at all region of the L6 spinal cord in normal rats (P < 0.05), while this reduction was not observed in SCI rats. This result demonstrated that administration of SNRI attenuates nocieptive transmission in the LUT, indicating that 5-HT and noradrenaline work as mediators of endogenous analgesic mechanisms through the supraspinal descending pain pathways. Supraspinal projections of descending serotonergic and noradrenergic pathways to the lower lumbar spinal cord modulate nocieptive transmission in the LUT. Administration of SNRI attenuates nocieptive transmission in the LUT, which could result from enhancement of modulating descending serotonergic and noradrenergic pathways.

Immunolocalization of Water Channel Proteins AQP1 and AQP4 in Rat Spinal Cord.

Aquaporin (AQP) is a water-selective channel protein. In the brain, AQPs play critical roles in the production of cerebrospinal fluid and in edema formation. In contrast, the expression and role of AQPs in spinal cord are unclear. We aimed to investigate the localization of AQP1 and AQP4 in normal rat spinal cord compared with the expression of marker proteins for astrocytes, neurons, and endothelial cells. Immunohistochemistry demonstrated that AQP1 and AQP4 are expressed along all levels of the spinal cord from the cervical to lumbar levels. AQP1 immunolabeling was observed in the dorsal horns in the gray matter, whereas the labeling was weak and mainly seen close to glia limitans in the white matter. AQP1 was co-labeled with marker proteins for unmyelinated neuronal fibers (peripherin) and endothelial cells (RECA-1) of blood vessels that had penetrated through the glia limitans. In contrast, AQP1 did not colocalize with GFAP, an astrocyte marker, at any level of the spinal cord. AQP4 was exclusively localized at the astrocytes, but AQP4 expression in spinal cord exhibited a less polarized and more spatial distribution than that of brain astrocytes. The observed characteristic localization and expression patterns of AQP1 and AQP4 could provide insights toward gaining an understanding of the role of AQPs in the spinal cord.

Increased Expression of TBP/TFIID after spinal cord injury in adult rats.

Transcription factor IID (TFIID), as a general transcription factor, plays a pivotal role in the preinitiation complex (PIC) assembly and transcription initiation by recruiting RNA polymerase II to the promoter. The TFIID complex contains the TATA-box binding protein (TBP) and a group of conserved TAF proteins. However, its distribution and function in the central nervous system (CNS) are more diverse than previously understood. Here, we mainly investigated the spatiotemporal expression and cellular localization of TBP/TFIID during spinal cord injury (SCI) in adult rats. Western blot analysis revealed that TBP/TFIID was present in normal rat's spinal cord. It gradually increased, reached a peak at the third day after SCI, and then decreased. We observed that TBP/TFIID was widely distributed in spinal cord, mainly in neurons and glial cells. In addition, Western blot detection also showed that the third day post-injury was the proliferation peak indicated by the elevated expression of proliferating cell nuclear antigen (PCNA), a marker of proliferating cells. Importantly, injury-induced expression of TBP/TFIID was colabelled by PCNA showed the increase of TBP/TFIID expression in proliferating astrocytes and microglia. Collectively, we hypothesize that TBP/TFIID may be implicated in the proliferation of astrocytes and microglia and the recovery of neurological outcomes.

Keratan sulfate expression in microglia is diminished in the spinal cord in experimental autoimmune neuritis

Experimental autoimmune neuritis (EAN) is an animal model of Guillain–Barré syndrome, an inflammatory demyelination disease of the peripheral nervous system. Although this disease has been extensively studied on peripheral nerves, the pathology of the central nervous system has not been fully understood. Previous studies demonstrate that expression of keratan sulfate (KS), the sugar chain of proteoglycan, is associated with activated microglia/macrophages accumulated after neuronal injuries. Unexpectedly, we found here that KS is rather diminished in rat EAN. KS was restrictively expressed in microglia in the spinal cord of normal rats. KS was positive in 50% microglia in the ventral horn and 20% in the dorsal horn. In EAN, microglia increased in number and expressed the activation marker CD68, but KS expression was abolished. Concomitantly, pro-inflammatory cytokines, i.e., interferon (IFN)-γ, interleukin (IL)-1β, and tumor necrosis factor (TNF)-α, were increased in the spinal cord of EAN rats, whereas anti-inflammatory cytokines, such as IL-4 and IL-10, were decreased. In addition, silencing of KSGal6ST attenuated KS expression on the primary cultured microglia and upregulated expression of some activation markers (TNF-α, IL-1β, and iNOS) under the stimulation with lipopolysaccharide and IFN-γ. This study demonstrates for the first time a close association of EAN and disappearance of KS on microglia. KS expression could be a useful marker to evaluate the status of polyneuropathy.

Altered rate-dependent depression of the spinal H-reflex as an indicator of spinal disinhibition in models of neuropathic pain

The unpredictable efficacy of current therapies for neuropathic pain may reflect diverse etiological mechanisms operating between, and within, diseases. As descriptions of pain rarely establish specific mechanisms, a tool that can identify underlying causes of neuropathic pain would be useful in developing patient-specific treatments. Rate-dependent depression (RDD), a measure of the change in amplitude of the Hoffman reflex over consecutive stimulations, is attenuated in diabetic rats that also exhibit impaired spinal γ-aminobutyric acid (GABA)A receptor function, reduced spinal potassium chloride co-transporter (KCC2) expression, and indices of painful neuropathy. To investigate whether loss of RDD is a reliable indicator of the contribution of spinal GABAergic dysfunction to neuropathic pain, we assessed RDD, tactile allodynia, and formalin-evoked hyperalgesia in 3 models: rats treated acutely with brain-derived neurotrophic factor (BDNF), diabetic rats treated with the BDNF-sequestering molecule tyrosine receptor kinase B/Fc (TrkB/Fc), and rats with paclitaxel-induced neuropathy. Delivery of BDNF to the spinal cord of normal rats produced RDD deficits and features of painful neuropathy associated with disrupted GABAA receptor-mediated inhibitory function and reduced dorsal spinal KCC2 expression. Treating diabetic rats with TrkB/Fc restored RDD and alleviated indices of painful neuropathy. In paclitaxel-treated rats, RDD was not impaired and behavioral indices of neuropathic pain were not associated with spinal GABAergic dysfunction or reduced dorsal spinal KCC2 expression. Our data reveal BDNF as part of the mechanism underlying spinal cord disinhibition caused by altered GABAA receptor function in diabetic rats and suggest that RDD deficits may be a useful indicator of neuropathic pain states associated with spinal disinhibition, thereby revealing specific therapeutic targets.

Angiotensin type 2 receptors in the intermediolateral cell column of the spinal cord: Negative regulation of sympathetic nerve activity and blood pressure

Our previous study demonstrated that AT2R in brainstem nuclei participated in the regulation of sympathetic outflow and cardiovascular function. However, the functional significance of AT2R in the intermediolateral cell column (IML) of the thoracic spinal cord in normal rats remains elusive. We hypothesized that AT2R activation in the IML exerts a sympatho-inhibitory effect. Using Western-blot analysis, immunohistochemical staining and quantitative real-time PCR, both AT1R and AT2R expressions were detected in the spinal cord. The highest AT2R protein expression was found in the IML, while AT1R expression didn't display regional differences within the gray matter. Microinjection of Ang II into the IML dose-dependently elevated mean blood pressure (MAP, employing a transducer-tipped catheter) and renal sympathetic nerve activity (RSNA, using a pair of platinum-iridium recording electrodes), which were completely abolished by Losartan, and attenuated by TEMPOL and apocynin. Activation of AT2R in the IML with CGP42112 evoked hypotension (ΔMAP: -21 ± 4 mmHg) and sympatho-inhibition (RSNA: 73 ± 3% of baseline), which were completely abolished by PD123319 and l-NAME. Blockade of AT2R in the IML with PD123319 significantly increased MAP (11 ± 1 mmHg) and sympathetic nerve activity (RSNA: 133 ± 13% of baseline). Moreover, PD123319 significantly enhanced the Ang II induced pressor response. Furthermore, in isolated IML neurons, CGP42112 treatment augmented potassium current and decreased resting membrane potential by employing whole-cell patch clamp. In the normal condition, AT2R in the IML tonically inhibits sympathetic activity through an NO/NOS dependent pathway and subsequent potassium channel activation.

275 SUPRASPINAL PROJECTION OF SEROTONERGIC AND NORADRENERGIC PATHWAYS MODULATES NOCICEPTIVE TRANSMISSION IN THE LOWER URINARY TRACT OF RATS

INTRODUCTION AND OBJECTIVES: Although 5-hydroxytryptamine (5-HT) and norepinephrine have been implicated as mediators of endogenous analgesic mechanisms in the supraspinal descending pain pathways, the effect of these supraspinal projection are still unknown in the lower urinary tract (LUT). In the present study, we therefore investigated the effect of descending serotonergic and noradrenergic pathways on nociception in the LUT. METHODS: Female Sprague-Dawley rats were used. Regarding spinal cord injury (SCI) rats, the spinal cord was completely cut with scissors and experiment was performed at 8 weeks after SCI. Following intraperitoneal administration of Vehicle or Milnacipram (30mg/kg), which is a serotonin-norepinephrine reuptake inhibitor (SNRI), 0.1% acetic acid (AA) was infused into the bladder in normal and SCI rats for 2 hours on consciousness, and c-fos, 5-HT and dopamine-beta-hydroxylase (D H) were stained using immunohistochemistry at the L6 spinal cord. RESULTS: In SCI rats, 5-HT or D H-positive fibers were not observed at the L6 spinal cord, while there were many 5-HT and D H-positive fibers in normal rats. The total number of c-fos-positive cells was significantly increased in SCI rats compared to Normal rats (209.4 7.1 in Normal, 336.4 28.9 in SCI, p 0.05), which indicated that interruption of supraspinal modulation enhances nocieptive transmission in the LUT. Regarding the effect of Milnacipram administration, the number of c-fos-positive cells was significantly decreased at all region of the L6 spinal cord in normal rats, while this reduction was not observed in SCI rats (Fig, *: p 0.05).This result demonstrated that administration of a SNRI attenuates nocieptive transmission in the LUT, which means that 5-HT and norepinephrine work as mediators of endogenous analgesic mechanisms in the supraspinal descending pain pathways. CONCLUSIONS: Supraspinal projections of descending serotonergic and noradrenergic pathways in to the lumbosacral spinal cord modulate nocieptive transmission in the LUT. Administration of a SNRI attenuates nocieptive transmission in the LUT, which could result from enhancement of modulating descending serotonergic and noradrenergic pathways. Source of Funding: None

Temporospatial expression and cellular localization of glutamine synthetase following traumatic spinal cord injury in adult rats

Glutamine synthetase (GS) is an enzyme involved in an endogenous mechanism of protection against glutamate neurotoxicity and is important in the regulation of astrocyte migration. To date, limited information is available concerning the expression of GS in normal spinal cords and following injury. In the present study, GS expression was identified in astrocytes, oligodendrocytes and microglia in normal rat spinal cords. Following traumatic spinal cord injury (SCI), the glutamate concentration increased rapidly at 1h and returned to baseline rapidly. However, the GS activity and protein levels were found to decrease at 4h and then increase gradually from day3 following SCI. The quantification of astrocytes, oligodendrocytes and activated microglia/macrophages, as well as immunohistochemistry staining of day7 post‑injured spinal cords, indicated that the astrocytes and microglia/macrophages contributed to the increase in GS. Collectively, the results provided evidence for the temporospatial expression and location of GS following SCI and suggested that the changes in GS levels may contribute to glutamate neurotoxicity and glial cell response following SCI.

Response of the rat spinal cord to X-ray microbeams

Background and purposeTo quantify the late dose-related responses of the rat cervical spinal cord to X-ray irradiations by an array of microbeams or by a single millimeter beam. Materials and methodsNecks of anesthetized rats were irradiated transversely by an 11mm wide array of 52 parallel, 35μm wide, vertical X-ray microbeams, separated by 210μm intervals between centers. Comparison was made with rats irradiated with a 1.35mm wide single beam of similar X-rays. Rats were killed when paresis developed, or up to 383days post irradiation (dpi). ResultsMicrobeam peak/valley doses of ≈357/12.7Gy to 715/25.4Gy to an 11mm long segment of the spinal cord, or single beam doses of ≈146–454Gy to a 1.35mm long segment caused foreleg paresis and histopathologically verified spinal cord damage; rats exposed to peak/valley doses up to 253/9Gy were paresis-free at 383dpi. ConclusionsWhereas microbeam radiation therapy [MRT] for malignant gliomas implanted in rat brains can be safe, palliative or curative, the high tolerance of normal rat spinal cords to similar microbeam exposures justifies testing MRT for autochthonous malignancies in the central nervous system of larger animals with a view to subsequent clinical applications.

Gene Expression in the Spinal Cord in Female Lewis Rats with Experimental Autoimmune Encephalomyelitis Induced with Myelin Basic Protein

BackgroundExperimental autoimmune encephalomyelitis (EAE), the best available model of multiple sclerosis, can be induced in different animal strains using immunization with central nervous system antigens. EAE is associated with inflammation and demyelination of the nervous system. Micro-array can be used to investigate gene expression and biological pathways that are altered during disease. There are few studies of the changes in gene expression in EAE, and these have mostly been done in a chronic mouse EAE model. EAE induced in the Lewis with myelin basic protein (MBP-EAE) is well characterised, making it an ideal candidate for the analysis of gene expression in this disease model.Methodology/Principal FindingsMBP-EAE was induced in female Lewis rats by inoculation with MBP and adjuvants. Total RNA was extracted from the spinal cords and used for micro-array analysis using AffimetrixGeneChip Rat Exon 1.0 ST Arrays. Gene expression in the spinal cords was compared between healthy female rats and female rats with MBP-EAE. Gene expression in the spinal cord of rats with MBP-EAE differed from that in the spinal cord of normal rats, and there was regulation of pathways involved with immune function and nervous system function. For selected genes the change in expression was confirmed with real-time PCR.Conclusions/SignificanceEAE leads to modulation of gene expression in the spinal cord. We have identified the genes that are most significantly regulated in MBP-EAE in the Lewis rat and produced a profile of gene expression in the spinal cord at the peak of disease.

Ephrin-B3 Decreases the Survival of Adult Rat Spinal Cord-Derived Neural Stem/Progenitor Cells In Vitro and After Transplantation into the Injured Rat Spinal Cord

Although transplantation of neural stem/progenitor cells (NSPC) encourages regeneration and repair after spinal cord injury (SCI), the survival of transplanted NSPC is limited. Ephrin-B3 has been shown to reduce the death of endogenous NSPC in the subventricular zone of the mouse brain without inducing uncontrolled proliferation. Due to similarities in the environment of the brain and spinal cord, we hypothesized that ephrin-B3 might reduce the death of both transplanted and endogenous spinal cord-derived NSPC. Both normal and injured (26 g clip compression) spinal cords were examined. Ephrin-B3-Fc was tested, and Fc fragments and phosphate-buffered saline (PBS) were used as controls. We found that EphA4 receptors were expressed by spinal cord-derived NSPC and expressed in the normal and injured rat spinal cord (higher expression in the latter). In vitro, ephrin-B3-Fc did not significantly reduce the survival of NSPC except at 1 μg/mL (P<0.05), but Fc fragments alone reduced NSPC survival at all doses in a dose-dependent fashion. In vivo, intrathecal infusion of ephrin-B3-Fc increased the proliferation of endogenous ependymal cells and the proportion of proliferating cells that expressed the glial fibrillary acidic protein astrocytic marker in the injured spinal cord compared with the infusion of PBS (P<0.05). However, in the injured spinal cord, the infusion of either ephrin-B3-Fc or Fc fragments alone caused a 20-fold reduction in the survival of transplanted NSPC (P<0.001). Thus, after SCI, ephrin-B3-Fc and Fc fragments are toxic to transplanted NSPC.

The spinal notch signaling pathway plays a pivotal role in the development of neuropathic pain

BackgroundThe Notch signaling pathway has been shown to be involved in the development of the nervous system. Recent studies showed that Notch receptors and ligands are also expressed in the nervous system of adult animals. However, whether the Notch signaling pathway has a function in adults is not fully understood. The present study is designed to investigate the function of the Notch signaling pathway in nociceptive transmission, especially during neuropathic pain in adult rats.ResultsWe found that the Notch intracellular domain (NICD) is expressed in the DRG (Dorsal Root Ganglia), sciatic nerve and spinal cord in normal rats, and is upregulated in the sciatic nerve and spinal cord after spared nerve injury (SNI). Moreover, we used the γ-secretase (a key enzyme of the Notch signaling pathway) inhibitor DAPT to observe the effect of the Notch signaling pathway after SNI. We found that intrathecal DAPT significantly increased paw withdrawal thermal latency and mechanical threshold. Mechanical hyperalgesia occurring after SNI could be significantly reversed by DAPT in a dose-dependent manner.ConclusionsThese results suggest that the Notch signaling pathway participates in the induction and maintenance of neuropathic pain, which indicates that the Notch pathway maybe a potential drug target for neuropathic pain treatment.

Antihypersensitivity Effects of Tramadol Hydrochloride in a Rat Model of Postoperative Pain

Tramadol is used to treat a wide range of acute and chronic pain. This drug induces analgesia by 2 mechanisms of action: opioid receptor activation and enhancement of noradrenaline (NA) and serotonin (5-HT) transmission. The effect of tramadol on NA and 5-HT concentrations in the spinal cord, however, have not been assessed. In the present study, we investigated the antihypersensitivity effect of tramadol using a rat model of postoperative pain. We also evaluated the increase in NA and 5-HT levels in the spinal cord after tramadol injection using in vivo microdialysis. We made a hindpaw incision in male Sprague-Dawley rats (postoperative pain model). Tramadol was administered intraperitoneally and intrathecally 24 hours after paw incision. Mechanical hypersensitivity was measured by determining the withdrawal threshold using von Frey filaments. Microdialysis studies from the dorsal horn of the lumbar spinal cord were performed to measure NA and 5-HT levels after intraperitoneal injection of tramadol. We also measured the NA and 5-HT content in the spinal cord in normal rats and rats with paw incision. Intraperitoneal (10, 20, and 40 mg/kg) and intrathecal (125, 250, and 500 μg) injection of tramadol produced an antihyperalgesic effect in a dose-dependent manner. The antihypersensitivity effect of tramadol was prevented by intrathecal pretreatment with methysergide (30 μg), a serotonin receptor antagonist; idazoxane (30 μg), a noradrenaline receptor antagonist; and naloxone (30 μg), a nonselective opioid receptor antagonist. Microdialysis study revealed that 5-HT and NA concentrations at the spinal dorsal horn were increased, peaking at 30 minutes after intraperitoneal injection of 20 mg/kg tramadol. Furthermore, the NA and 5-HT content in the ipsilateral dorsal half of the lumbar spinal cord was increased 1 day and 3 days after paw incision, respectively. These findings indicate that tramadol inhibits postoperative hypersensitivity by increasing NA and 5-HT levels in the spinal cord and activating opioid receptors. Tramadol might be more effective in the early postoperative period when spinal NA and 5-HT levels are increased.

Neuroendocrine Mechanisms of Acupuncture

Neuroendocrine Mechanisms of Acupuncture

Expression of bone morphogenetic proteins and their receptors in the normal adult rat spinal cord.

To observe the expression distribution of bone morphogenetic proteins (BMP) in the spinal cord of normal adult rats. Expression of BMP2, BMP4, and BMP7, and their receptors BMPR Ia, BMPR Ib, and BMP II were detected by immunochemistry analysis in the spinal cord of normal adult rats. Expression of BMPR Ia or BMPR Ib was observed in the motor neurons of the anterior horn, sensory neurons of the dorsal horn, oligodentrocytes, some microglia, and some astrocytes. Expression of receptor BMPR II was found in the oligodentrocytes and motor neurons in the gray matter of anterior horn. It was also expressed in some glial fibrillary acidic protein (GFAP)-positive astrocytes in the white matter but not in the gray matter. BMP2 and BMP4 were not expressed in the spinal cord of normal adult rats by immunohistochemistry. BMP7 was expressed in all the APC-positive oligodentrocytes, all the NeuN-positive motor neurons in the anterior horn, and some astrocytes in the normal spinal cord. Phosphated pSmad 1/5/8 protein was expressed in all the oligodentrocytes, all the neurons, and some astrocytes, especially in the GFAP-positive astrocytes which were RC2-positive radial glia in the subventricular zone. BMP7, BMP receptors, and phosphated pSmad 1/5/8 are expressed in many types of cells whereas BMP2 and BMP4 are not expressed in the spinal cord of normal adult rats, which suggests an important function of BMP signal pathway in the neuron and glia of spinal cord.