Neurons In Lumbar Spinal Cord Research Articles

Changes in the number of macrophages, T-lymphocytes, activity of antioxidant enzymes in the brain, behavior and structure of the central nervous system neurons in adult and aging mice of different strains with the MPTP-induced model of parkinsonism

Background. Oxidative stress and immune cell (T-lymphocytes, macrophages) products are important for the development of morpho-functional disorders of the nervous system in Parkinson’s disease. Connection of Parkinson’s disease with age and functioning of the major histocompatibility complex genes are also shown. The purpose was to assess changes in the brain of T-lymphocytes, macrophages, malondialdehyde (MDA) contents, the activity of antioxidant enzymes, the structure of brain and spinal cord neurons, as well as behavior in adult and aging mice with different H-2 haplotypes and toxic model of parkinsonism. Materials and methods. Adult and aging female mice of FVB/N (genotype H-2q) and 129/Sv (genotype H-2b) strains were once injected with the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) at the dose of 30 mg/kg. Contents of CD3+, CD3–CD11b+, CD3+CD11b+ cells, MDA and the activity of antioxidant enzymes in the brain were evaluated. The structure of neurons of the substantia nigra, lumbar spine, behavior in the open field test, as well as in rigidity and rotarod performance tests were studied. Results. In adult mice of both strains, motor and non-motor (spatial-exploratory, emotional) activity is impaired under the influence of MPTP. At the same time, linear differences in behavior changes were revealed in their directions and expressiveness. In aging FVB/N experimental mice, motor behavior disorders prevailed and were combined with non-motor changes in 129/Sv mice. Violations in the structure of substantia nigra neurons after MPTP administration were more severe in FVB/N mice while those in the lumbar spinal cord neurons were more pronounced in the 129/Sv mice. In aging experimental mice of both strains, the percentage of damaged neurons in the brain and spinal cord was significantly lower than in adult animals. After MPTP administration to adult mice, the direction of changes in the brain T-lymphocytes and macrophages (increase or decrease) depended on their strains. In aging experimental mice, the linear heterogeneity of changes was preserved for the T-lymphocyte content, while the macrophage level was increased regardless of the animal strains. Under MPTP influence, the MDA content increased in the brain of mice of all experimental groups. The direction of changes in superoxide dismutase, catalase, glutathione peroxidase and glutathione reductase activities (decrease or increase) depended on the strains of animals and their age. Conclusions. MPTP-induced changes (directions, expressiveness) in T-lymphocyte, macrophage contents, antioxidant enzymes activity, the structure of neurons of the substantia nigra and lumbar spine, as well as behavioral reactions largely depended on the adult mice H-2 haplotype. There were age-related effects of the neurotoxin on changes in the studied indicators in mice of different strains. At the same time, dependence of changes in most above indicators on the H-2 haplotype preserved.

Neuroregeneration of injured peripheral nerve by fraction B of catfish epidermal secretions through the reversal of the apoptotic pathway and DNA damage.

Introduction: Crush injuries occur from acute traumatic nerve compression resulting in different degrees of neural damage leading to permanent functional deficits. Recently, we have shown that administration of Fraction B (FB) derived from catfish epidermal secretions accelerates healing of damaged nerve in a sciatic nerve crush injury, as it ameliorates the neurobehavioral deficits and enhances axonal regeneration, as well as protects spinal neurons and increases astrocytic activity and decreasing GAP-43 expression. The present study aimed to investigate the role of FB treatment on the apoptotic pathway in the neuroregeneration of the sciatic nerve crush injury. Methods: Male Wistar rats were randomly assigned into five groups: (I) SHAM, (II) CRUSH, (III) CRUSH + (1.5mg/kg) FB, (IV) CRUSH + (3mg/kg) FB, and (V) CRUSH + (4.5mg/kg) FB. Rats underwent sciatic nerve crush surgery, followed by treatment with FB administered intraperitoneally (IP) daily for two weeks and then sacrificed at the end of the fourth week. Results: FB improved the recovery of neurobehavioral functions with a concomitant increase in axonal regeneration and neuroprotective effects on spinal cord neurons following crush injury. Further, FB enhanced Schwann cells (SCs) proliferation with a significant increase in myelin basic protein expression. FB-treated animals demonstrated higher numbers of neurons in the spinal cord, possibly through ameliorating oxidative DNA damage and alleviating the mitochondrial-dependent apoptotic pathway by inhibiting the release of cytochrome c and the activation of caspase-3 in the spinal cord neurons. Conclusion: FB alleviates the neurodegenerative changes in the lumbar spinal cord neurons and recovers the decrease in the neuronal count through its anti-apoptotic and DNA antioxidative properties.

Spinal Cord Stimulation Attenuates Below-Level Mechanical Hypersensitivity in Rats After Thoracic Spinal Cord Injury

ObjectivesThe burden of pain after spinal cord injury (SCI), which may occur above, at, or below injury level, is high worldwide. Spinal cord stimulation (SCS) is an important neuromodulation pain therapy, but its efficacy in SCI pain remains unclear. In SCI rats, we tested whether conventional SCS (50 Hz, 80% motor threshold [MoT]) and 1200 Hz, low-intensity SCS (40% MoT) inhibit hind paw mechanical hypersensitivity, and whether conventional SCS attenuates evoked responses of wide-dynamic range (WDR) neurons in lumbar spinal cord. Materials and MethodsMale rats underwent a moderate contusive injury at the T9 vertebral level. Six to eight weeks later, SCS or sham stimulation (120 min, n = 10) was delivered through epidural miniature electrodes placed at upper-lumbar spinal cord, with using a crossover design. Mechanical hypersensitivity was examined in awake rats by measuring paw withdrawal threshold (PWT) to stimulation with von Frey filaments. WDR neurons were recorded with in vivo electrophysiologic methods in a separate study of anesthetized rats. ResultsBoth conventional SCS and 1200 Hz SCS increased PWTs from prestimulation level in SCI rats, but the effects were modest and short-lived. Sham SCS was not effective. Conventional SCS (10 min) at an intensity that evokes the peak Aα/β waveform of sciatic compound action potential did not inhibit WDR neuronal responses (n = 19) to graded or repeated electrical stimulation that induces windup. ConclusionsConventional SCS and 1200 Hz, low-intensity SCS modestly attenuated below-level mechanical hypersensitivity after SCI. Inhibition of WDR neurons was not associated with pain inhibition from conventional SCS.

Differential modulation of excitatory and inhibitory populations of superficial dorsal horn neurons in lumbar spinal cord by Aβ-fiber electrical stimulation.

Activation of Aβ-fibers is fundamental to numerous analgesic therapies, yet its effects on dorsal horn neuronal activity remain unclear. We used multiphoton microscopy of the genetically encoded calcium indicator GCaMP6s to characterize the effects of Aβ-fiber electrical stimulation (Aβ-ES) on neural activity. Specifically, we quantified somatic responses evoked by C-fiber intensity stimulation before and after a 10-minute train of dorsal root Aβ-ES in superficial dorsal horn (SDH) neurons, in mouse lumbar spinal cord. Aβ-ES did not alter C-fiber-evoked activity when GCaMP6s was virally expressed in all neurons, in an intact lumbar spinal cord preparation. However, when we restricted the expression of GCaMP6s to excitatory or inhibitory populations, we observed that Aβ-ES modestly potentiated evoked activity of excitatory neurons and depressed that of inhibitory neurons. Aβ-ES had no significant effects in a slice preparation in either SDH population. A larger proportion of SDH neurons was activated by Aβ-ES when delivered at a root rostral or caudal to the segment where the imaging and C-fiber intensity stimulation occurred. Aβ-ES effects on excitatory and inhibitory populations depended on the root used. Our findings suggest that Aβ-ES differentially modulates lumbar spinal cord SDH populations in a cell type- and input-specific manner. Furthermore, they underscore the importance of the Aβ-ES delivery site, suggesting that Aβ stimulation at a segment adjacent to where the pain is may improve analgesic efficacy.

Motor Neurons Pathology After Chronic Exposure to MPTP in Mice.

The neurotoxin 1-methyl,4-phenyl-1,2,3,6-tetrahydropiridine (MPTP) is widely used to produce experimental parkinsonism in rodents and primates. Among different administration protocols, continuous or chronic exposure to small amounts of MPTP is reported to better mimic cell pathology reminiscent of Parkinson's disease (PD). Catecholamine neurons are the most sensitive to MPTP neurotoxicity; however, recent studies have found that MPTP alters the fine anatomy of the spinal cord including motor neurons, thus overlapping again with the spinal cord involvement documented in PD. In the present study, we demonstrate that chronic exposure to low amounts of MPTP (10mg/kg daily, × 21days) significantly reduces motor neurons in the ventral lumbar spinal cord while increasing α-synuclein immune-staining within the ventral horn. Spinal cord involvement in MPTP-treated mice extends to Calbindin D28 KDa immune-reactive neurons other than motor neurons within lamina VII. These results were obtained in the absence of significant reduction of dopaminergic cell bodies in the Substantia Nigra pars compacta, while a slight decrease was documented in striatal tyrosine hydroxylase immune-staining. Thus, the present study highlights neuropathological similarities between dopaminergic neurons and spinal motor neurons and supports the pathological involvement of spinal cord in PD and experimental MPTP-induced parkinsonism. Remarkably, the toxic threshold for motor neurons appears to be lower compared with nigral dopaminergic neurons following a chronic pattern of MPTP intoxication. This sharply contrasts with previous studies showing that MPTP intoxication produces comparable neuronal loss within spinal cord and Substantia Nigra.

Lumbar spinal cord neurons putatively involved in ejaculation are sexually dimorphic in early postnatal mice.

A crucial role in ejaculation is thought to be played by a population of lumbar spino-thalamic neurons (LSt), which express galanin and other neuropeptides. In rats, these neurons are activated with ejaculation and their lesion selectively abolishes ejaculation but not other mating behaviors. Consistently with their role, in adult rats and humans, LSt neurons are sexually dimorphic, being more numerous in males. Here we examined whether sexual dimorphism arises early in development, using a transgenic mouse line in which the expression of fluorescent protein is driven by the galanin promoter. We focused on postnatal day 4, shortly after a transient perinatal androgen surge in males that could play an organizational role in LSt development. We found a population of brightly fluorescent neurons organized in bilateral columns dorsolateral to the central canal in segments L1-L5, the expected location of the LSt group. Their number was close to that of adult preparations and significantly greater in male than in female siblings (+19%; CI95% : +13% to +27%; p < .01). This was not due to a generalized higher galanin expression in the male since fluorescent L4 DRG neurons, innervating the hindlimbs and lower back, were not significantly dimorphic (-4%; CI95% : -10% to +8%; p = .92). Unexpectedly, we found in cervical segments a population of fluorescent neurons having a location relative to the central canal similar to the LSt. Thus, the LSt group is sexually dimorphic soon after birth. However, it is possible that only a subset of its neurons participate in the control of ejaculation.

PS-04-009 The lumbar spinal cord neurons putatively involved in ejaculation are sexually dimorphic already in pre-pubertal mice

The expression of ejaculation is thought to require a well-defined population of lumbar spino-thalamic (LSt) neurons which co-express several neuropeptides, including galanin. In adult rats and humans LSt neurons are significantly more numerous in males than in females. Here we investigated whether this sexual dimorphism is established already before puberty. A transgenic mouse line expressing green fluorescent protein driven by the galanin promoter, was used to directly visualize galaninergic neurons and fibers in the postnatal spinal cord (P4). Transverse sections of the cord were made to map the segmental distribution of galanin expression. Wholemount spinal cords were fixed with paraformaldehyde and cleared with a fructose-based protocol to view and count entire neuron pools, comparing male vs. female siblings from the same litter. Image stacks were acquired with a brightfield fluorescence microscope, deconvolved and analyzed in ImageJ software. Fluorescent reporter expression in the early postnatal mouse largely matched what previously described for galanin using immunohistochemistry in adult tissue. A notable exception was evidence for galanin expression in motoneurons, a phenomenon known to occur transiently during ontogenesis in rat and human. Crucially, brightly fluorescent neurons were present in the expected location of the LSt population (the lumbar enlargement, dorsolaterally of the ependymal canal) in both males and females. Moreover, in each pair of siblings (n=6) they were more numerous in the male than in the female (by 22±4% mean±sd; p=0.03 Wilcoxon paired test; median=1029 in males, 855 in females).

Protective effects of Withania somnifera extract in SOD1G93A mouse model of amyotrophic lateral sclerosis

Withania somnifera (WS; commonly known as Ashwagandha or Indian ginseng) is a medicinal plant whose extracts have been in use for centuries in various regions of the world as a rejuvenator. There is now a growing body of evidence documenting neuroprotective functions of the plant extracts or its purified compounds in several models of neurodegenerative diseases including amyotrophic lateral sclerosis (ALS). Based on the extract's beneficial effect in a mouse model of ALS with TDP-43 proteinopathy, the current study was designed to test its efficacy in another model of familial ALS. Our results show that administration of WS extracts by gavage to mice expressing G93A mutant form of superoxide dismutase (SOD1) resulted in increased longevity, improved motor performance and increased number of motor neurons in lumbar spinal cord. The WS treatment caused substantial reduction in levels of misfolded SOD1whereas it enhanced expression of cellular chaperons in spinal cord of SOD1G93A mice. WS markedly reduced glial activation and prevented phosphorylation of nuclear factor kappaB (NF-κB). The overall immunomodulatory effect of WS was further evidenced by changes in expression of multiple cytokines/chemokines. WS also served as an autophagy inducer which may be beneficial at early stages of the disease. These results suggest that WS extracts might constitute promising therapeutics for treatment of ALS with involvement of misfolded SOD1.

Effects of resveratrol on motor function and anterior horn neuron of spinal cord following acute sciatic nerve compression injury in rats

Objective To investigate the effects of resveratrol (Res) on motor function and the anterior horn neuron of lumbar spinal cord after acute sciatic nerve compression injury in rats. Methods The rat model with acute sciatic nerve compression injury was established in 32 Sprague Dawley (SD) rats, which were randomly divided into four groups: Res group, Dimethyl Sulfoxide (DMSO) group, Normal Saline (NS) group and sham-operation group. Res, DMSO and saline were successively injected by intraperitoneal for 10 days after established crush acute sciatic nerve compression injury model, while sham-operation group was sutured only after exposure to the sciatic nerve. The weight, the change of toe extension angle, and the sciatic functional index (SFI) of rats were observed at the 1st day before operation and the 1st, 3rd, 7th, and 10th days after surgery. The expressions of neuron-specific nuclear protein (NeuN) and microtubule-associate protein 2 (MAP2) were detected by immunofluorescent staining of L4-L6 spinal cord anterior horn on the 10th day after surgery. Results No significant changes were found in the weight of rats among four groups. Compared to the sham, the motor function of the injured limb in Res, DMSO, and NS rats was impaired, and the anterior horn neurons were seriously damaged. But the differences of the change of toe extension and the sciatic functional index of rats were significantly higher in Res group than that of the DMSO group and NS group (P<0.01) at the 3rd, 7th, and 10th days after surgery. The expressions of NeuN and MAP2 in the anterior horn of rat lumbar spinal cord were up-regulated in Res group relative to DMSO and NS, and the number of neurons in the lumbar spinal cord was significantly relieved at the 10th days. Conclusions Res was significant to rat model of acute sciatic nerve injury, which could increase the number of neurons in the anterior horn of the spinal cord and help the recovery of motor function. Key words: Veratrum alkaloids/PD; Sciatic nerve/IN; Motor activity/DE; Anterior horn cells/DE

Disabled-1 dorsal horn spinal cord neurons co-express Lmx1b and function in nociceptive circuits.

The Reelin-signaling pathway is essential for correct neuronal positioning within the central nervous system. Mutant mice with a deletion of Reelin, its lipoprotein receptors, or its intracellular adaptor protein Disabled-1 (Dab1), exhibit nociceptive abnormalities: thermal (heat) hyperalgesia and reduced mechanical sensitivity. To determine dorsal horn alterations associated with these nociceptive abnormalities, we first characterized the correctly positioned Dab1 neurons in wild-type and mispositioned neurons in Reelin-signaling pathway mutant lumbar spinal cord. Using immunofluorescence, we found that 70% of the numerous Dab1 neurons in Reln+/+ laminae I-II and 67% of those in the lateral reticulated area and lateral spinal nucleus (LSN) co-express the LIM-homeobox transcription factor 1 beta (Lmx1b), an excitatory glutamatergic neuron marker. Evidence of Dab1- and Dab1-Lmx1b neuronal positioning errors was found within the isolectin B4 terminal region of Reln-/- lamina IIinner and in the lateral reticulated area and LSN, where about 50% of the Dab1-Lmx1b neurons are missing. Importantly, Dab1-Lmx1b neurons in laminae I-II and the lateral reticulated area express Fos after noxious thermal or mechanical stimulation and thus participate in these circuits. In another pain relevant locus - the lateral cervical nucleus (LCN), we also found about a 50% loss of Dab1-Lmx1b neurons in Reln-/- mice. We suggest that extensively mispositioned Dab1 projection neurons in the lateral reticulated area, LSN, and LCN and the more subtle positioning errors of Dab1 interneurons in laminae I-II contribute to the abnormalities in pain responses found in Reelin-signaling pathway mutants.

Evidence for the involvement of descending pain-inhibitory mechanisms in the attenuation of cancer pain by carvacrol aided through a docking study

AimsThe present study evaluated the carvacrol (CARV) effect on hyperalgesia and nociception induced by sarcoma 180 (S180) in mice. Main methodsCarvacrol treatment (12.5–50mg/kgs.c.) once daily for 15days was started 24h after injection of the sarcoma cells in the hind paw (s.c.). Mice were evaluated for mechanical sensitivity (von Frey), spontaneous and palpation-induced nociception, limb use and tumor growth on alternate days. CARV effects on the central nervous system were evaluated through immunofluorescence for Fos protein. Molecular docking studies also were performed to evaluate intermolecular interactions of the carvacrol and muscimol, as ligands of interleukin-10 and GABAA receptors. Key findingsCARV was able to significantly reduce mechanical hyperalgesia and spontaneous and palpation-induced nociception, improve use paw, decrease the number of positively marked neurons in lumbar spinal cord and activate periaqueductal gray, nucleus raphe magnus and locus coeruleus. CARV also caused significant decreased tumor growth. Docking studies showed favorable interaction overlay of the CARV with IL-10 and GABAA. SignificanceTogether, these results demonstrated that CARV may be an interesting option for the development of new analgesic drugs for the management of cancer pain.

Prenatal effects of retinoic acid on lumbar spinal cord development and liver antioxidants in rats

During embryonic and early postnatal development, retinoic acid (RA) regulates genes that control neuronal differentiation and neurite outgrowth from the neural tube. The effects of high levels of RA on the CNS can be detected via nitric oxide (NO), which plays a crucial role in neural transmission. The aim of the study was to investigate the prenatal influence of high levels of RA on postnatal development of nitrergic structures in lumbar spinal cord and antioxidant status. RA was administered orally at a dose of 10mg/kg body weight to pregnant female Wistar rats during days 8–10 of gestation. Neuronal nitric oxide synthase (nNOS) of lumbar spinal cord sections was processed for visualization via nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) histochemistry on postnatal day one, day twenty-one and in adults. The results suggest that prenatal administration of high levels of RA is not associated with postnatal morphological changes in nNOS-positive neurons in the rat lumbar spinal cord. An estimation of the activity of enzymes related to the storage of retinoid in the liver showed possible side effects. Suppression and deepening of superoxide dismutase activity persisted into adulthood, and a concurrent downregulation of glutathione reductase was noted. A decrease in reduced glutathione persisted until adulthood when other compensatory mechanisms were probably active to maintain an appropriate level.

Morphological changes and altered expression of antioxidant proteins in a heterozygous dynein mutant; a mouse model of spinal muscular atrophy.

There is increased evidence that oxidative stress is involved in exacerbations of neurodegenerative diseases and spinal muscular atrophies. We examined changes in morphology and expression of antioxidant proteins and peroxiredoxins in motor neurons of lumbar spinal cord, dorsal root ganglion sensory neurons, macroglial cells and quadriceps muscles of newborn heterozygous Loa/+ mice ("legs at odd angles"), a mouse model for early onset of the spinal muscular atrophy with lower extremity predominance (SMA-LED). Our data indicate that newborn Loa-mice develop: neuroinflammation of the sensory and motor neurons; muscular inflammation with atrophic and denervated myofibers; increased expression of neuronal mitochondrial peroxiredoxins (Prxs) 3, 5 and cytoplasmic Prx 6 in motor and sensory neurons, myofibers, fibroblasts of perimysium and chondrocytes of cartilage; and decreased expression of Prx 6 by glial cells and in extracellular space surrounding motor neurons. The decrease in expression of Prx 6 by glial cells and extracellular Prx 6 secretion in early stages of the pathological conditions is consistent with the hypothesis that chronic oxidative stress may lead to neurodegeneration of motor neurons and exacerbation of the pathology.

Sensory denervation with capsaicin reduces ovarian follicular development and delays the onset of puberty in guinea pigs

Introduction: It has been documented that mammalian ovaries receive sympathetic, parasympathetic and sensory nerve fibers. The aim of this work was to investigate the effects of sensory denervation with capsaicin at the first vaginal opening (FVO) on follicular development and the expression of TRPV1 receptors in ovary cells as well as in the dorsal root ganglia (DRGs) and lumbar dorsal spinal cord neurons of guinea pigs. The DRGs and lumbar dorsal spinal cord neurons serve as a nerve connection from the ovaries to the CNS. Materials and Methods: Female guinea pigs received a subcutaneous injection of capsaicin (30 mM) at 10 days of age (P10), while control animals were injected with vehicle. Using light microscopy, we counted healthy preantral follicles (HPF), healthy antral follicles (HAF), atretic preantral follicles (APF), and atretic antral follicles (AAF) in the ovaries at the FVO, and the numbers of TRPV1-positive cells were counted in the ovarian follicles, DRGs, and lumbar dorsal spinal cord (L2-L4) neurons by immunohistochemistry. Results: Guinea pigs treated with capsaicin showed a significant delay of FVO in comparison with the control animals (36 vs. 44 days). In the ovaries, the number of preantral and antral follicles decreased significantly. Additionally, the number of TRPV1-positive thecainterstitial cells of the antral follicles was reduced significantly, and the number of TRPV1-positive neurons in the DRGs and lumbar dorsal spinal cord (L2-L4) decreased. Thus, we showed that TRPV1 receptors throughout the sensory fibers modulate ovarian follicular development and the onset of puberty in guinea pigs. Conclusion: Sensory denervation decreases ovarian follicular development and delays the onset of puberty of guinea pigs. Our data support the idea that through TRPV1 receptors, ovarian afferent fibers sense local stimuli that are sent to the CNS.

The effect of olfactory ensheathing cell transplantation combined with walking training on neurofunction recovery in rats after spinal cord contusion

Objective To explore the effect of olfactory ensheathing cell （OEC） transplantation combined with walking training on neurofunction recovery in rats after spinal cord contusion. Methods Forty adult female rats aged （75 ± 1 ） days were subjected to experimental spinal cord contusion at the T10 level using a New York University impactor at a height of 25 mm. They were then divided into 4 groups： （ 1 ） an OEC transplantation combined with walking training （OEC-walking training） group, （2） an OEC transplantation （OEC） group, （3） a walking training combined with Dulbecco＇s modified Eagle medium injection （DMEM） （walking training-DMEM） group, and （4） aDMEM injection （SCI-DMEM） group. The OEC transplants and DMEM injections were performed 2 weeks post-injury. Walking training began at the 7th day post-injury and consisted of daily sessions （once daily, 5 days a week for 10 weeks） of quadrupedal treadmill training, starting from 15 min and gradually increasing to 30 min daily, at speeds starting from 3 m/min and gradually increasing in accordance to the condition of the rats. Locomotor function recovery of the rats＇ hindlimbs was evaluated weekly using the Basso, Beattie and Bresnahan （BBB） locomotor rating scale.The expression of tyrosine hydroxylase （ TH ） was detected in the injured region of the lumbar spinal cord. Results The BBB scores of rats in the OEC-walking training group and the walking training-DMEM group improved significantly from the 4th week post-injury compared to the SCI-DMEM injection group. Rats in the OEC transplantation group had a significant improvement in BBB scores at the 5th to 8th weeks post-injury. At the end of the 11th week post-injury, the average BBB scores were 13.14 ± 0.24 in the OEC-walking training group, 11. 64 ± 0.56 in the OEC transplantation group, 12.29 ±0.64 in the walking training-DMEM group and 11.07 ± 0.84 in the SCI-DMEM group.The OEC-walking training group scored significantly higher than the other 3 groups. Although the number of TH-positive neurons in the lumbar spinal cord was not significantly different among the groups, the morphology of TH-positiveneurons in the OEC-walking training group and the walking training-DMEM group was different from those in the OEC transplantation group and the SCI-DMEM group. Conclusions OEC transplantation combined with walking training can effectively promote the functional recovery of the hindlimb. The plasticity of the descending TH system and of motoneurons of the ventral horn of the lumbar spinal cord might mediate the changes. Key words: Spinal cord contusion; Olfactory ensheathing cells transplantation; Walking training; Neurofunction; Tyrosine hydroxylase

Effect of different programs of hyperbaric oxygen preconditioning on spinal cord ischemia- reperfusion injury in rabbits

Objective To investigate the effect of different programs of preconditioning with hyperbaric oxygen (HBO) on spinal cord ischemia-reperfusion injury (I/R) in rabbits. Methods Forty-five New Zealand rabbits aged 4-5 months weighing 2.0-2.5 kg were randomly divided into 5 groups: group S, sham operation ( n = 5);group IR, spinal cord I/R injury (n = 10);group H_(1～3) , the animals were pretreated with 100% O_2 at 2.5 ATA 1 h/d for 5 (group H_1 ), 10 (group H_2 ) , or 20 (group H_3 ) consecutive days respectively 24 h before spinal cord I/R. The animals were anesthetized with iv pentobarbital sodium 30 mg/kg. The artery in the ear and left femoral artery were cannulated for proximal and distal mean blood pressure monitoring. Spinal cord ischemia was produced by cross-clamping of abdominal aorta distal to renal artery for 20 min. Hind-limb motor function was assessed at 48 h after reperfusion according to the modified criteria established by Tarlov (0 = no spontaneous movement, 4= normal motor function) . The animals were then killed and the L_5 segment of the spinal cord was removed for detection of neuronal survival (by HE staining), apoptosis (by TUNEL) and degeneration (by Fluoro-Jade B staining). Results Preconditioning with 5 or 10 d of HBO improved the hind-limb motor function and preserved more normal neurons in the spinal cord after I/R injury. Both apoptotic and degenerative cell death were attenuated in H_1 and H_2 groups. There was no significant difference in hind-limb motor dysfunction and the number of normal neurons in the lumbar spinal cord between H_3 group and I/R group. Conclusion Preconditioning with 5 d or 10 d HBO induces tolerance against spinal cord I/R injury, whereas preconditioning with 20 d of HBO fails to protect the spinal cord from I/R injury. Key words: Hyperbaric oxygenation; Ischemic preconditioning; Reperfusion injury; Spinal cord

Androgen Regulates the Sexually Dimorphic Gastrin-Releasing Peptide System in the Lumbar Spinal Cord that Mediates Male Sexual Function

A collection of neurons in the upper lumbar spinal cord of male rats projects to the lower lumbar spinal cord, releasing gastrin-releasing peptide (GRP) onto somatic and autonomic centers known to regulate male sexual reflexes such as erection and ejaculation. Because these reflexes are androgen dependent, we asked whether manipulating levels of androgen in adult rats would affect GRP expression in this spinal center. We found that castration resulted, 28 d later, in a profound decrease in the expression of GRP in the spinal cord, as reflected in immunocytochemistry and competitive ELISA for the protein as well as real-time quantitative PCR for the transcript. These effects were prevented if the castrates were treated with testosterone propionate. Genetically male (XY) rats with the dysfunctional testicular feminization allele for the androgen receptor (AR) displayed GRP mRNA and protein levels in the spinal cord similar to those of females, indicating that androgen normally maintains the system through AR. We saw no effect of castration or the testicular feminization allele on expression of the receptor for GRP in the spinal cord, but castration did reduce expression of AR transcripts within the spinal cord as revealed by real-time quantitative PCR and Western blots. Taken together, these results suggest that androgen signaling plays a pivotal role in the regulation of GRP expression in male lumbar spinal cord. A greater understanding of how androgen modulates the spinal GRP system might lead to new therapeutic approaches to male sexual dysfunction.

Neuroprotective effect of sonic hedgehog up‐regulated in Schwann cells following sciatic nerve injury

The physiological roles of sonic hedgehog (Shh) have been intensively characterized in development of various organs. However, their functions in adult tissues have not been fully elucidated. We investigated the expression and the potential function of Shh in crush-injured adult rat sciatic nerves. The Shh expression was up-regulated in Schwann cells adjacent to the injured site. The time-course analyses of various neurotrophic factors revealed the up-regulation of Shh mRNA followed by that of brain-derived neurotrophic factor (BDNF) mRNA. The continuous administration of cyclopamine, a hedgehog signal inhibitor, to the injured site suppressed the increase of BDNF expression and deteriorated the survival of motor neurons in lumbar spinal cord. Treatment of exogenous Shh in cultured Schwann cells enhanced the BDNF expression. The BDNF promoter activity (exon I and II) was increased in IMS32 cells co-transfected with Shh and its receptor Smoothened. These findings imply that the up-regulated expression of Shh in Schwann cells may play an important role in injured motor neurons through the induction of BDNF.

Training improves the electrophysiological properties of lumbar neurons and locomotion after thoracic spinal cord injury in rats

The aim of the present study was to evaluate the effect of a stepping-based rehabilitation program in voluntary wheel cages on the functional recovery and electrophysiological properties of neurons in the rat lumbar spinal cord after compressive thoracic (T10) spinal cord injury (SCI). A significant decrease in stance/swing duration and the number of limbs simultaneously in the stance phase was seen in trained compared to sedentary rats at 28 days after SCI ( p < 0.05). These kinematic improvements were associated with a significant increase in the amplitude of extracellular recordings from the tibial motoneuron pool in response to descending neuronal drive as well as significant amelioration of electrophysiological properties assessed from intracellular recordings. In fact, electrophysiological properties were not significantly different between uninjured controls and SCI-trained rats. Brain-derived neurotrophic factor (BDNF) levels were significantly elevated in the lumbar spinal cord of SCI-trained rats compared to SCI-sedentary controls. The data support a therapeutic role of increased neuromuscular activity in promoting functional recovery and suggest that it might occur via the beneficial effects of neurotrophic factors on neuronal plasticity.

Tight Glycemic Control by Insulin, Started in the Preischemic, but Not Postischemic, Period, Protects Against Ischemic Spinal Cord Injury in Rabbits

It is not well established whether insulin protects against ischemic spinal cord injury. We examined the effects of a single dose of insulin that corrects mild hyperglycemia on the outcome after transient spinal cord ischemia in rabbits. We assigned rabbits to four groups (n = 8 in each); untreated control (C) group, preischemic insulin (Pre-I) group, preischemic insulin with glucose (GI) group (glucose concentrations were maintained at levels similar to the C group by the administration of glucose), and postischemic insulin (Post-I) group. Insulin (0.5 IU/kg) was administered 30 min before ischemia in the Pre-I and GI groups, and just after reperfusion in the Post-I group. Spinal cord ischemia was produced by occluding the abdominal aorta for 13 min. Neurologic and histopathologic evaluations were performed 7 days after ischemia. The mean blood glucose concentration before ischemia in the Pre-I group (118 mg/dL) was significantly lower than in the other three groups (158-180 mg/dL) and those of 30 min after reperfusion in the Pre-I (92 mg/dL) and Post-I (100 mg/dL) groups were significantly lower than in the C (148 mg/dL) and GI (140 mg/dL) groups. The motor function score and number of normal neurons in the anterior lumbar spinal cord in the Pre-I group were significantly greater than in the other three groups. These results suggest that a relatively small dose of preischemic insulin protects against ischemic spinal cord injury, and that the protective effects result from tight glycemic control before ischemia.