Loss of host axons in nerve allografts after abolishing immunological tolerance in rats

Because host axons regenerate through long nerve allografts and become ensheathed by allogeneic cells in tolerant rats, we determined what would happen to these axons if tolerance were abolished. This was achieved by inoculating lymphoid cells from specifically sensitized isogenic donors into tolerant hosts. As a consequence of the immunological reaction which ensued, the host axons in the graft degenerated, indicating that permanent immunosuppression is required to maintain these fibers.     

The fate and functional capacity of neonatal neurons in allografts of ganglia in normal and immunologically tolerant rats

Previous studies have demonstrated that neurons in Ag-B histoincompatible allografts of adult ganglia are rejected unless the recipients have been rendered immunologically tolerant. Because neonatal grafts sometimes survive in situations where adult grafts of the same genotype are rejected, the fate and functional capacity of neurons in Ag-B histoincompatible allografts of neonatal ganglia was investigated. Sensory or sympathetic ganglia from neonatal Lewis rats were transplanted to the anterior chamber of the eye or into sternomastoid muscle of normal adult Brown Norway (BN) rats or to adult BN rats previously made immunologically tolerant to Lewis antigens. Normal BN rats rejected neonatal Lewis sensory ganglia (i.e., no neurons were present) at 70 days, whereas tolerant animals accepted these grafts for the 120 days they were followed. Furthermore, surviving Lewis neurons in tolerant BN rats were functional in that they regenerated nerve fibers into BN isografts of tongue and induced the development of taste buds. On the other hand, no neurons survived in neonatal sympathetic allografts to normal or tolerant BN rats and none or very few survived for even 2 weeks in sympathetic isografts. These results demonstrate that immunological factors (Ag-B antigens) are responsible for the ultimate loss of sensory neurons in neonatal allografts of ganglia whereas nonimmunological factors (i.e., transection of nerve fibers, separation from end-organ, nerve growth factor dependency) cause the loss of neurons in neonatal sympathetic ganglia. We conclude that both immunological and nonimmunological factors are important in determining the survival of neonatal neurons after transplantation.     

Survival of nerve and Schwann cells in allografts after cyclosporin A treatment

Because nerve and Schwann cells in allografts are rejected by normal rats, we investigated whether or not these neurological cells would survive if rats were treated with the new immunosuppressive drug, Cyclosporin A. Untreated rats rejected nerve and Schwann cells in allografts of ganglia or nerve. On the other hand, nerve and Schwann cells survived in allografts in Cyclosporin A-treated rats even after drug therapy was terminated. These results indicate that Cyclosporin A may be of value if allogenic nerve or Schwann cells are needed to aid in the repair of injured nerve tissue.     

Descending projection neurons to the spinal cord of the goldfish, Carassius auratus

The sources of descending spinal tracts in the goldfish, Carassius auratus, were visualized by retrograde transport of horseradish peroxidase (HRP) administered to the hemisected spinal cord. In the diencephalon, HRP-positive neurons were identified in the nucleus preopticus magnocellularis pars magnocellularis and ventromedial nucleus of the thalamus of the ipsilateral side. In the mesencephalic tegmentum, a few somata of the contralateral nucleus ruber and several ipsilateral neurons of the nucleus of the median longitudinal fasciculus were labeled. The reticular formation of the rhombencephalon was the major source of descending afferents to the spinal cord. A larger number of neurons were retrogradely labeled in the ipsilateral superior, middle, and inferior nuclei than in the contralateral nuclei. A few raphe neurons and the contralateral Mauthner neuron were also HRP-positive. The octaval area showed retrogradely labeled neurons in the anterior, magnocellular, descending, and posterior octaval nuclei of the ipsilateral side. A large number of neurons in the facial lobe and a few somata located adjacent to the descending trigeminal tract were labeled on the ipsilateral side. The pattern of descending spinal projections in goldfish is comparable to that of tetrapods and suggests that the spinal tracts have originated quite early in the course of vertebrate evolution.     

Self-mutilation after dorsal rhizotomy in rats: Effects of prior pain and pattern of root lesions

Self-mutilation of a forelimb often occurs in rats after the dorsal roots from the limb are transected. The present study investigated some of the conditions that affect the onset of chewing of the paw. It was found that painful irritation of the forepaw for 1 h prior to root transection significantly accelerated the onset of chewing and shifted the site of origin of chewing to the more distal portions of the paw. This observation suggests that the painful irritation triggers central neural activity characteristic of normal pain processing, and that deafferentation interrupts the input to brain stem inhibitory regions which would ordinarily suppress the pain-related activity. This “pain memory” may thus continue unchecked, thereby accelerating the onset of the abnormal central neural activity which triggers biting or chewing responses. The surprising observation that painful irritation of the paw after rhizotomy also accelerated the onset of chewing suggests that nonneural factors may contribute to the abnormal neural activity that underlies self-mutilation. The effects of different configurations of dorsal root transection on self-mutilation are described and discussed.     

Estimate of the total number of neurons and glial and endothelial cells in the rat spinal cord by means of the optical disector

The total numbers of neurons and glial and endothelial cells in five rat spinal cords were estimated by stereological techniques. Each spinal cord was divided into 12 slabs of equal length. One transverse and one oblique slice was cut from each slab. The volumes of gray and white matter of each cord were then estimated by point-counting techniques on the transverse slices. By means of optical disectors and systematic sampling, the numerical densities of different cell types were estimated on 35 m?-thick plastic sections from the oblique slices. The total cell number was calculated by multiplying the numerical density by the total volume of gray and white matter. On average there were 15.1 and 21.1 million cells in white and gray matter, respectively. Of the cells in gray matter, 6.4 million were judged to be neurons, 4.3 million to be endothelial, and 10.3 million to be glial. Of the neurons, 1.7 million were located in the cervical region, 2.5 million in the thoracic, 1.6 million in the lumbar, and 0.6 million in the sacro-coccygeal region. The methods used are simple to perform, and the counting necessary to obtain a reliable estimate of cell number from one spinal cord can be carried out during the course of 1 day. The only major problem is reliable criteria for unambiguous cell classification. © 1993 Wiley-Liss, Inc.     

Somatostatin-like neurons are expressed in fetal neocortical homografts in adult rat spinal cord

Fetal central nervous system homografts to adult spinal cord are considered a potential aid for recovery of function after paraplegia. This study utilizes somatostatin (SOM) immunohistochemistry to study the organization of an embryonic day 14 (E14) neocortical homograft into the spinal cord of an adult host over 6 postoperative months. Although the E14 homograft does not contain SOM-positive cells, SOM-reactive neurons are expressed by 30 days postimplantation and are still present in 6-month-old homografts. SOM-immunoreactive neurons are bitufted or multipolar and have dendrites that are confined to the graft. The homograft contains SOM-immunoreactive axons entering and/or exiting from lamina II in the host dorsal horn and SOM-positive homografted neurons send axons into the host ventral columns. These data show that the SOM peptide neocortical phenotype is preserved in homografts to spinal cord but there is anatomical host-homograft integration.     

Isolation of myelinoid Marchi-positive bodies from normal rabbit spinal cord

Normal rabbit spinal cord was homogenized in sucrose and fractionated by centrifugation in a sucrose density gradient system slightly modified after the Norton-Poduslo method for the isolation of myelin. The following fractions were recovered: the fraction floating on 0.32 M sucrose, the myelin fraction at the 0.32 M/0.85 M interface and the pellet. After fixation in glutaraldehyde the fractions were subjected to Marchi staining, a histochemical method used for the demonstration of degenerating myelin. The floating fraction was enriched in Marchi-positive bodies as compared to the homogenate while the myelin fraction and the pellet contained low amounts. No esterified cholesterol was found in the floating fraction. Since histochemical and electron microscopical studies have shown that Marchi-positive myelinoid bodies in the normal CNS are associated with node-paranode regions our results indicate a possibility to isolate and biochemically characterize a presumably closely myelin-related fraction of known anatomical origin. The absence of esterified cholesterol in the floating fraction shows that biochemical or biophysical properties other than a content of esterified cholesterol may give rise to a positive Marchi reaction.     

Evidence of normal mitosis with complete cytokinesis in central nervous system neurons during sustained depolarization with ouabain

Ouabain-induced depolarization of culture-matured spinal cord neurons from chick embryos causes the activation of DNA synthesis with subsequent mitotic nuclear division in a large fraction of the cells. Many of the activated neurons do not complete cytokinesis, and the binucleate cells thus formed have served as a convenient index for assaying the effectiveness of mitogenesis induction. However, the fact that the frequency of neurons induced to initiate DNA synthesis is consistently much larger than the frequency of binucleate neurons ultimately observed suggested the possibility that the difference may represent activated neurons that do complete cytokinesis, but go undetected in the binucleate assay cultures. This possibility was explored in the present study by determining the respective frequencies of induced neurons entering mitotic metaphase and completing mitotic telophase, and comparing them with the frequency of binucleate neurons ultimately formed. The results provide further evidence that normal mitogenic and mitotic processes are induced in central nervous system neurons by the depolarization treatment, and indicate that between 40 and 60% of the neurons which complete telophase subsequently undergo complete cytokinesis.     

Relationship of NK3 receptor- immunoreactivity to subpopulations of neurons in rat spinal cord

The distribution of immunoreactivity to the neurokinin₃ receptor (NK3R) was examined in segments C7, T11-12, L1-2, and L4-6 of the rat spinal cord. NK3R immunoreactivity was visualized by using two antisera generated against sequences of amino acids contained in the C-terminal region of the NK3R. NK3R-immunoreactive cells were numerous in the substantia gelatinosa of all spinal segments examined as well as the dorsal commissural nucleus of spinal segments L1-2. Isolated, immunoreactive cells were scattered throughout other regions of the spinal cord. The relationship of NK3R-immunoreactivity with neurons was demonstrated by colocalization with microtubule associated protein 2-immunoreactivity in individual cells. Within neurons, NK3R-immunoreactivity was associated predominately with the plasma membrane of cell bodies and dendrites. Within the substantia gelatinosa, 86% of nitric oxide synthase (NOS)-immunoreactive neurons were also NK3R-immunoreactive. Although NOS-immunoreactive neurons were found throughout all other regions of the spinal cord in the segments examined, these were not NK3R-immunoreactive. When preganglionic sympathetic neurons in spinal segments T11-12 and L1-2 were visualized by intraperitoneal injection of Fluorogold, less than 1% of the Fluorogold-labeled neurons were also immunoreactive for NK3R. The large number of NK3R-immunoreactive neurons in the substantia gelatinosa suggests that some effects of tachykinins on somatosensation may be mediated by NK3R. J. Comp. Neurol. 381:439-448, 1997. © 1997 Wiley-Liss, Inc.     

srGAP3在正常成年大鼠脑内的表达

目的 探讨slit—robo GTP激活蛋白3(slit-robo GTP activated protein 3，srGAP3)在正常成年大鼠中枢神经系统中的分布。方法 应用免疫组织化学ABC法研究了srGAP3蛋白在正常成年大鼠脑内的分布和定位。结果 srGAP3蛋白在正常成年大鼠脑中分布非常广泛，在脑内各主要结构如大脑皮质、海马、杏仁核、丘脑和下丘脑的部分核团、中脑、脑桥、小脑及延髓等均有表达。srGAP3免疫反应阳性物质位于神经元的细胞核周围。结论 srGAP3蛋白在正常成年大鼠脑中有很广泛的表达，提示srGAP3蛋白在成年大鼠中枢神经系统的功能活动中可能也起重要作用。     

Immunohistochemical demonstration of serotonin neurons in autonomic regions of the rat spinal cord

The immunohistochemical distribution of serotonin neurons in normal and transected spinal cords of rats was examined. Intraspinal serotonin neurons were immunostained as far rostral and caudal as T3 and Co1, respectively. All serotonin neurons were located in lamina VII and X, and most were located in spinal autonomic areas. Both bipolar and multipolar neurons were observed with many of the neurons oriented longitudinally to the long axis of the cord. Spinal neurons immunostained for serotonin were visible with and without L-tryptophan and monoamine oxidase inhibitor pretreatment.     

The fate of cryopreserved nerve isografts and allografts in normal and immunosuppressed rats

Donor Schwann cells, perineurial cells, and vasculature are known to survive in grafts of peripheral nerve. In the present study, we attempted to cryopreserve nerve to determine whether these cellular components of nerve would survive after transplantation and support host axonal regeneration through the graft. Four-centimeter lengths of peroneal nerves were removed from inbred adult American Cancer Institute (ACI) rats and placed into vials that contained a cryoprotective mixture of dimethyl sulfoxide and formamide (DF) at room temperature. Each vial with nerves in DF was cooled at a rate of 1–1.5°C/minute down to –40°C at which point the vials were plunged into liquid nitrogen at –196°C. After 5 weeks of storage, the nerves were thawed and DF removed. Some of the cryopreserved-thawed ACI nerves were transplanted as isografts into the legs of ACI rats. Other ACI nerves were used as allografts and inserted into immunologically normal Fischer (FR) rats that were untreated or were immunosuppressed with the drug Cyclosporin A (Cy-A). At surgery, only one end of the nerve graft was joined to the cut proximal end of the peroneal nerve of the host. The cellular elements of ACI grafts were present at 5 weeks in grafts removed from ACI rats and FR rats treated with Cy-A. Non-immunosuppressed FR rats rejected ACI nerves as did FR rats in whom Cy-A was stopped after 5 weeks of treatment. All surviving ACI grafts underwent Wallerian degeneration and consisted of columns of Schwann cells, which in their proximal portion were associated with regenerating host axons. The donor perineurial sheath and vasculature were also present in surviving grafts. ACI isografts only were examined 20 weeks postoperatively. All normal tissue components survived in these older grafts and contained regenerated and myelinated host axons throughout their 4 cm lengths. These results demonstrated that the cellular elements of nerve can be cryopreserved, and after transplantation, survive and function. Because nerves survived after prolonged cryopreservation, it seems feasible to establish a nerve bank from which grafts can be withdrawn to repair gaps in injured nerves. However, cryopreserved nerves used as allografts remain immunogenic and require immunosuppression for their survival. Published in 1993 by Wiley-Liss, Inc.     

Bombesin-induced hypothermia in rats tested at normal ambient temperatures: Contribution of the sympathetic nervous system

Rats infused centrally with bombesin become hypothermic at normal ambient temperatures when acutely deprived of food, but not while allowed unrestrained access to food. Ad lib-fed rats, tested at normal ambient temperatures, become hypothermic after receiving intracerebroventricular (ICV) bombesin when they have ventromedial hypothalamic lesions or when administered insulin or 2-deoxy-d-glucose peripherally. All of these conditions have been linked to reductions of sympathetic nervous system activity to brown adipose tissue (BAT), a major thermogenic mechanism of many homeotherms. A between group design was used to examine the effects of ICV bombesin infusions on the response to peripheral injections of a) the sympathetic ganglionic blocker chlorisondamine (2.5 mg/kg, IP) in ad lib-fed rats, b) the nonspecific β-agonist isoproterenol (30 mg/kg, IP) in food-deprived rats, and c) the combination of isoproterenol and chlorisondamine in ad lib-fed rats. Ad libfed rats receiving ICV bombesin (100 ng/5 μl), in combination with peripheral chlorisondamine injection, became hypothermic 60 min postbombesin administration (−2.84 ± 0.33°C), while ad lib-fed rats receiving ICV bombesin infusion and peripheral injections of saline did not (−0.08 ± 0.37°C). Isoproterenol blocked hypothermia in ad lib-fed rats injected with chlorisondamine and ICV bombesin. Food-deprived rats receiving ICV bombesin infusion and peripheral saline injection exhibited hypothermia 60 min postbombesin administration (−2.51 ± 0.29°C). Peripheral injections of isoproterenol prevented bombesin-induced hypothermia in food-deprived rats. These data suggest that bombesin induces hypothermia at normal ambient temperatures when the sympathetic nervous system drive to BAT cannot be (or is not) activated.     

Distribution of bradykinin B2 receptors in sheep brain and spinal cord visualized by in vitro autoradiography

Bradykinin B₂ receptors were localized in the sheep brain and spinal cord by quantitative in vitro autoradiography using a radiolabelled and specific bradykinin B₂ receptor antagonist analogue, 3-4-hydroxyphenyl-propionyl-D-Arg⁰-[Hyp³,Thi⁵,D-Tic⁷,Oic⁸]bradykinin, (HPP-HOE140). This radioligand displays high affinity and specificity for bradykinin B₂ receptors. The respective K_i values of 0.32, 1.37 and 156 nM were obtained for bradykinin, HOE140 and D-Arg[Hyp³,D-Phe⁷,Leu⁸]bradykinin competing for radioligand binding to lamina II of sheep spinal cord sections. Using this radioligand, we have demonstrated the distribution of bradykinin B₂ receptors in many brain regions which have not been previously reported. The highest density of bradykinin B₂ receptors occur in the pleoglial periaqueductal gray, oculomotor and trochlear nuclei and the circumventricular organs. Moderate densities of receptors occur in the substantia nigra, particularly the reticular part, the posterior thalamic and subthalamic nuclei, zona incerta, the red and pontine nuclei, some of the pretectal nuclei and in discrete layers of the superior colliculus. In the hindbrain, moderate levels of bradykinin B₂ receptor binding occur in the nucleus of the solitary tract, and in spinal trigeminal, inferior olivary, cuneate and vestibular nuclei. Laminae II, X and dorsal root ganglia display the most striking binding densities in the spinal cord, while the remainder of the dorsal and ventral horn display a low and diffuse density of binding. Bradykinin B₂ receptors are extensively distributed throughout the sheep brain and spinal cord, not only to sensory areas but also to areas involved in motor activity. J. Comp. neurol. 381:203-218, 1997. © 1997 Wiley-Liss, Inc.     

Autoradiographic distribution of [3H]YM-09151-2, a high-affinity and selective antagonist ligand for the dopamine D2 receptor group,in the rat brain and spinal cord

We determined the regional distribution of the dopamine D₂ receptor group in the rat central nervous system by quantitative receptor autoradiography with a high-affinity and selective antagonist, [³H]YM-09151-2. Saturation and competition experiments demonstrated that the binding of [³H]YM-09151-2 to striatal sections was saturable (Bmax=37.3 fmol/section), of high affinity (Kd=0.315 nM), and was inhibited selectively by prototypic D₂ ligands. The anatomical localization of binding sites was determined by comparison of autoradiograms and the original ³H-ligand-exposed sections stained with cresyl violet. Very high levels of [³H] YM-09151-2 binding were found in the caudate-putamen, nucleus accumbens, tuberculum olfactorium and the insula of Calleja, to each of which midbrain dopaminergic neurons project densely. High levels of binding were also observed in other regiions rich in dopaminergic neurons and fibers including the glomerular layer of the olfactory bulb, the intermediate lobe of the pituitary, lateral septum, substantia nigra pars compacta, interfascicular nucleus, dorsal raphe nucleus, locus coeruleus, and nucleus of the solitary tract. Some regions poor in dopaminergi innervation, however, had high levels of [³H]YM-09151-2 binding including the molecular layer of gyrus dentatus, all layers of CA1 and the nonpyramidal layer of CA4 of hippocampus, and the deeper layer of medial entorhinal cortex. Motor neurons present in brainstem motor nuclei and spinal ventral horn were also strongly labeled. Neocortical, cerebellar, and thalamic regions had low levels of binding, except lobules 9–10 of the cerebellum, the olivary pretectal nucleus, zona incerta and lateral mammillary nucleus, in which moderate to high levels of binding were detected. Our findings concerning the widespread but region-specific localization of [³]YM-09151-2 binding sites in the brain and spinal cord may prove useful for analyzing varoius dopaminergic functions in the central nervous system. © 1994 Wiley-Liss, Inc.     

脊髓髓内海绵状血管瘤的显微外科治疗及预后

目的 探讨脊髓髓内海绵状血管瘤(ISCC)的临床过程、影像学特点、治疗策略.方法 对宣武医院神经外科1993年5月至2007年11月手术治疗的81例ISCC资料进行回顾性分析.结果 ISCC占同期脊髓血管病的8.7%;男:女=1.6:1;平均发病年龄为35.7岁;病灶位于胸段56例(69%),颈段23例(28%),腰段2例(3%);64例患者获得长期随访,平均随访48个月,神经功能改善23例,未变化35例,恶化6例.术后复发4例.结论 (1)ISCC好发于青壮年,胸段多见,颈段次之;(2)大部分ISCC在MRI有特征性的表现;(3)对症状明显、病灶位于脊髓背侧、非出血急性期的ISCC提倡显微手术治疗.     

Immunohistochemical identification of neurons in ganglia of the guinea pig sphincter of oddi

The sphincter of Oddi is a smooth muscle sphincter that regulates the flow of bile into the duodenum. To identify potential chemical coding in sphincter of Oddi neurons, immunohistochemistry and histochemistry were employed to assay for putative neurotransmitters and related synthetic enzymes in wholemount preparations, with and without colchicine treatment. Immunoreactivities for enkephalin-endorphin (ENK-END), substance P (SP), nitric oxide synthase, vasoactive intestinal peptide (VIP), neuropeptide Y (NPY), and calcitonin generelated peptide (CGRP) were demonstrated within the ganglionated plexus. Roughly half of the neurons in the sphincter of Oddi expressed immunoreactivity for both SP and ENK-END, but not for nitric oxide synthase. About 25% of the neurons expressed nitric oxide synthase immunoreactivity as well as NADPH-diaphorase activity. This contingent of neurons was made up of two subgroups: one that expressed immunoreactivity for VIP, the other for NPY. Neurons that expressed CGRP immunoreactivity were sparse in sphincter of Oddi ganglia; however, many axons immunoreactive for both CGRP and SP were present in the ganglionated plexus. The CGRP/SP fibers are probably visceral afferents that may influence ganglionic output through axon reflex circuits. These results, along with studies of the actions of these neuroactive compounds on sphincter tone, support the view that ganglia of the sphincter of Oddi are largely comprised of excitatory (SP/ENK-END-immunoreactive) and inhibitory (nitric oxide synthase/VIP- or NPY-immunoreactive) neurons, and that sphincter of Oddi tone is controlled by the regulation of the outputs of these two groups of cells. © 1995 Wiley-Liss, Inc.     

Preoptic area unit activity during sleep and wakefulness in the cat

The spontaneous discharge of 86 preoptic area (POA) neurons was recorded extracellularly in chronically prepared cats during wakefulness (W), slow-wave sleep (SWS), and REM sleep. Of these, the percentage of units exhibiting maximal discharge rates in SWS and REM sleep (84%) was significantly greater than that of those exhibiting a maximal discharge rate in W (16%). Furthermore, those neurons that discharged rapidly in sleep (fast units) generally had a reduced discharge rate in W. Sixteen of the 86 units showed a strong tendency to discharge in bursts during SWS but not during W or REM sleep. The mean coefficient of variation and the mean discharge rate for these bursting cells in SWS were significantly greater than the corresponding values for the same cells in W and REM sleep, and for the nonbursting cells in SWS. Because POA stimulation is known to initiate behavioral and electrocortical signs of sleep, it is suggested that "fast units" in SWS with reduced discharge rates in W, may be "hypnogenic" cells.     

Quantification of myelin basic protein in the human fetal spinal cord during the midtrimester of gestation

The amount of myelin basic protein (MBP) was quantified in human fetal spinal cords from 12 to 24 gestational weeks (GW). MBP expression was determined by Northern blot, quantitative immunoblot, and immunocytochemistry. The development of compact myelin was analyzed by electron microscopy. Thirty-eight human fetal spinal cords were obtained after elective termination of intrauterine pregnancies from healthy women. Northern blot analysis showed a 15.8-fold increase in MBP mRNA between 12 and 18 GW. From 18 to 24 GW, MBP mRNA increased by 2.2-fold. The mRNA data paralleled immunoblot results that showed a 90.5-fold increase in MBP (0.147 ng/mg to 13.3 ng/mg tissue) between 12 and 18 GW and an approximately 11.5-fold increase between 18 and 24 GW (13.3 ng/mg to 154 ng/mg tissue). Immunocytochemical analysis also showed increased staining for MBP with advancing gestational age. At 12 GW, MBP immunoreactivity was observed in all three spinal cord funiculi. By 18 GW, MBP was expressed throughout the spinal cord white matter with the exception of the lateral corticospinal tracts and in the rostral levels of the fasciculus gracilis. With respect to myelin, at 12 GW, rare, noncompacted myelin lamellae were observed by electron microscopy. By 18 GW, discrete areas of compact myelin were observed in areas that showed MBP immunoreactivity, and at 24 GW, compact myelin was prominent throughout the white matter of the spinal cord. This study demonstrates a quantitative increase in MBP expression that is associated with myelin formation during the second trimester of human gestation. This information may provide normative data that can aid in the diagnosis of myelin disorders of the preterm, neonatal, and pediatric spinal cord. © 1996 Wiley-Liss, Inc.