An electron microscope study of glycine-like immunoreactivity in laminae I-III of the spinal dorsal horn of the rat

The ultrastructural distribution of glycine-like immunoreactivity in laminae I-III of rat spinal dorsal horn was examined by using pre-embedding immunocytochemistry. Immunoreactive axons, dendrites and cell bodies were observed in all three laminae, but were most common in lamina III. The axons were presynaptic at axodendritic and axosomatic synapses, but also at axo-axonic synapses in laminae II and III, where the postsynaptic boutons frequently resembled the terminals of myelinated primary afferents. Some vesicle-containing dendrites in lamina II also showed glycine-like immunoreactivity. Immunoreactive dendrites in laminae II and III were postsynaptic to the central axons of type II, but not type I glomeruli, which suggests that glycinergic neurons receive a major monosynaptic input from myelinated primary afferents. These results support the suggestion that GABA and glycine co-exist in some neurons in laminae I-III of rat dorsal horn, and confirm that glycine is involved in somatosensory processing involving low threshold myelinated cutaneous primary afferents.     

Catecholaminergic innervation of the spinal dorsal horn: a correlated light and electron microscopic analysis of tyrosine hydroxylase-immunoreactive fibres in the cat.

The ultrastructural organization of presumed catecholamine-containing boutons, in the dorsal horn of the cat lumbosacral spinal cord, was examined in an immunocytochemical study using an antiserum against tyrosine hydroxylase. The study was restricted to the first four laminae of Rexed. Light microscopic inspection revealed numerous, varicose, tyrosine hydroxylase-immunoreactive axons throughout this region of the spinal cord. Within laminae I and II the fibres exhibited a prominent rostrocaudal orientation, while in laminae III and IV they were organized predominantly dorsoventrally. Correlated ultrastructural analysis confirmed that these varicosities were synaptic boutons. Forty-five of these structures were examined through serial sections and they were found to form symmetrical (Gray type II) synaptic junctions with dendrites (95%) and somata (5%). Immunoreactive boutons were not observed to be either presynaptic or postsynaptic to axon terminals. These findings suggest that catecholamines within the spinal dorsal horn act through a postsynaptic action upon dorsal horn neurons.     

Morphology and synaptic relationships of physiologically identified low-threshold dorsal root axons stained with intra-axonal horseradish peroxidase in the cat and monkey

The arborizations and synaptic relationships of intra-axonally stained horseradish peroxidase- (HRP) labeled primary afferent fibers to the dorsal horn of the cat and monkey spinal cord have been studied by light and electron microscopic methods. The light microscopic arborizations of the afferent fiber types (hair follicle afferents, pacinian corpuscle afferents, type I and type II slowly adapting afferents) are similar to those described by Brown and his colleagues (1) in the cat. The synaptic profiles formed by labeled afferents contain rounded synaptic vesicles. In serial thin sections, it was found that single dorsal root axons may make hundreds or thousands of synapses with neuronal structures of the dorsal horn. The vast majority of synaptic contacts are on the dendritic trees of dorsal horn neurons. The synapses made by these low-threshold afferent axons are almost all in the deeper laminae (III-VI) of the dorsal horn. The hair follicle afferent axons and the pacinian corpuscle afferents have numerous vesicle-containing structures that synapse on them to form either axoaxonal synapses or dendroaxonal synapses. The slowly adapting afferent axons are less often found to be postsynaptic to axons or dendrites. It is concluded that different physiological classes of primary afferent axons have different morphological characteristics, both at the light and electron microscopic level.     

脊髓侧角区5-HT、TH、SP和L-ENK免疫反应纤维和终末的超微结构

本文应用免疫细胞化学ABC法,在电镜下观察脊髓侧角区单胺能和某些肽能纤维及末梢的突触组合。大鼠侧角内的5-HT、TH、SP和L-ENK免疫反应纤维均为无髓纤维。在侧角细胞簇内,这些纤维穿行于胞体之间,有的与胞体相邻,但很少与胞体形成轴-体突触。这些单胺和肽类纤维也与树突伴行,在树突束内数量最多。有时一小束无髓纤维都含同一种免疫反应物质。轴-树突触是各种免疫反应纤维终末所形成突触的主要形式。各种纤维终末所含的小泡多为圆清亮小泡,或兼有少数大颗粒泡。SP和L-ENK纤维膨体内的小泡与其终末内者不同,大颗粒泡较多,有时约占半数。各种免疫反应终末所组成参与的突触,对称或非对称型均不显现优势。     

Substance P-immunoreactive elements in laminae I and II of the chicken spinal cord: a light- and electron-microscopic study

Substance P (SP)-immunoreactive (IR) elements were studied in laminae I and II of the chicken spinal cord in conjunction with an anti-SP monoclonal antibody at light- and electron-microscopic levels by means of the indirect antibody peroxidase-antiperoxidase technique. At the light-microscopic level, SP-IR elements were most intensely observed in the dorsolateral portion of the dorsal horn, laminae I and II. Electron-microscopically, SP-IR boutons contained large spherical dense-cored vesicles (diameter range: 60-125 nm) and spherical clear vesicles. They were subdivided into two groups: large SP-IR boutons, which were the central terminals in synaptic glomeruli, and small SP-IR boutons. In the synaptic glomerulus, two kinds of non-IR presynaptic profiles made axo-axonic synapses with the SP-IR central terminal: one was the presynaptic profile containing pleomorphic clear vesicles and the other was the presynaptic profile containing large dense-cored vesicles. A 'septate junction'-like structure was observed between large SP-IR boutons in synaptic glomeruli. The present results suggest that SP-containing primary afferents are modulated presynaptically by two different neurotransmitter or modulator systems.     

Inhibitory zinc-enriched terminals in mouse spinal cord

The ultrastructural localization of zinc transporter-3, glutamate decarboxylase and zinc ions in zinc-enriched terminals in the mouse spinal cord was studied by zinc transporter-3 and glutamate decarboxylase immunohistochemistry and zinc selenium autometallography, respectively.The distribution of zinc selenium autometallographic silver grains, and zinc transporter-3 and glutamate decarboxylase immunohistochemical puncta in both ventral and dorsal horns as seen in the light microscope corresponded to their presence in the synaptic vesicles of zinc-enriched terminals at ultrastructural levels. The densest populations of zinc-enriched terminals were seen in dorsal horn laminae I, III and IV, whereas the deeper laminae V and VI contained fewer terminals. At ultrastructural levels, zinc-enriched terminals primarily formed symmetrical synapses on perikarya and dendrites. Only relatively few asymmetrical synapses were observed on zinc-enriched terminals. In general, the biggest zinc-enriched terminals contacted neuronal somata and large dendritic elements, while medium-sized and small terminals made contacts on small dendrites. The ventral horn was primarily populated by big and medium-sized zinc-enriched terminals, whereas the dorsal horn was dominated by medium-sized and small zinc-enriched terminals.The presence of boutons with flat synaptic vesicles with zinc ions and symmetric synaptic contacts suggests the presence of inhibitory zinc-enriched terminals in the mammalian spinal cord, and this was confirmed by the finding that zinc ions and glutamate decarboxylase are co-localized in these terminals. The pattern of zinc-enriched boutons in both dorsal and ventral horns is compatible with evidence suggesting that zinc may be involved in both sensory transmission and motor control.     

Distribution of thyrotropin-releasing hormone in the rat spinal cord with special reference to sympathetic nuclei: A light- and electron-microscopic immunocytochemical study

Summary This paper deals with the distribution of thyrotropin-releasing hormone-like immunoreactivity in the spinal cord of the rat, and particularly in the sympathetic nuclei, at light and electron microscopic levels. In the dorsal horn, the inner part of laminae II and III displayed thin thyrotropin-releasing hormone immunoreactive profiles. Electron microscopy revealed small immunoreactive varicosities which made synaptic contact with small dendrites or dendritic spines. Dense thyrotropin-releasing hormone-like immunoreactivity was observed in all sympathetic nuclei (nucleus intermediolateralis pars fascicularis and principalis, nucleus intercalatus and dorsal commissural nucleus) except the nucleus intercalatus pars ependymalis. Electron microscopy showed many immunoreactive varicosities which were often in synaptic contact with dendrites (proximal or distal), rarely with perikarya and never with axons. Sometimes, the same immunoreactive varicosity made axodendritic contacts with two dendrites and, conversely one dendrite was sometimes synaptically contacted by two or more immunoreactive varicosities. The ventral horn displayed a diffuse thyrotropin-releasing hormone-like immunoreactivity except for the cremaster nucleus (at lumbar level) which was densely outlined by immunoreactive profiles. Occasionally a large cell body in lamina IX (a putative motoneuron) was outlined by immunoreactive profiles but ultrastructural studies revealed very few immunoreactive axosomatic synapses, while immunoreactive symmetrical or asymmetrical axodendritic synapses were observed. The present study clearly confirms the existence of thyrotropin-releasing hormone immunoreactive synapses, thus substantiating the physiological role of this hormone in the spinal cord.     

Anatomical distribution and ultrastructural organization of the gabaergic system in the rat spinal cord. An immunocytochemical study using anti-GABA antibodies

γ-Aminobutyric acid (GABA)-containing elements have been studied by light and electron microscopy in the rat spinal cord, using immunocytochemistry with anti-GABA antibodies. Light microscopy showed immunoreactive somata localized principally in laminae I–III, and occasionally in the deeper laminae of the dorsal horn and in the ventral horn. Small somata were also observed around the central canal. Punctate GABA-immunoreactive profiles were particularly concentrated in laminae I–III, and moderately abundant in the deeper laminae and in the ventral horn where they were observed surrounding the unlabelled motoneurons.

At the ultrastructural level, the punctate profiles corresponded to GABA-containing axonal varicosities or small dendrites. GABA-immunoreactive varicosities were presynaptic to labelled or unlabelled dendrites and cell bodies. Some unlabelled terminals presynaptic to unlabelled dendrites received symmetrical synaptic contacts from GABA-immunoreactive terminals.

These results confirm data obtained withl-glutamate decar☐ylase immunocytochemistry, and support the role of GABA in pre- and postsynaptic inhibition in the spinal cord, respectively via axoaxonal and axosomatic or axodendritic synapses.     

Cholecystokinin-like immunoreactivity in the dorsal horn of the spinal cord of the rat: a light and electron microscopic study

Cholecystokinin-like immunoreactivity was investigated with an indirect immunoperoxidase technique in the whole spinal cord with the light microscope and in the dorsal horn with the electron microscope. Intraparenchymal injections of colchicine were performed to allow the detection of cholecystokinin-like immunoreactive cell bodies. Rats treated at birth with capsaicin were also studied at the light microscope. Numerous cholecystokinin-like immunoreactive fibres and varicosities were found in the two superficial layers of the dorsal horn and in the intermedio-medial nucleus; cholecystokinin-like immunoreactive cell bodies were also present in these two regions. After neonatal capsaicin treatment, the number of cholecystokinin-like immunoreactive fibres and varicosities was strongly reduced in the dorsal horn. At the electron microscope level, cholecystokinin-like immunoreactivity was localized in numerous neurites often filled with vesicles (axon terminals and dendrites containing vesicles) and in few cell bodies and dendrites. The immunoreaction was found mainly associated with ribosomes, granular reticulum, neurotubules and vesicles. Large granular vesicles were filled with the reaction product whereas small and medium-sized vesicles showed a varying degree of immunoprecipitate around their membrane. In addition dense "granules" of precipitate were observed in numerous presynaptic neurites. Cholecystokinin-like immunoreactive axons were of small calibre and mostly unmyelinated. Cholecystokinin-like immunoreactive axon terminals made asymmetric synaptic contacts with generally unlabelled dendrites or dendritic spines. A single labelled nerve terminal could contact several different dendrites in structures resembling glomeruli. Few axo-somatic synapses but a relatively high number of axo-axonic contacts were seen. About half of these axo-axonic contacts involved pre- and postsynaptic profiles. Both light and electron microscopic observations led us to the conclusion that some of the cholecystokinin-like immunoreactive fibres of the dorsal horn originate in the spinal ganglia via capsaicin-sensitive C afferents; and some from intrinsic neurons, particularly islet cells. Other fibres may come from supraspinal centres, other local neurons or capsaicin-insensitive afferents from the spinal ganglia. The results are discussed with regard to data in the literature, particularly those concerned with the specificity of the cholecystokinin antibodies; it is hypothesized that several types of cholecystokinin-like immunoreactive peptides may be present in the dorsal horn, depending on their origin (supraspinal, intrinsic or peripheral).(ABSTRACT TRUNCATED AT 400 WORDS)     

Enkephalin-, thyrotropin-releasing hormone- and substance P-immunoreactive axonal innervation of the ventrolateral dendritic bundle in the cat sacral spinal cord: An ultrastructural study

The distribution and synaptic arrangement of thyrotropin-releasing hormone-, substance P- and enkephalin-immunoreactive axonal boutons have been studied in the ventrolateral nucleus (Onuf's nucleus) of the upper sacral spinal cord segments in the cat. For this purpose, the peroxidase-antiperoxidase immunohistochemical technique was used. Immunoreactive axonal boutons were traced in complete series of sections in order to reveal synaptic contacts with the bundled dendrites of the ventrolateral nucleus. As judged from the cross-sectional diameter of the postsynaptic dendrites, the distribution of immunoreactive boutons was non-random. Enkephalin-immunoreactive axonal boutons, presumed to be mostly of segmental origin, displayed a rather restricted distribution to mainly (> 80%) medium-to-large dendrites. Thyrotropin-releasing hormone-immunoreactive boutons, that derive from supraspinal levels, were also found to impinge on medium-to-large dendrites (> 80%), indicating a proximal location within the dendritic trees. The skewness toward large postsynaptic dendrites was even more marked for thyrotropin-releasing hormone- than for enkephalin-immunoreactive boutons. Substance P-immunoreactive boutons, that are of either supraspinal or spinal origin, showed a more even distribution throughout the dendritic trees, including both thin distal branches and thick proximal dendrites. In view of the well-known fact that virtually all thyrotropin-releasing hormone-immunoreactive boutons in the ventral horn cocontain substance P (and serotonin) it was assumed that substance P-immunoreactive boutons in synaptic contact with the finest-calibre dendrites as well as most of those with a very proximal juxtasomatic location on the dendritic trees were of segmental origin, while those impinging on medium-to-large dendrites could be of either spinal or supraspinal origin. Fine-calibre dendrites (< 1 μm) represent about 25% of the dendritic branches in the ventrolateral nucleus, but receive, with the exception of substance P (8%), very little (< 3%) peptidergic or GABAergic (Ramírez-León and Ulfhake, 1993) input, although the degree of dendritic membrane covering by bouton profiles in the ventrolateral nucleus does not seem to vary much with the calibre of the postsynaptic dendrite (Ramírez-León and Ulfhake, 1993). Both substance P- and enkephalin-immunoreactive axonal boutons established synaptic contact with more than one dendrite. Furthermore, one and the same bouton could be found in contact with two dendrites that were coupled to each other by a dendro-dendritic contact of desmosomal or puncta adherentia type. This synaptic arrangement was, however, not seen among thyrotropin-releasing hormone-immunoreactive boutons, indicating that these axonal boutons act on a single postsynaptic element, while inputs intrinsic to the spinal cord can show a divergence also at the terminal level.     

Substance P neuronal organization in the median region of the interpeduncular nucleus of the cat: an electron microscopic analysis

The localization of substance P-like immunoreactivity in the interpeduncular nucleus using the peroxidase-antiperoxidase technique, revealed that the median region of the interpeduncular nucleus was one area rich in substance P-like immunoreactive processes. The ultrastructural characteristics of these substance P-like immunoreactive processes and their organization within the middle zone of the median region of the interpeduncular nucleus was studied. Substance P-like immunoreactivity was found in the perikaryon of small neurons, and in proximal and small dendrites. The substance P-like immunoreactive cell bodies and proximal dendrites receive a variety of unlabeled synaptic terminals. The immunoreactive small dendrites usually formed the central component of a "rosette"-like formation with unlabeled terminals. A few immunoreactive small unmyelinated axons and boutons were also present in the neuropil. The substance P-like immunoreactive boutons contained mainly small round vesicles with some large dense-core vesicles. These substance P-like immunoreactive boutons were presynaptic to unlabeled dendritic profiles, and frequently to substance P-like immunoreactive dendritic profiles. They were also seen in apposition to unlabeled boutons. Substance P-like immunoreactive boutons were not found to synapse with the crest-like dendritic processes in this part of the interpeduncular nucleus. It is suggested on the basis of morphological features, that some of the unlabeled terminals synapsing on substance P-like immunoreactive dendrities, may be cholinergic in nature.     

Presynaptic localization of the carboxy-terminus epitopes of the mu opioid receptor splice variants MOR-1C and MOR-1D in the superficial laminae of the rat spinal cord.

Opioids inhibit nociceptive transmission at the level of the spinal cord, possibly through inhibition of neurotransmitter release by presynaptic mu opioid receptors (MORs) thus preventing the activation of ascending pathways and the perception of pain. Most nociceptive primary afferents are unmyelinated fibers containing peptides such as substance P and/or calcitonin gene-related peptide. However, few terminals contain both substance P and MOR. Recently, we identified new carboxy-terminal MOR splice variants that are localized in the superficial laminae of the dorsal horn. We now report the precise cellular distribution of two of these MOR-1 variants, MOR-1C (exon 7/8/9 epitope) and MOR-1D (exon 8/9 epitope), at the ultrastructural level. In the superficial laminae of the dorsal horn, the majority of the labeling of MOR-1C and MOR-1D was found in unmyelinated axons. This distribution contrasts with that of MOR-1 (exon 4 epitope), in which labeling is equally found in dendrites and soma, as well as in axons. The presence of dense core vesicles in many of the MOR-1C-like immunoreactive terminals implies that this splice variant might be involved in presynaptic inhibition of transmitter release from peptide-containing afferents to the dorsal horn. Consistent with this finding, confocal microscopy analyses showed that many MOR-1C profiles in laminae I-II also contained calcitonin gene-related peptide, whereas fewer MOR-1 profiles contained either substance P or calcitonin gene-related peptide in this same region.From these findings we suggest that there are differential distributions of MOR-1 splice variants as well as distinct peptide colocalizations in the dorsal horn.     

Ultrastructural visualization of glutamate and aspartate immunoreactivities in the rat dorsal horn, with special reference to the co-localization of glutamate, substance P and calcitonin-gene related peptide.

Antisera raised against the fixation products of L-glutamate and L-aspartate were used, singly or in combination, to study the ultrastructural localization of the amino acids in the rat dorsal horn, with post-embedding immunogold techniques. Immunostaining for each of the amino acids was also combined with immunolocalization of GABA, an important inhibitory neurotransmitter in the spinal cord, or synaptophysin, a synaptic vesicle glycoprotein. In addition, we examined the localization of glutamate immunoreactivity in relation to that of calcitonin-gene related peptide and substance P, two neuropeptides present in high concentrations in the dorsal horn. Glutamate- and aspartate-immunoreactive neuronal cell bodies, dendrites, axons and terminals were apparent in the first three laminae of the dorsal horn. In somatic and dendritic profiles, the immunolabel was present over the general cytoplasm and mitochondria; in the terminals, it was found over small, agranular vesicles, mitochondria and, at times, synaptic densities. Quantitative estimation indicated that the colloidal gold density in the glutamate-immunoreactive terminals was five-fold more than in any other neuronal profile. Both glutamate- and aspartate-immunopositive terminals made asymmetric synaptic contacts onto unlabelled dendrites; glutamate-positive terminals often formed the core of type I and II glomeruli. After double labelling of the same sections, glutamate and aspartate immunoreactivities consistently occurred in different axonal and terminal profiles. In these preparations, it was clearly seen that glutamate-immunoreactive terminals were far more numerous than (more than 10-fold) those immunoreactive for aspartate. Double labelling for glutamate or aspartate and GABA also revealed distinct staining of different terminals. Simultaneous immunolocalization of each of the amino acids and synaptophysin showed the amino acid and glycoprotein immunoreactivities co-localized in small, agranular vesicles in immunoreactive terminals. Finally, triple labelling of the same sections for glutamate, calcitonin gene-related peptide and substance P revealed that glutamate was often co-localized with either of the two neuropeptides in the same axonal boutons; terminals that showed simultaneous labelling for glutamate, calcitonin gene-related peptide and substance P were also noted. In all cases, the glutamate immunoreactivity was restricted to small, clear vesicles whereas the neuropeptide immunoreactivities were present in larger, dense-cored vesicles. Our observations demonstrate that there is an abundant glutamate immunoreactivity in the superficial layers of the rat dorsal horn, localized in neuronal profiles distinct from those containing aspartate or GABA.(ABSTRACT TRUNCATED AT 400 WORDS)     

Substance P immunoreactivity in the rat interpeduncular nucleus: synaptic interactions between substance P-positive profiles and choline acetyltransferase- or glutamate decarboxylase-immunoreactive structures.

The subnuclear and synaptic distribution of substance P immunoreactivity was examined in the rat interpeduncular nucleus at the light and electron microscope level. The nucleus possessed a prominent substance P-immunoreactive axonal plexus in the lateral and dorsomedial subnuclei, and in the dorsal cap of the rostral subnucleus. The density of substance P-immunoreactive axons in the remaining subnuclear divisions was sparse to moderate. Terminals of immunoreactive axons contained spherical vesicles and formed asymmetric contacts on dendritic processes exclusively. Immunoreactive neurons, restricted to the rostral subnucleus, possessed long, sparsely branched dendrites. Unlabelled terminals containing either spherical or pleomorphic vesicles contacted substance P-immunoreactive dendritic profiles. Axodendritic and axosomatic synapses containing substance P immunoreactivity pre- and postsynaptically were not observed. Ultrastructural evidence for synaptic relationships between substance P-containing profiles and those containing either choline acetyltransferase or glutamate decarboxylase was obtained by means of double antigen immunohistochemistry. Terminals of fasciculus retroflexus axons stained for choline acetyltransferase immunoreactivity formed asymmetric synaptic contacts with substance P-immunoreactive dendritic profiles. Few substance P-positive dendrites in the rostral subnucleus received terminals possessing glutamate decarboxylase activity. Unlabelled terminals containing either spherical or pleomorphic vesicles contacted substance P- and glutamate decarboxylase-immunoreactive dendritic profiles simultaneously. Terminals possessing either substance P or glutamate decarboxylase immunoreactivity formed synaptic contacts with dendritic processes of neurons in the lateral subnucleus. Many of the neurons within this subnuclear division contained glutamate decarboxylase. This study provides direct evidence of synaptic relationships between choline acetyltransferase-immunoreactive axons and substance P-immunoreactive dendritic profiles, and between substance P-positive axons and glutamate decarboxylase-immunoreactive dendrites. These findings reveal that two types of transmitter-specific axons of the fasciculus retroflexus innervate neuronal populations of the interpeduncular nucleus stained immunohistochemically for either substance P or glutamate decarboxylase.     

大鼠脊髓P物质受体定位分布的免疫细胞化学研究

用免疫细胞化学技术对大鼠脊髓和脊神经节内Ｐ物质受体的定位分布进行了系统的研究。结果证明，Ｐ物质受体阳性胞体和树突主要密集地分布于脊髓全长的Ⅰ层，此外，还发现Ⅱ层的外侧部出现少量阳性胞体和树突，来自Ⅲ层的阳性树突穿过Ⅱ层后进入Ⅰ层；Ⅲ～Ⅳ和Ⅹ层也可见中等密度的阳性胞体和树突；Ⅵ层和Ⅶ层仅见少量阳性胞体和树突，但胸髓中间带外侧核、骶髓副交感运动核、骶髓后连合核内可见大量浓染的阳性胞体和树突；Ⅷ、Ⅸ层、Ｏｎｕｆ氏核、外侧颈核和外侧脊索核也有阳性胞体和树突．灰质内的阳性胞体的树突还伸向前索和外侧索，有时可达脊髓的边缘．此外，脊神经节内也可见少量散在且均匀分布的小型阳性胞体．     

Opioid neurons and pain modulation: an ultrastructural analysis of enkephalin in cat superficial dorsal horn

To clarify the circuitry through which opioid compounds modulate spinal and trigeminal nociceptive transmission, we have examined the synaptic associations formed by leucine-enkephalin-containing (enkephalin) neurons in the superficial dorsal horn of the cat. As described previously, punctate enkephalin immunoreactivity is concentrated in the marginal layer (lamina I) and in both the outer and inner layers of the substantia gelatinosa (lamina IIo and IIi). In colchicine treated cats, enkephalin perikarya are most numerous in lamina I and at the border between laminae I and II. Ultrastructural analysis reveals that enkephalin cells receive a diverse afferent input. The majority of afferent inputs are presynaptic to the enkephalin dendrites; few axosomatic synapses are seen. Among these presynaptic axonal profiles are unlabeled axons which resemble primary afferent terminals, including the characteristic central axonal varicosity. Enkephalin dendrites are also postsynaptic to enkephalin immunoreactive axons. Two types of enkephalin axonal profiles appear in the superficial dorsal horn. Class I profiles are only found in lamina I. These are large profiles which form few synapses; those synapses made are axodendritic. Class II enkephalin axons are smaller and are distributed in both layers I and II. While Class II axons most commonly form axo-dendritic synapses, they also form axo-axonic synapses with flat vesicle-containing profiles; the latter are generally presynaptic to the enkephalin terminals. Serial analysis further revealed that both the enkephalin and the flat vesicle-containing profile synapse onto a common dendrite. Although enkephalin axons frequently lie adjacent to round vesicle-containing profiles, anatomical evidence that opioid axons form synapses with this type of ending was not found. An additional type of enkephalin vesicle containing-profile is found in layer IIi; its morphological features do not clearly distinguish its axonal or dendritic origin. These endings are typically postsynaptic to unlabelled central endings, and provide minimal presynaptic input to other elements in the neuropil. Like some class II axons, these labelled profiles contain vesicles which cluster at the membrane immediately adjacent to unlabelled central axons. These results indicate that spinal enkephalin neurons receive a variety of synaptic inputs. These include inputs which may derive from primary afferent axons. Enkephalin neurons, in turn, influence nociceptive transmission predominantly through postsynaptic mechanisms. Finally, while we did not observe enkephalin terminals presynaptic in an axoaxonic relationship, the possibility that enkephalin neurons modulate the excitability of fine fiber nociceptive and nonnociceptive afferents via "nonsynaptic interactions" is discussed.     

鸡脊髓后角Ⅰ、Ⅱ层内大神经元的免疫细胞化学研究

本研究采用心脏灌流法固定鸡标本 ,切取鸡脊髓颈膨大部分 ,利用免疫细胞化学法 ( PAP法 ) ,在光镜下观察了后角大神经元 ,其周围有大量 SP样免疫反应终扣。大神经元的胞体主要分布在 层、 层外侧区和 层内 ;树突伸向 层、 层外侧区以及 层的深部。电镜下观察 ,在 层内 ,大神经元的树突与突触小球的 P物质阳性中心终末形成突触。这一发现提示 ,大神经元直接接受来自初级感觉传入终末的信息。结合以前的研究推测 :这些大神经元是脊髓丘脑束的起始神经元     

Interferon-gamma receptors are expressed at synapses in the rat superficial dorsal horn and lateral spinal nucleus

Interferon-gamma can facilitate the spinal nociceptive flexor reflex and may elicit neuropathic pain-related behavior in rats and mice. Immunoreactivity for the interferon-gamma receptor (IFN-gamma R) occurs in the superficial layers of the dorsal horn and the lateral spinal nucleus in the rat and mouse spinal cord, as well as in subsets of neurons in the dorsal root ganglia. The aim of the present study was to examine the cellular localization and origin of the IFN-gamma R in the spinal cord. As viewed by confocal microscopy, the immunopositivity for the IFN-gamma R was co-localized with that of the presynaptic marker synaptophysin and with neuronal nitric oxide synthase in the lateral spinal nucleus, whereas only a minor overlap with these molecules was observed in laminae I and II of the dorsal horn. There was no co-localization of the IFN-gamma R with markers for astrocytes and microglial cells. Ultrastructurally, the IFN-gamma R was found predominantly in axon terminals in the lateral spinal nucleus, but at postsynaptic sites in dendrites in laminae I and II. The IFN-gamma R expressed in neurons in dorsal root ganglia was transported in axons both centrally and peripherally. Hemisection of the spinal cord caused no reduction in immunolabelling of the IFN-gamma R in the dorsal horn or the lateral spinal nucleus. Since rhizotomy does not affect the immunolabelling in the lateral spinal nucleus, our observation indicates that the presynaptic receptors in this nucleus are derived from intrinsic neurons. The localization of the IFN-gamma R in the spinal cord differed from that of the AMPA glutamate receptor subunits 2 and 3 and the substance P receptor (NK1). Our results, showing localization of IFN-gamma R to pre- and postsynaptic sites in the dorsal horn and lateral spinal nucleus indicate that IFN-gamma can modulate nociception at the spinal cord level.     

Fine structure of synapses associated with characterized postsynaptic dorsal column neurons in the cat

Fourteen dorsal horn neurons with axons projecting through the dorsal columns were identified either by electrophysiological methods (and subsequently injected with horseradish peroxidase) or by retrograde labelling with horseradish peroxidase in cats. All neurons were contacted by small (less than 2 micron) boutons containing spherical or elongated agranular vesicles. One neuron with its soma located in lamina III received additional contacts from central elements of glomerular complexes. Neurons with somata located more ventrally (deep lamina IV and V) were also postsynaptic to large (greater than 2 microns) electron lucent profiles which formed multiple synapses with the labelled cells. Some boutons presynaptic to postsynaptic dorsal column neurons were themselves postsynaptic to profiles containing pleiomorphic agranular vesicles at axoaxonic synapses. They also occasionally participated in triadic complexes. It is concluded that the synaptic arrangements formed by boutons in association with postsynaptic dorsal column neurons differ significantly from those associated with spinocervical neurons. Such differences might provide the anatomical substrate for the observed receptive field characteristics of these neurons.