Three-dimensional topographic analysis of spinal accessory motoneurons under chronic mechanical compression: an experimental study in the mouse

We investigated the effect of chronic mechanical compression of the cervical spinal cord on the number of spinal accessory motoneurons in 25 tiptoe-walking Yoshimura mice. The animals had calcified deposits in the atlantoaxial membrane at the C1–C2 vertebral level, compressing the spinal cord posterolaterally. Motoneurons of the spinal accessory nerve between C1 and C5 segments were labelled using wheat germ agglutinin-horseradish peroxidase (WGA-HRP) injected into the sternocleidomastoid muscles. The counted cells were processed into a three-dimensional computer display to analyse the cytoarchitectonic changes caused by external cord compression. The number of WGA-HRP-labelled spinal accessory motoneurons was significantly reduced on the affected side. The number of motoneurons in compromised C2 and C3 cord segments correlated linearly with the extent of mechanical compression, but no such relationship was present on the contralateral side. There was an increase in the number of WGA-HRP-labelled spinal accessory motoneurons in the medial cell pools of the anterior grey horn at a level most rostral to the compression, and in the ventrolateral cell pools at levels immediately rostral to the compression. Our findings suggest that the spinal accessory motoneurons translocate rostral to the area of external compression in order to avoid mechanical injury. Received: 12 March 1996 Received in revised form: 18 November 1996 Accepted: 8 December 1996     

Histological investigation of spinal cord lesions in the spinal hyperostotic mouse (twy/twy): morphological changes in anterior horn cells and immunoreactivity to neurotropic factors

We examined the morphology of spinal accessory motoneurons and immunoreactivity to neurotrophins, brain-derived neurotropic factor (BDNF) and neurotrophin (NT)-3, as well as the presence of reactive astrocytosis in 70 tiptoe walking Yoshimura (twy) mice that develop calcification at C1-C2 vertebral level compressing the spinal cord. At the level of compression, the area of neuronal soma and total length of dendrites of wheat germ agglutinin-horseradish peroxidase (WGA-HRP)-labelled accessory motoneurons in the medial cell pool decreased significantly with decrement in motoneuron population, relative to the control. In contrast, at sites rostral to the compressive lesion, a significant enlargement of the neuron soma and dendritic elongation were noted, associated with increased motoneuron population and decreased transverse area of the cord at the level of compression. At this site, enhanced BDNF and NT-3 immunoreactivities were evident in the anterior horn cells. In mice with a more severe degree of compression, astrocyte-like cells showing BDNF immunoreactivity became abundant and axons in the anterior column demonstrated a marked NT-3 immunoreactivity. Our results suggest increased functional activity of anterior horn cells at levels rostral to the site of compression. We speculate that the presence of BDNF and NT-3 in neurons and astrocyte-like cells is proportionate to the severity of chronic mechanical compression and may contribute to the heterotropic neuronal reserve and survival. Received: 18 August 1997ƒReceived in revised form: 17 February 1998ƒAccepted: 6 March 1998     

Eosinophilic granuloma of C4 causing spinal cord compression

A rare case of vertebral eosinophilic granuloma (C4) causing spinal cord compression is reported. The clinical, histological and radiological features of this pathological entity are discussed. After surgery a complete neurological recovery was observed. The value, in selected cases, of surgical treatment with total removal of the tumour and reconstruction of the spine to ensure spinal stability and to prevent irreversible neurological deficit is emphasized.     

Progesterone restores retrograde labeling of cervical motoneurons in Wobbler mouse motoneuron disease

The Wobbler mouse, a mutant characterized by motoneuron degeneration in the cervical spinal cord, has been used to test the efficacy of novel treatments for human motoneuron diseases (HMD). Previous reports have shown that slow axonal transport is impaired in Wobblers and other models of HMD. Since progesterone (PROG) corrects some morphological, molecular, and functional abnormalities of Wobbler mice, we studied if steroid exposure for 8 weeks restored retrograde axonal transport by measuring motoneuron labeling after injection of fluorogold into the limb muscles. The dye was injected into forelimb biceps bracchii and flexor or into the rearlimb gastrocnemius muscles; 6 days later, the number of fluorescent motoneurons and the total number of cresyl violet stained motoneurons were counted in the cervical (C5-T1) or lumbar (L3-L5) spinal cord regions. A pronounced reduction (- 42.2%) of the percent of fluorescent motoneurons in Wobbler mice cervical cord was noted, which was significantly corrected after PROG treatment. In contrast, labeling of lumbar motoneurons was not reduced in Wobbler mice and was not affected by PROG treatment. In no case PROG showed an effect in control mice. Concomitantly, PROG slightly but significantly increased biceps weight of Wobbler mice. Behaviorally, PROG-treated Wobblers performed better on a motor test (hanging time from a horizontal rope) compared to untreated counterparts. We postulate a dual role for PROG in the Wobbler mouse, in part by prevention of motoneuron degeneration and also by enhancement of axonal transport. The latter mechanism could improve the traffic of neurotrophic factors from the forelimb muscles into the ailing motoneurons, improving neuromuscular function in this murine model of HMD.     

Motor neuron destruction in guinea pigs immunized with bovine spinal cord ventral horn homogenate: experimental autoimmune gray matter disease

Guinea pigs immunized with bovine spinal cord ventral horn homogenate develop muscle weakness with electromyographic and morphologic evidence of denervation. Pathological examination demonstrates a loss of motoneurons and scattered inflammatory foci primarily localized to the spinal cord. Immunohistochemical techniques document the presence of immunoglobulin G at the motor end plate and around the external membrane and within the cytoplasm of motoneurons. This syndrome of experimental autoimmune gray matter disease (EAGMD) differs from experimental autoimmune motor neuron disease induced by inoculation with purified motoneurons and also differs from experimental autoimmune encephalomyelitis. The existence of two different forms of immune-mediated motoneuron destruction suggests that a number of cytoplasmic and membrane antigens may give rise to an immunologically based attack on the motor system.     

引起脊髓压迫的脊椎转移瘤的手术治疗体会

目的总结引起脊髓压迫的脊椎转移瘤的临床特点及治疗经验。方法回顾性分析手术治疗的8例引起脊髓压迫的脊椎转移瘤的临床资料。8例病人均行椎板减压及椎管内肿瘤切除术，手术中充分切除椎板，硬膜一般不打开，用取瘤钳咬除侧方及前方的硬膜外肿瘤，使脊髓充分得到减压。对神经根受累或椎体转移灶压迫神经根导致的严重根性疼痛，可切断一侧神经根，术后进行放疗等综合治疗。结果8例均经手术治疗，无手术死亡，随访1—19个月。1例术后4个月因肝癌死亡，其余均存活至今。手术前后神经功能评价采用Frankel分级，术前4例A级的1例无变化，3例恢复到C级；术前2例B级恢复到D级；术前2例C级恢复勉强达E级。所有疼痛均消失，二便功能均有改善。结论部分引起脊髓压迫的脊椎转移瘤经后正中入路肿瘤部分切除、椎板减压治疗可以取得良好的治疗效果。     

Introduction of the MASH1 gene into mouse embryonic stem cells leads to differentiation of motoneuron precursors lacking Nogo receptor expression that can be applicable for transplantation to spinal cord injury

ES cells transfected with the MASH1 gene yielded purified spinal motoneuron precursors expressing HB9 and Islet1. The cells lacked the expression of Nogo receptor that was of great advantage for axon growth after transplantation to an injured spinal cord. After transplantation, mice with the complete transection of spinal cord exhibited excellent improvement of the motor functions. Electrophysiological assessment confirmed the quantitative recovery of motor-evoked potential in the transplanted spinal cord. In the grafted spinal cord, gliosis was inhibited and Nogo receptor expression was scarcely detected. The transplanted cells labeled with GFP showed extensive outgrowth of axons positive for neurofilament middle chain, connected to each other and expressed Synaptophysin, Lim1/2 and Islet1. Thus, the in vivo differentiation into mature spinal motoneurons and the reconstitution of neuronal pathways were suggested. The grafted cell population was purified for neurons and was free from teratoma development. These therapeutic strategies may contribute to a potent treatment for spinal cord injury in future.     

Quantitative autoradiographic distribution of glutamate receptors in the cervical segment of the spinal cord of the wobbler mouse

The reduction of glutamate content has been observed in the spinal cord of the wobbler mouse, a purported model of amyotrophic lateral sclerosis (ALS). To elucidate glutamate receptors in the wobbler spinal cord, we measured densities ofN-methyl-d-aspartate (NMDA), kainate, -amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) and metabotropic glutamate (mGlu) binding sites using in vitro autoradiography. In wobbler mice, NMDA, kainate, and AMPA binding sites were increased in the dorsal horn and kainate binding sites were also increased in the intermediate zone. However, mGlu binding was unchanged. These results disagree with those observed in ALS spinal cords, in which NMDA and kinate binding sites are decreased. The wobbler mouse may have the glutamate dysfunction, but in a different way from ALS.     

Methodological analysis of an experimental spinal cord compression model in the rat

A technique for producing graded spinal cord compression injuries in rats is described. A metal plate 2.2 x 5.0 mm in size is applied to the exposed spinal dura and loaded with weights. Neurological function is tested postoperatively on an inclined plane. Reading of the maximal angle of rat performance on this plane was found reproducible on comparison between different observers. Laminectomy per se had a minimal effect on the performance. Compression of 35 g for 5 min caused a pronounced but incomplete injury, with almost total recovery within 14 days and with no difference between animals on artificial respiration and those breathing spontaneously. Animals whose spine was fixed during the compression had a better outcome than those without such fixation.     

脊髓慢性压迫性损伤后细胞凋亡和BDNF表达的观察

目的 探讨大鼠脊髓在遭受到持续进行性压迫损伤后神经细胞凋亡以及脑源性神经营养因子（BDNF）及其TrkB受体的变化。方法 将75只SD大鼠分为模型组、对照组和正常组,各25只;根据造模后取材时间,各组再分为1、7、14、21、28 d 5亚组,每亚组5只。胸11~12椎板和硬脊膜之间置入缓膨材料（3 mm×5 mm,厚0.8 mm）制作大鼠慢性压迫性脊髓损伤模型,BBB评分评估行为学变化,TUNEL染色检测细胞凋亡,免疫组化染色检测BDNF及其TrkB受体表达变化。结果 造模后7、14、21、28 d,模型组大鼠BBB评分均明显低于对照组和正常组（P<0.05）,而对照组和正常组均无统计学差异（P>0.05）。模型组大鼠可观察到从脊髓受压开始,神经细胞开始出现凋亡,中央管及前角区域的神经细胞凋亡明显,邻近灰质的白质部分神经胶质细胞凋亡明显;而对照组和正常大鼠未见明显凋亡细胞。模型组大鼠脊髓内BDNF及其TrkB受体呈强阳性,尤其是神经元部位,BDNF及其受体TrkB表达明显,且主要表达在运动类神经元中,随压迫进行,表达逐渐增强,至相对稳定;对照组和正常组大鼠脊髓内BDNF及其TrkB受体表达较少。结论 大鼠脊髓在受到慢性压迫性损伤时,神经细胞凋亡明显,BDNF、TrkB受体表达明显增强。     

Thyrotropin-releasing hormone (TRH) neurons sprout in cervical spinal cord of Wobbler mouse

The present study was undertaken to quantify the immunocytochemical changes for thyrotropin-releasing hormone (TRH) within the ventral horn of the cervical spinal cord from Wobbler (wr / wr) mice selected at postnatal ages 3 weeks to 5 months compared with the normal phenotype (NFR/wr) littermates as well as mice from two related normal mouse strains: the NFR/N parent strain, and the closely related C57B1/6N mouse strain. The immunoreactive (IR) neuronal processes containing TRH appeared in all specimens within Rexed's laminae VIII, IX, and X. Compared with the normal (C57B1/6N, NFR/N) specimens, the pair-matched normal phenotype (NFR/wr) and Wobbler (wr / wr) specimens possessed significantly greater numbers of IR-TRH containing processes at every age studied. Compared with the normal phenotype (NFR/wr) specimens, greater numbers of IR-TRH containing processes appeared in the ventral horn region studied from the Wobbler (wr / wr) specimens taken early (Stage 1) as well as later (Stages 3 and 4) in the motoneuron disease. An age-related decline in the number of IR-TRH processes was apparent among the specimens from the Wobbler mouse strain (NFR/wr, wr / wr), but not the normal (NFR/N, C57B1/6N) mouse strains. The data suggest that TRH may play a significant role in the Wobbler disease, possibly even before the symptoms become apparent. In addition strain-related differences exist which may be important to the etiology of the Wobbler disorder.     

慢性渐进性压迫对大鼠行为功能及脊髓运动神经递质表达的影响

目的 :观察脊髓慢性受压后实验动物的行为功能与运动神经递质表达的变化情况。方法 :采用大鼠后路渐进性脊髓压迫动物模型 ,观察联合行为评分 (CBS) ,常规病理及免疫组化检测胆碱乙酰转移酶 (ChAT)的变化。结果 :免疫组织化学染色显示 ,在正常大鼠脊髓前角运动神经元及大小神经元ChAT均表达阳性 ,脊髓损伤后ChAT阳性细胞数减少 ,CBS升高 ,二者具有相关关系。结论 :脊髓受压抑制大鼠脊髓神经元合成ChAT ,从而影响实验动物的行为功能     

Activation of 5-HT1C/2 receptors depresses polysynaptic reflexes and excitatory amino acid-induced motoneuron responses in frog spinal cord

Sucrose gap recordings from the ventral roots of isolated, hemisected frog spinal cords were used to evaluate the effects of high concentrations of serotonin (5-HT) and alpha-methyl-5-HT (alpha-Me-5-HT) on the changes in motoneuron potential produced by dorsal root stimulation and by excitatory amino acids and agonists. Bath application of 5-HT in concentrations of 10 microM or greater produced a concentration-dependent motoneuron depolarization. Polysynaptic ventral root potentials evoked by dorsal root stimuli were reduced in both amplitude and area by 5-HT or alpha-Me-5-HT (both 100 microM). This may result from a reduction of the postsynaptic sensitivity of motoneurons to excitatory amino acid transmitters because 5-HT significantly depressed motoneuron depolarizations produced by addition of L-glutamate and L-aspartate to the superfusate. Similarly, 5-HT reduced depolarizations produced by the excitatory amino acid agonists N-methyl-D-aspartate (NMDA), quisqualate, alpha-amino-3-hydroxy-5-methyl-4-isoxazoleproprionic acid (AMPA), and kainate. alpha-Me-5-HT reduced NMDA depolarizations. Tetrodotoxin (TTX) did not affect the ability of 5-HT to attenuate NMDA or kainate depolarizations, but did eliminate the 5-HT-induced attenuation of quisqualate and AMPA depolarizations. The glycine receptor site associated with the NMDA receptor did not appear to be affected by 5-HT because saturation of the site by excess glycine did not alter the 5-HT-induced depression of NMDA responses. The 5-HT1C/2 antagonist ketanserin and the 5-HT1A/2 antagonist spiperone significantly attenuated the 5-HT-induced depression of NMDA-depolarizations.(ABSTRACT TRUNCATED AT 250 WORDS)     

Demyelination in the spinal cord of murine globoid cell leukodystrophy (the twitcher mouse)

Summary Chronologic events of demyelination were investigated in the spinal cord of the twicher mouse, an authentic murine model of human globoid cell leukodystrophy (GLD) from 5 to 45 days postnatal. There was very little evidence of myelin degeneration before day 25 although clustered or scattered globoid cells were already noted in the dorsal columns and intramedullary portion of the ventral roots.Globoid cells contained typical cytoplasmic inclusions and in those which were found adjacent to degenerating myelin and naked axons, myelin debris were conspicuous in their cytoplasm.Vesiculation of myelin and a feature of globoid cells stripping myelin lamellae were noted in the area of demyelination. Myelin and oligodendroglial degeneration became pronounced throughout the spinal white matter after day 40 but globoid cells tended to be more concentrated in the dorsal columns.Our observations suggest that the emergence of globoid cells in GLD is in response to the changes in biochemical environment (i.e., excessive presence of galactosylceramide in the tissue?), and these cells appear to have a role as phagocytic cells in removing myelin lamellae.Supported in part by research grants NS-03356, NS-10803, and HD-01799 and a training grant for experimental neuropathology, NS-07098, from NINCDS     

Hormone-sensitive periods for the control of motoneuron number and soma size in the dorsolateral nucleus of the rat spinal cord

The dorsolateral nucleus (DLN) of the rat lumbosacral spinal cord is sexually dimorphic, with males having more and larger DLN motoneurons than do females. The development of this dimorphism depends on the presence of perinatal androgens. The present study sought to determine the periods in development during which the DLN is sensitive to the masculinizing effects of the androgen testosterone propionate (TP). The size and number of DLN motoneurons in neonatally ovariectomized female rats that were exposed to TP during either the late prenatal, early postnatal, or late postnatal period were compared to control males and females. Both late prenatal and early postnatal TP injections significantly increased DLN number by 48% and 50%, respectively, but the sensitive period for TP masculinization of soma size seems to be primarily postnatal, because prenatal TP injections had little or no effect on that measure. The sensitive period for TP masculinization of DLN neuron number is similar to that of the sexually dimorphic spinal nucleus of the bulbocavernosus (SNB). However, the sensitive period for TP masculinization of DLN soma size appears to begin later than for the SNB.     

不同类型脊髓损伤患者和正常成人外周血白细胞糖皮质激素受体的比较及意义

目的 比较正常成人和急性脊髓损伤患者、慢性脊髓压迫症患者外周血白细胞塘皮质激素受体的结合位点数并探讨其意义。方法 采用放射配体结合法测定15例正常成人、20例急性脊髓损伤患者和21例慢性脊髓压迫症患者外周血白细胞上糖皮质激素受体结合位点数。结果 正常成人外周血白细胞精皮质激素受体结合位点数为4462±891.6个/细胞,慢性脊髓压迫症患者为4225±1271个/细胞,急性脊髓损伤患者为2517±857.8个/细胞,经统计学比较正常成人组和慢性脊髓压迫症患者组没有显著性差异,急性脊髓损伤组与其他两者相差均有显著性意义。急性脊髓损伤组中,全瘫患者为2279±921个/细胞,不全瘫患者为2806±718个/细胞,两者无统计学差异。结论 外周血白细胞上的糖皮质激素受体有高亲和力和低亲和力两种结合位点,急性脊髓损伤后外周血白细胞的精皮质激素受体结合位点数的减少主要是高亲和力位点的减少,低亲和力位点维持不变。大剂量的糖皮质激素和白细胞的低亲和力位点结合,抑制白细胞的趋向移动,减少白细胞进入损伤脊髓区,减轻损伤后的急性炎症反直,起到神经保护作用。慢性脊髓压迫症患者予以糖皮质激素治疗无疗效。     

Decreased immunoreactive (IR) calcitonin gene-related peptide correlates with sprouting of IR-peptidergic and serotonergic neuronal processes in spinal cord and brain nuclei from the Wobbler mouse during motoneurons disease

The Wobbler mouse possesses an inherited form of motoneuron disease that expresses itself most dramatically in the forelimbs. Previous immunocytochemical (ICC) studies have shown that neuronal processes containing substance P (SP), thyrotropin releasing hormone (TRH) and serotonin (5-HT) seem to sprout in the ventral horn of the cervical spinal cord taken from the Wobbler mouse. By radioimmunoassay, increased concentrations of spinal SP, TRH, and 5-HT, as well as leucine and methionine enkephalins (LE, ME) have been documented. The present ICC study quantifies the numbers of neuronal processes in the Wobbler cervical spinal cord and brainstem which contain SP, 5-HT, LE, ME and other neuropeptides (cholecystokinin, CCK; neuropeptide Y; galanin; calcitonin gene-related peptide, CGRP). It is proposed that those processes that sprout early in the mononeuron disease (5-HT, LE, ME, CCK and also TRH according to other studies) may be involved in the etiology. In addition, it is hypothesized that the loss of CGRP within the ventral horn may represent the loss of a trophic factor that is important to the survival motoneurons and may influence the increase of fiber densities around the dying motoneurons.     

Assessment of axonal dysfunction in an in vitro model of acute compressive injury to adult rat spinal cord axons

An in vitro model of spinal cord injury was developed to study the pathophysiology of posttraumatic axonal dysfunction. A 25 mm length of thoracic spinal cord was removed from the adult male rat (n = 27). A dorsal column segment was isolated and pinned in a recording chamber and superfused with oxygenated (95%O₂/5% CO₂) Ringer. The cord was stimulated with a bipolar electrode, while two point responses were recorded extracellularly. Injury was accomplished by compression with a modified aneurysm clip which applied a 2 g force for 15 s. With injury the compound action potential (CAP) amplitude decreased to 53.7 ± 5.4% (P < 0.001), while the latency increased to 115.6 ± 3.1% (P < 0.0025) of control values. The absolute refractory period increased with injury from 1.7 ± 0.1 ms to 2.1 ± 0.1 ms (P < 0.001). With train stimulation (200 and 400 Hz), injured axons showed evidence of high frequency conduction failure (P < 0.05). The infusion of 5 mM 4-aminopyridine (4-AP), a blocker of voltage-sensitive ‘fast’ K channels confined to internodal regions, resulted in broadening of the CAP of injured axons to 114.9 ± 3.1% of control (P < 0.05). Ultrastructural analysis of the injured dorsal column segments revealed marked axonal and myelin pathology, including considerable myelin disruption.In conclusion, we have developed and characterized an in vitro model of mammalian spinal cord injury which simulates many of the features of in vivo trauma. Injured axons display characteristic changes in physiological function including a shift in refractory period and high frequency conduction failure. The ultrastructural data and response of injured axons to 4-AP suggest that myelin disruption with exposure of ‘fast’ K⁺ channels contributes to posttraumatic axonal dysfunction.     

Permanent alterations in the rat spinal cord following prenatal exposure to N-ethyl-N-nitrosourea

Pregnant rats between gestational stages E14–E22 were given a single injection of N-ethyl-N-nitrosourea (ENU). Pups born of these females were sacrificed 60 days after birth and their spinal cords examined qualitatively and quantitatively. Quantitative analysis involved measurement of spinal cord length and volume, estimation of neuron number, and the measurement of individual cell dendritic number and length. Cytoarchitecturally spinal cords appeared normal in all animals regardless of the age when they were exposed to ENU. Animals exposed during the latter portion of neurogenesis in the spinal cord (E14–E16) had significantly (p < 0.05) reduced volumes of gray matter and reduced cell counts. Cellular analysis showed that all animals exhibited some stunting of dendritic length, although the number of dendritic branches was significantly (p < 0.01) higher than normal for neurons of the intermediate gray and the substantia gelatinosa. Increase in the number of dendrites per cell suggests a mechanism of structural compensation by the surviving neuronal cells following their exposure to the teratogen.     

Soma size and oxidative enzyme activity in normal and chronically stimulated motoneurones of the Cat''s spinal cord

In normal adult cats we measured the density of staining for the activity of succinate dehydrogenase (SDH staining) in ventral horn cells of different sizes. The measurements were restricted to that part of the lumbar ventral horn (L6-L7) which is known to contain motoneurones of the peroneal nerve. A statistically significant tendency was found for the SDH staining to be denser in smaller than in larger neurones within the size range of a motoneurones (soma diameter greater than 40 microns). These results are consistent with recently published evidence for ventral horn cells of rats and qualitatively similar relationships between size and SDH staining have also been observed among skeletal muscle fibres (confirmed for mixed muscle of cat in present study). In hindlimb muscles, size as well as SDH staining are known to be markedly activity-dependent. We tested whether this is the case for peroneal motoneurones as well by analyzing the effects of chronic nerve stimulation on the properties of neurones within the appropriate region of the ventral horn. Prior to the final acute experiment, these cats had been subjected to a left-side dorsal rhizotomy and hemispinalization. By aid of a portable mini-stimulator, the left-side common peroneal nerve was activated by repetitive pulses during 50% of total time per day (intra-activity rate: 10, 20 or 40 Hz). After 8 weeks of such treatment, cell sizes as well as the densities of SDH staining showed hardly any differences between peroneal ventral horn cells of the experimental and control sides of the spinal cord.(ABSTRACT TRUNCATED AT 250 WORDS)