GABA and glycine in synaptic microcircuits associated with physiologically characterized primary afferents of cat trigeminal principal nucleus

Previous studies suggest that sensory information conveyed through trigeminal afferents is more strongly controlled at the level of the first synapse by GABA-mediated presynaptic mechanisms in the trigeminal principal sensory nucleus (Vp) than other sensory nuclei. However, it is unknown if such a mechanism is common to functionally different classes of primary afferent in the same nucleus or across the nuclei. To address these issues, the present study focused on synaptic microcircuits associated with slowly adapting (SA) mechanosensory afferents innervating the periodontal ligaments in the cat Vp and attempted to examine GABA, glycine, and glutamate immunoreactivity in axon terminals involved in the circuits. Afferents were physiologically characterized before injection of horseradish peroxidase (HRP) and preparation for electron microscopy. HRP-labeled afferent boutons were serially sectioned and immunostained with antibodies against GABA, glycine, and glutamate using a postembedding immunogold method. All the afferent boutons examined contacted non-primary dendrites and they were frequently postsynaptic to unlabeled axons (p-endings). Axodendritic and axoaxonic contacts per afferent bouton were 1.3 (46/35) and 2.0 (70/35), respectively. Most p-endings were immunoreactive for GABA (63/70) and also glycine was co-stained in the majority of the p-endings (49/63). Thirty percent of p-endings with the colocalization of GABA and glycine participated in synaptic triads where a p-ending formed a synapse with the same dendrite as the afferent bouton. None of the p-endings was immunoreactive for glutamate. Most afferent boutons were enriched with glutamate but were immunonegative for GABA and glycine. This study provides evidence suggesting that transmission from SA afferents is strongly controlled presynaptically by GABAergic interneurons with colocalized glycine, and that a proportion of these interneurons, involved in synaptic triads, may also have postsynaptic inhibitory actions on target neurons of the SA afferents.     

Colocalization of neurotransmitters in presynaptic boutons of inhibitory synapses in the lamprey spinal cord

Electron-microscopic immunocytochemical studies were performed to detect GABA and glycine immunoreactivity in presynaptic axon terminals in the central gray matter of the spinal cord of the lampreyLampetra fluviatilis. The immunopositive presynaptic terminals contacting identified dendrites of motoneurons and unidentified postsynaptic profiles included terminals immunopositive for GABA only (44%) and glycine only (26%), as well as terminals containing GABA and glycine (30%). Glycine-immunopositive presynaptic terminals contained flattened synaptic vesicles. Large synaptic vesicles with dense cores were present along with classical synaptic vesicles in 74% of GABA-immunopositive boutons. Translated from Rossiiskii Fiziologicheskii Zhurnal imeni I. M. Sechenova, Vol. 85, No. 4, pp. 515–522, April, 1999.     

Ultrastructural subtypes of glutamate-immunoreactive terminals on rat trigeminal motoneurones and their relationships with GABA-immunoreactive terminals

 Electron-microscopic immunolabelling methods were used to study the relationships between glutamate-immunoreactive and γ-aminobutyric acid (GABA)-immunoreactive synapses on trigeminal motoneurones labelled by the retrograde transport of horseradish peroxidase. Serial sections were cut through the motor nucleus, alternate sections were incubated with antibodies to glutamate and GABA, and the immunopositive nerve terminal profiles were recognized using a quantitative, postembedding immunogold method. Boutons exhibiting high levels of glutamate immunoreactivity and GABA-immunoreactive boutons both formed axo-dendritic and axo-somatic synaptic contacts on labelled motoneurones. Boutons strongly immunopositive for glutamate were not immunopositive for GABA, and vice versa. Strongly glutamate immunoreactive boutons received axo-axonic synaptic contacts but did not form such contacts, while GABA-immunoreactive boutons formed axo-axonic synapses but did not receive them. The presynaptic elements at all axo-axonic synapses on to glutamate-immunoreactive boutons sampled were GABA-immunopositive. These data provide ultrastructural evidence in support of the roles of glutamate and GABA as transmitters at synapses on trigeminal motoneurones, and for presynaptic control of transmission at glutamatergic synapses by GABA acting at receptors at axo-axonic synapses. The vast majority (more than 90%) of strongly glutamate immunoreactive boutons contained spherical synaptic vesicles, in contrast to GABA-immunoreactive boutons, which contained pleomorphic vesicles. Most of the glutamate-immunoreactive boutons (67%) formed asymmetrical synaptic active zones, many of which (47% of total) were associated with subsynaptic dense ”Taxi” bodies (T-terminals), while a smaller population of boutons (21%) formed symmetrical synapses, and a few (11%) made synapses associated with subsynaptic cisternae (C-terminals). The heterogeneity of active zone ultrastructure of boutons identified as being glutamatergic on the basis of their high levels of immunolabelling is discussed in relation to possible differences in co-transmitters released, origins of the synaptic input or post-synaptic receptor subtypes activated. Received: 13 May 1996 / Accepted: 9 September 1996     

Quantitative ultrastructure of synapses on functionally identified primary afferent neurons in the cat trigeminal mesencephalic nucleus

Though a number of studies have reported the presence of synapses on neurons in the trigeminal mesencephalic nucleus (Vmes), there have been no quantitative studies of either the density of innervation, or the ultrastructure, of the synapses on single, physiologically identified neurons in this nucleus. In this study we recorded from single neurons in the Vmes, identified them as being either muscle spindle afferents (MS) or periodontal ligament mechanoreceptor afferents (PL), and then labeled the neurons by intra-axonal injection of horseradish peroxidase (HRP). The material was first processed to reveal the HRP activity, following which ultrathin sections through the labeled somata were cut and examined under the electron microscope. Complete serial reconstructions were made through the soma of one MS neuron and one PL neuron, and the contacts on the neurons reconstructed. Boutons were found on the soma, spines, appendages and the axon hillock and the initial segment of the axon. The numbers of boutons terminating on the two neurons were 198 (PL) and 424 (MS), giving a packing density of 4.4 and 10.7 boutons respectively (i.e., number of boutons/100 micron 2 of the postsynaptic membrane). Boutons could be separated into two types on the basis of their vesicles: those containing clear, round vesicles (i.e., S-type) and those containing a mixture of round, oval and flattened vesicles (P-type). Ninety-five (PL neuron) and 99% (MS neuron) of terminals on the two neurons were P-type. All the S-type boutons and 80% of the P-type boutons formed asymmetric synaptic contacts while 10% of the P-type boutons made symmetric contacts. Quantitative measurements of the P-type boutons on the labeled neurons, in which the data of MS and PL neurons were pooled, revealed that bouton volume was highly correlated with bouton surface area, active zone number, total active zone area, vesicle number, and mitochondrial volume. However, comparing the quantitative measurements of the P-type boutons with those of previously reported vibrissa afferent terminals and their associated axon terminals revealed that all the parameters were smaller for the P-type boutons (on Vmes neurons) than those of the vibrissa afferent terminals but similar to those of axon terminals presynaptic to the vibrissa afferents. Taken together, our results emphasize the wide scope for synaptic interactions in the Vmes and suggest that it may be more fruitful to view the Vmes as an integrating center.     

GABA能神经成分对大鼠咀嚼肌本体觉传入进行初级整合的形态学基础──Ricin跨节溃变、HRP逆行追踪与抗GABA免疫组化法结合的电镜观察

本文作者曾证明,三叉神经感觉主核背内侧区和三叉上核尾外侧部是大鼠三又神经本体觉三级传入通路中继站。为证实此二处的ＧＡＢＡ能丘脑投射神经元和中间神经元是否参与三叉神经本体觉的传导和初级整合并阐明其整合机制,用Ｒｉｃｉｎ毁损三叉神经中脑核神经元及其终末,ＨＲＰ逆行标记此区的丘脑投射神经元以及抗ＧＡＢＡ免疫组化三者结合的方法,在电镜下发现：（１）中脑核神经元的溃变终末与该二区神经毯中的ＧＡＢＡ能三叉—丘脑投射神经元的胞体和树突形成轴—体和轴—树突触。（２）溃变终末与ＧＡＢＡ样中间神经元形成突触。（３）ＧＡ—ＢＡ样终末与ＧＡＢＡ能三叉—丘脑投射神经元形成突触。（４）ＧＡＢＡ样终末也与ＧＡＢＡ样神经元的胞体和树突形成自突触。此外还观察到ＧＡＢＡ能三叉—丘脑投射神经元的回返侧支与ＧＡＢＡ能中间神经元间的突触连接和大的“贝壳状”ＧＡＢＡ样终末与周围的神经成分形成复杂的突触复合体。证实该二区的ＧＡＢＡ能三叉—丘脑投射神经元直接参与三叉神经本体党的向心传导;同时,ＧＡＢＡ能中间神经元和其它来源的ＧＡＢＡ能神经成分也参与本体觉传导的初级整合作用,其间存在着突触前、后抑制,去抑制,突触前易化,回返抑制等复杂整合机制的结构基础。     

Central connectivity of the chorda tympani afferent terminals in the rat rostral nucleus of the solitary tract

The rostral nucleus of the solitary tract (rNST) receives gustatory input via chorda tympani (CT) afferents from the anterior two-thirds of the tongue and transmits it to higher brain regions. To help understand how the gustatory information is processed at the 1st relay nucleus of the brain stem, we investigated the central connectivity of the CT afferent terminals in the central subdivision of the rat rNST through retrograde labeling with horseradish peroxidase, immunogold staining for GABA, glycine, and glutamate, and quantitative ultrastructural analysis. Most CT afferents were small myelinated fibers (<5 µm² in cross-sectional area) and made simple synaptic arrangements with 1–2 postsynaptic dendrites. It suggests that the gustatory signal is relayed to a specific group of neurons with a small degree of synaptic divergence. The volume of the identified synaptic boutons was positively correlated with their mitochondrial volume and active zone area, and also with the number of their postsynaptic dendrites. One-fourth of the boutons received synapses from GABA-immunopositive presynaptic profiles, 27 % of which were also glycine-immunopositive. These results suggest that the gustatory information mediated by CT afferents to the rNST is processed in a simple and specific manner. They also suggest that the minority of CT afferents are presynaptically modulated by GABA- and/or glycine-mediated mechanism.     

The distribution of inhibitory and excitatory synapses on single, reconstructed jaw-opening motoneurons in the cat

In a previous study, we reported that the distribution of inhibitory input, in contrast to excitatory input, decreased somatofugally along dendrites of cat jaw-closing alpha-motoneurons [J Comp Neurol 414 (1999) 454]. The present study examined the distribution of GABA, glycine, and glutamate immunopositive boutons covering horseradish peroxidase-labeled cat jaw-opening motoneurons. The motoneurons were divided into four compartments: the soma, and primary, intermediate, and distal dendrites. Ninety-seven percent of the total number of studied boutons had immunoreactivity for at least one of the three amino acids. The proportion of boutons immunoreactive for GABA and/or glycine was lower than the proportion of boutons immunoreactive for glutamate. Boutons immunoreactive to glycine alone were more numerous than boutons double-labeled for GABA and glycine, which, in turn, occurred more frequently than boutons immunoreactive to GABA alone. The percentage synaptic covering (proportion of membrane covered by synaptic boutons) of the putatively excitatory (glutamate containing) and putatively inhibitory (GABA and/or glycine containing) boutons decreased somatofugally along the dendrites. Such systematic variations were not seen in the packing density (number of boutons per 100 microm(2)); the packing density showed a distinct drop between the soma and primary dendrites but did not differ significantly among the three dendritic compartments. Overall, the packing density was slightly higher for the putatively excitatory boutons than for the inhibitory ones. When taken together with previous analyses of jaw-closing alpha-motoneurons the present data on jaw-opening alpha-motoneurons indicate that the two types of neuron differ in regard to the nature of synaptic integration in the dendritic tree.     

GABAergic neurons in the mouse superficial dorsal horn with special emphasis on their relation to primary afferent central terminals

Immunocytochemical studies were carried out on the morphological relation between primary afferent central terminals (C-terminals) and GABAergic neurons in the mouse superficial dorsal horn. The superficial dorsal horn is composed of many synaptic glomeruli comprising two types: Type I with centrally located CI-terminals surrounded by several dendrites and few axonal endings, and Type II with centrally located CII-terminals surrounded by several dendrites and a few axonal endings. The CI-terminals are sinuous or scalloped with densely packed agranular synaptic vesicles, a few granular synaptic vesicles and mitochondria, and show an electron dense axoplasm, whereas the CII-terminals are large and round or rectangular with evenly distributed agranular synaptic vesicles, a number of granular synaptic vesicles and mitochondria, and show an electron opaque axoplasm. The immunoreaction of GABA was remarkable in the superficial laminae of the dorsal horn. Many interneuronal somata in the substantia gelatinosa showed GABAergic immunoreactivity. The immunoreaction was seen in the entire GABAergic neuroplasm, but not in the nucleus and its envelope. Most GABAergic features appeared as dendrites making postsynaptic contact with CI- or CII-terminals; i.e., numerous C-terminals made presynaptic contact with GABAergic dendrites. GABA immunoreactivity was seen over round synaptic vesicles and mitochondrial membranes. A few CII-terminals made presynaptic contact with GABAergic interneuronal somata. Previous physiological and anatomical studies have suggested that not only the cutaneous nociceptive primary afferent C-terminals but also mechanoreceptive primary afferent C-terminals make presynaptic contact with the GABAergic dendrites, boutons and soma. The presynaptic relation of these primary afferents with GABAergic neurons seems to provide morphological support for the essential feature of the gate control theory: primary afferent fibers may play a part in the modulation of nociceptive information via GABAergic neurons in the superficial dorsal horn. Small GABAergic terminals were found to make contact with blood capillaries suggesting the release of GABA into circulation.     

NEURAL PATHWAYS OF TRIGEMINAL PROPRIOCEPTIVE AFFERENTS COORDINATE ORAL MOTOR BEHAVIORS

Neural pathways and synaptic connections from the trigeminal mesencephalic nucleus (Vme) neurons to the cranial motor nuclei were studied in the rat using double labelling methodologies of intracellular Neurobiotin staining combined with retrograde horseradish peroxidase (HRP) transport, anterograde biotinylated dextran amine (BDA) tracing combined with retrograde HRP transport, and a dual fluorescent labelling of BDA anterograde combined tracing with Cholera Toxin B (CTB) retrograde transport. Direct projections and synapses were demonstrated from Vme neuronal boutons to motoneurons (MNs) of the trigeminal motor nucleus (Vmo), the hypoglossal nucleus (XⅡ) and the ambiguus nucleus (Amb). Indirect projections and pathways from Vme neurons to the cranial motor nuclei including Vmo, XⅡ, the facial nucleus (VⅡ) and the cervical spinal cord (C1～5) were seen to relay on their premotor neurons. The premotor neurons of above cranial motor nuclei were overlapped in bilateral premotor neuronal pool including the parvocellular reticular formation (PCRt) and its alpha division (PCRtA), the dorsomedial part of the spinal trigeminal nucleus oralis (Vodm), and interpolaris (Vidm), the medullary reticular nucleus dorsal division (MdD), the supratrigeminal region (Vsup) and the dorsomedial part of the principal trigeminal sensory nucleus (Vpdm). Synapses between Vme neuronal boutons and Vmo and XⅡ MNs and XII premotor neurons were predominantly asymmetric. There were four types of synaptic organizations, i.e. synaptic convergence; synaptic divergence presynaptic inhibition and afferent feedforward inhibition seen between Vme boutons and Vmo, XⅡ MNs and between Vme boutons and XⅡ premotor neurons. The results of present studies have demonstrated direct pathways from the trigeminal proprioceptive afferents to Vmo, XⅡ and Amb MNs, and indirect pathways from the trigeminal proprioceptive afferents to bilateral Vmo, XⅡ, VⅡ and C1～5 via their premotor neurons. It provides neuroanatomical network to elucidate trigeminal proprioceptive afferents coordinate oral motor behaviors.     

GABA- and glycine-immunoreactive terminals contacting motoneurons in lamprey spinal cord

Double postembedding GABA- and glycine-immunostaining was performed on the lamprey (Lampetra fluviatilis) spinal cord after previous HRP labeling of motoneurons. Immunopositive boutons contacting motoneurons were counted and distinguished as GABA (39%), glycine (30%) and both GABA+glycine-immunopositive (31%). Densely-packed, flattened synaptic vesicles were only observed in glycine-immunopositive boutons while GABA-immunoreactive and GABA+glycine-immunoreactive boutons contained rounded or oval synaptic vesicles. Dense-core vesicles of different diameters were associated with conventional synaptic vesicles in 74% of GABA-only-immunopositive boutons, 50% of double GABA+glycine-immunopositive boutons, but were only observed in 9% of glycine-only-immunopositive boutons. The presence of terminals immunoreactive to either GABA or glycine contacting the motoneurons suggests that there is a morphological substrate for both GABAergic and glycinergic postsynaptic inhibition of motoneurons in the lamprey spinal cord.     

Axo-axonic GABA-immunopositive synapses on the primary afferent fibers in frogs.

In three frog species Rana esculenta, Rana temporaria and Xenopus laevis, the contacts established by gamma-aminobutyric acid and glutamate decarboxylase immunoreactive (-ir) terminals upon primary afferent fibers were studied using confocal and electron microscopy. For confocal microscopy, the primary afferent fibers were labeled through the dorsal root with Dextran-Texas Red, whereas gamma-aminobutyric acid and glutamate decarboxylase immunoreactivity were revealed with fluorescein isothiocyanate. Appositions of gamma-aminobutyric acid and glutamate decarboxylase immunoreactive profiles onto primary afferent fibers were observed and were considered as putative axo-axonic contacts of GABAergic terminals upon primary afferents. The latter was confirmed by the ultrastructural finding of axo-axonic synapses from gamma-aminobutyric acid immunopositive boutons upon the HRP-labeled primary afferent fibers in postembedding immunoelectron microscopic study. Such synapses may represent the morphological basis of GABAergic presynaptic inhibition of primary afferent fibers.     

Morphological evidence for GABA/glycine-cocontaining terminals in synaptic contact with neurokinin-1 receptor-expressing neurons in the sacral dorsal commissural nucleus of the rat

Previous studies have shown that neurons in the sacral dorsal commissural nucleus (SDCN) express neurokinin-1 receptor (NK1R) and can be modulated by the co-release of GABA and glycine (Gly) from single presynaptic terminal. These results raise the possibility that GABA/Gly-cocontaining terminals might make synaptic contacts with NK1R-expressing neurons in the SDCN. In order to provide morphological evidence for this hypothesis, the triple-immunohistochemical studies were performed in the SDCN. Triple-immunofluorescence histochemical study showed that some axon terminals in close association with NK1R-immunopositive (NK1R-ip) neurons in the SDCN were immunopositive for both glutamic acid decarboxylase (GAD) and glycine transporter 2 (GlyT2). In electron microscopic dual- and triple-immunohistochemistry for GAD/GlyT2, GAD/NK1R, GlyT2/NK1R, or GAD/GlyT2/NK1R also revealed dually labeled (GAD/GlyT2-ip) synaptic terminals upon SDCN neurons, as well as GAD- and/or GlyT2-ip axon terminals in synaptic contact with NK1R-ip SDCN neurons. These results suggested that some synaptic terminals upon NK1R-expressing SDCN neurons co-released both GABA and Gly.     

GABA-, glycine-, and glutamate-immunoreactive bouton profiles in apposition to neurons of the central cervical nucleus in the rat

The neurons of the central cervical nucleus (CCN) convey information about the position and movements of the head, and receive excitatory input from dorsal neck muscles and the labyrinth. Both of these afferent sources form glutamatergic synaptic contacts with CCN neurons. However, these sensory afferent sources can also inhibit CCN neurons. To further elucidate the synaptic organization, we made an electron microscopic investigation, identifying and evaluating the relative frequency of bouton profiles containing the inhibitory transmitters GABA and glycine in apposition to identified CCN neurons. In addition, labeling for glutamate was performed. The identification of the CCN neurons was made possible by injections of retrograde tracer substances into the cerebellum. These substances were made visible by preembedding immunocytochemistry or postembedding immunogold staining. Such staining was also used to detect the three amino acids that were found in boutons apposed to the identified neurons (cf. Ornung et al., J. Comp. Neurol. 1996;365:413-426; Lind? et al., J. Comp. Neurol. 2000;425:10-23). Due to the relatively poor transport of the tracer substances into dendrites of the CCN neurons, the analysis was restricted to the cell body and included bouton profiles in direct apposition to the soma membrane. Data from 10 CCN neurons revealed that about 50% of the apposing bouton profiles were immunoreactive for GABA, and about 34% for glycine. In four neurons, the degree of colocalization of GABA and glycine was determined to be close to 30%. Thus, the vast majority of glycine-labeled profiles also contained GABA, while a considerable fraction of the profiles were immunoreactive for only GABA. The values for glycine immunoreactive bouton profiles presented here may represent somewhat low estimates, depending on the method used. Data from four neurons showed that about 18% of the profiles were labeled for glutamate. The large fraction of purely GABA immunoreactive profiles, or at least a substantial group of them, is suggestive of their derivation from axons descending from the brainstem.     

GAD67-GFP转基因小鼠三叉神经脊束核吻侧亚核和极间亚核内GABA和甘氨酸共存神经元的观察

本文以GAD67-GFP基因敲入小鼠为研究工具,利用免疫荧光组织化学双重染色技术观察了小鼠三叉神经脊束核(spi-nal trigeminalnucleus,V)的吻侧亚核(Vo)和极间亚核(Vi)神经元内谷氨酸脱羧酶67[GAD67,γ-氨基丁酸(GABA)能神经元的标识物质]和甘氨酸(glycine,Gly)的定位分布和共存情况。结果显示,在Vo和Vi内均有比较密集分布的GAD67和Gly阳性神经元。此外,在Vo和Vi内也可见一些GAD67阳性神经元同时呈Gly阳性反应,即存在GAD67/Gly双标神经元。结果表明,Vo和Vi内有GABA和Gly共存神经元,它们可能在初级传入信息的传递和调制中发挥重要作用。     

Jaw-muscle spindle afferent feedback to the cervical spinal cord in the rat

Putative synaptic contacts between masticatory-muscle spindle afferents and brainstem neurons which project to the cervical spinal cord were studied in rats by combining retrograde and intracellular neuronal labeling. Spinal cord projecting neurons were retrogradely labeled via injection of horseradish peroxidase unilaterally or bilaterally into cervical spinal cord segments C2 through C5. Twenty-four hours after the injection of horseradish peroxidase, one to five jaw-muscle spindle afferent axons were physiologically identified and intracellularly stained with biotinamide on each side of the brainstem. Horseradish-peroxidase-labeled neurons were found bilaterally in the supratrigeminal region, trigeminal principal sensory nucleus, parvicellular reticular nucleus including its alpha division, spinal trigeminal subnuclei oralis and interpolaris and the medullary reticular formation. Retrogradely labeled neurons were most numerous in the spinal trigeminal subnucleus oralis, parvicellular reticular formation and the ventral part of the spinal trigeminal subnucleus interpolaris. A small number of horseradish-peroxidase-labeled neurons were also present in the trigeminal mesencephalic nucleus and spinal trigeminal subnucleus caudalis. Appositions between jaw-muscle spindle afferent boutons and spinal projecting neurons were found in the supratrigeminal region, dorsomedial portions of the trigeminal principal sensory nucleus and spinal trigeminal subnuclei oralis and interpolaris, and the parvicellular reticular formation including its alpha division. Putative synaptic contacts were most frequent in the parvicellular reticular formation and the dorsomedial portion of the trigeminal subnucleus oralis. These results indicate that some orofacial proprioceptive feedback transmitted via the mesencephalic trigeminal nucleus reaches the cervical spinal cord directly and suggests that jaw-muscle spindle afferent feedback reaches the cervical spinal cord predominately via relays in the dorsomedial part of the spinal trigeminal subnucleus oralis and the parvicellular reticular formation. It is hypothesized that these pathways are primarily involved in the coordination of jaw and neck movement during mastication and biting. Materials and methods 27 January 1999 / Accepted: 9 May 1999     

Physiological and morphological correlates of presynaptic inhibition in primary afferents of the lamprey spinal cord.

Patch-clamp recordings in a whole-cell mode were performed on dorsal sensory cells enzymatically isolated from the spinal cord of two lamprey species, Ichthyomyzon unicuspis and Lampetra fluviatilis. The voltage-activated currents through calcium channels were analysed. GABA and the specific GABA(B) receptor agonist baclofen reduced the peak amplitude of inward Ba2+ current, as a robust alternate charge carrier through voltage-dependent Ca2+ channels. These effects were dose-dependent and reversible. GABA(B) receptor antagonists, 2-hydroxysaclofen and delta-amino-n-valeric acid, blocked the reduction of Ba2+ currents by GABA and baclofen, while bicuculline, a GABA(A) receptor antagonist, had no blocking action. GABA and baclofen did not modify the dorsal sensory cell membrane conductance, indicating that they did not activate ligand-gated channels. However, GABA, but not baclofen, considerably increased membrane conductance and induced Cl- currents in isolated multipolar neurons (presumably interneurons and/or motoneurons). These findings suggest that GABA and baclofen action on lamprey dorsal sensory cells is mediated by GABA(B) receptors. We concluded that GABA-mediated presynaptic inhibition of lamprey dorsal sensory cell fibers results from GABA(B) receptor activation followed by a decrease of inward voltage-activated calcium currents. Appositions of GABA-immunoreactive boutons to horseradish peroxidase-labeled fibers from the dorsal root were observed at the ultrastructural level in the dorsal column using postembedding immunogold cytochemistry. It seems likely that these appositions represent the morphological substrate of dorsal sensory cell fiber presynaptic inhibition. In very rare cases, ultrastructural features were observed which could be interpreted as synaptic specializations between the GABA-immunoreactive boutons and the primary afferent fibers. The extrasynaptic action of GABA as a basis of presynaptic inhibition of this population of primary afferent neurons is discussed.     

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.     

Ultrastructural localization of nitric oxide synthase immunoreactivity in the cat ventrobasal complex

This study describes the ultrastructural localization of nitric oxide synthase (NOS) immunoreactivity in the cat ventrobasal complex. NOS immunoreactivity was found in the cell bodies and dendrites of local circuit neurons and in vesicle-containing profiles. The vesicle-containing profiles could be divided into two classes, those of dendritic origin (presynaptic dendrite boutons) and those of axonal origin. The NOS labelled axon terminals varied in size and packing density and were principally located in the extra-glomerular neuropil. These boutons presented a range of morphologies and it was not possible to determine the probable source based on morphological criteria. The NOS immunoreactive presynaptic dendrite boutons were found both within and outside glomeruli and established both pre- and post-synaptic relationships with other elements. Post-embedding GABA immunocytochemistry showed that some NOS immunoreactive axonal boutons and presynaptic dendrites were also immunopositive for GABA. This finding suggests that some of the NOS labelled axonal boutons are of local circuit neuron origin. These results suggest that local circuit neurons in the cat ventrobasal complex might be involved in specific, short range interactions using GABA and longer, more global interactions using nitric oxide.     

Presynaptic inhibition in the vertebrate spinal cord revisited

The present review examines the experimental evidence supporting the existence of central mechanisms able to modulate the synaptic effectiveness of sensory fibers ending in the spinal cord of vertebrates. The first section covers work on the mode of operation and the synaptic mechanisms of presynaptic inhibition, in particular of the presynaptic control involving axo-axonic synapses made by GABAergic interneurons with the terminal arborizations of the afferent fibers. This includes reviewing of the ionic mechanisms involved in the generation of primary afferent depolarization (PAD) by GABAergic synapses, the ultrastructural basis underlying the generation of PAD, the relationship between PAD and presynaptic inhibition, the conduction of action potentials in the terminal arborizations of the afferent fibers, and the modeling of the presynaptic inhibitory synapse. The second section of the review deals with the functional organization of presynaptic inhibition. This includes the segmental and descending presynaptic control of the synaptic effectiveness of group-I and group-II muscle afferents, the evidence dealing with the local character of PAD as well as the differential inhibition of PAD in selected collaterals of individual muscle-spindle afferents by cutaneous and descending inputs. This section also examines observations on the presynaptic modulation of large cutaneous afferents, including the modulation of the synaptic effectiveness of thin myelinated and unmyelinated cutaneous fibers and of visceral afferents, as well as the presynaptic control of the synaptic actions of interneurons and descending tract neurons. The third section deals with the changes in PAD occurring during sleep and fictive locomotion in higher vertebrates and with the changes of presynaptic inhibition in humans during the execution of a variety of voluntary movements. In the final section, we examine the non-synaptic presynaptic modulation of transmitter release, including the possibility that the intraspinal endings of primary afferents also release colocalized peptides in a similar way as in the periphery. The outcome of the studies presently reviewed is that intraspinal terminals of sensory fibers are not hard-wired conductors of the information generated in their peripheral sensory receptors, but dynamic systems that convey information that can be selectively addressed by central mechanisms to specific neuronal targets. This central control of information flow in peripheral afferents appears to play an important role in the generation of integrated movements and processing of sensory information, including nociceptive information. Received: 16 December 1998 / Accepted: 1 June 1999