Further evidence for the topography and connectivity of the corpus callosum: an FMRI study of patients with partial callosal resection.

BACKGROUND AND PURPOSE: This functional MRI study was designed to describe activated fiber topography and trajectories in the corpus callosum (CC) of six patients carrying different degree of partial callosal resection. METHODS: Patients receiving gustatory, tactile, and visual stimulation according to a block-design protocol were scanned in a 1.5 Tesla magnet. Diffusion tensor imaging (DTI) data were also acquired to visualize spared interhemispheric fibers. RESULTS: Taste stimuli evoked bilateral activation of the primary gustatory area in all patients and foci in the anterior CC, when spared. Tactile stimuli to the hand evoked bilateral foci in the primary somatosensory area in patients with an intact posterior callosal body and only contralateral in the other patients. Callosal foci occurred in the CC body, if spared. In patients with an intact splenium central visual stimulation induced bilateral activation of the primary visual area as well as foci in the splenium itself. CONCLUSION: Present data show that interhemispheric fibers linking sensory areas crossed through the CC at the sites where the different sensory stimuli evoked activation foci, and that topography of callosal foci evoked by sensory stimulation in spared CC portions is consistent with that previously observed in subjects with intact CC.

Heterogeneity of glutamatergic and GABAergic release machinery in cerebral cortex: analysis of synaptogyrin, vesicle-associated membrane protein, and syntaxin.

To define whether cortical glutamatergic and GABAergic release machineries can be differentiated on the basis of the nature and amount of proteins they express, we studied the degree of co-localization of synaptogyrin (SGYR) 1 and 3, vesicle-associated membrane protein (VAMP) 1 and 2, syntaxin (STX) 1A and 1B in vesicular glutamate transporter (VGLUT)1-, VGLUT2- and vesicular GABA transporter (VGAT)-positive (+) puncta and synaptic vesicles in the rat cerebral cortex. Co-localization studies showed that SGYR1 and 3 were expressed in about 90% of VGLUT1+, 70% of VGLUT2+ and 80% of VGAT+ puncta; VAMP1 was expressed in approximately 45% of VGLUT1+, 55% of VGLUT2+, and 80% of VGAT+ puncta; VAMP2 in about 95% of VGLUT1+, 75% of VGLUT2+, and 80% of VGAT+ puncta; STX1A in about 65% of VGLUT1+, 30% of VGLUT2+, and 3% of VGAT+ puncta, and STX1B in approximately 45% of VGLUT1+, 35% of VGLUT2+, and 70% of VGAT+ puncta. Immunoisolation studies showed that while STX1A was completely segregated and virtually absent from VGAT synaptic vesicles, STX1B, VAMP1/VAMP2, SGYR1/SGYR3 showed a similar pattern with the highest expression in VGLUT1 immunoisolated vesicles and the lowest in VGAT immunoisolated vesicles. Moreover, we studied the localization of STX1B at the electron microscope and found that a population of axon terminals forming symmetric synapses were STX1B-positive.These results extend our previous observations on the differential expression of presynaptic proteins involved in neurotransmitter release in GABAergic and glutamatergic terminals and indicate that heterogeneity of glutamatergic and GABAergic release machinery can be contributed by both the presence or absence of a given protein in a nerve terminal and the amount of protein expressed by synaptic vesicles.

Heterogeneity of glutamatergic and GABAergic release machinery in cerebral cortex.

We investigated whether cortical glutamatergic and GABAergic release machineries can be differentiated on the basis of the proteins they express, by studying the degree of co-localization of synapsin (SYN) I and II, synaptophysin (SYP) I and II, synaptosomal-associated protein (SNAP)-25 and SNAP-23 in vesicular glutamate transporter (VGLUT) 1-, VGLUT2- and vesicular GABA transporter (VGAT)-positive (+) puncta in the rat cerebral cortex. Co-localization studies showed that SYNI and II were expressed in approximately 90% of VGLUT1+, approximately 30% of VGLUT2+ and 30-50% of VGAT+ puncta; SYPI was expressed in approximately 95% of VGLUT1+, 30% of VGLUT2+, and 45% of VGAT+ puncta; SYPII in approximately 7% of VGLUT1+, 3% of VGLUT2+, and 20% of VGAT+ puncta; SNAP-25 in approximately 94% of VGLUT1+, 5% of VGLUT2+, and 1% of VGAT+ puncta, and SNAP-23 in approximately 3% of VGLUT1+, 86% of VGLUT2+, and 22% of VGAT+ puncta. Since SYPI, which is considered ubiquitous, was expressed in about half of GABAergic axon terminals, we studied its localization electron microscopically and in immunoisolated synaptic vesicles: these studies showed that approximately 30% of axon terminals forming symmetric synapses were SYPI-negative, and that immunoisolated VGAT-positive synaptic vesicles were relatively depleted of SYPI as compared with VGLUT1+ vesicles. Overall, the present investigation shows that in the cerebral cortex of rats distinct presynaptic proteins involved in neurotransmitter release are differentially expressed in GABAergic and in the two major types of glutamatergic axon terminals in the cerebral cortex of rats.

The glial glutamate transporter GLT-1 is localized both in the vicinity of and at distance from axon terminals in the rat cerebral cortex.

Glutamate transporter-1 (GLT-1) is responsible for the largest proportion of glutamate transport in the brain and the density of GLT-1 molecules inserted in the plasma membrane is highest in regions of high demand. Previous electron microscopic studies in the hippocampus and cerebellum have shown that GLT-1 is concentrated both in the vicinity of and at considerable distance from the synaptic cleft [Chaudry et al., Neuron 15 (1995) 711-721], but little is known about its distribution in the neocortex. We therefore studied the spatial relationships between elements expressing the presynaptic marker synaptophysin and those containing GLT-1 in the rat cerebral cortex using confocal microscopy. Preliminary studies confirmed that GLT-1 positive puncta were exclusively astrocytic processes; moreover, they showed that in most cases GLT-1 positive processes either completely surrounded asymmetric synapses or had no apparent relationship with synapses; occasionally, they were apposed to terminals containing pleomorphic vesicles. In sections double-labeled for GLT-1 and synaptophysin, codistribution analysis revealed that 61.2% of pixels detecting fluorescent emission for GLT-1 immunoreactivity overlapped with pixels detecting synaptophysin. The percentages of GLT-1/synaptophysin codistribution were significantly different from controls. In sections double-labeled for GLT-1 and the vesicular GABA transporter, codistribution analysis revealed that 27% of pixels detecting GLT-1 overlapped with those revealing the vesicular GABA transporter.The remarkable 'synaptic' localization of GLT-1 provides anatomical support for the hypothesis that in the cerebral cortex GLT-1 contributes to shaping fast, point-to-point, excitatory synaptic transmission. Moreover, the considerable fraction of GLT-1 immunoreactivity localized at sites distant from axon terminals supports the notion that glutamate spillout occurs also in the intact brain and suggests that 'extrasynaptic' GLT-1 regulates the diffusion of glutamate escaped from the cleft.

Glutamate transporter EAAC1 in the cat periaqueductal gray matter.

The neuron-specific glutamate (Glu) transporter, excitatory amino acid carrier 1 (EAAC1), plays an important role in regulating Glu levels in the synaptic cleft. Using a specific EAAC1 monoclonal antibody, we investigated its regional distribution and ultrastructural localization in cat periaqueductal gray matter. In light microscopy EAAC1 immunoreactivity was randomly distributed to neurons and punctate structures. In electron microscopy, it was observed in the soma of many neurons, dendrites, in a discrete number of axon terminals, in ependymal cells and in few distal astrocytic processes. EAAC1 is thus not restricted to neurons, but could play an important role in glial cells. Moreover, EAAC1 protein could participate both in regulating the action of the Glu released at synaptic level and in other aspects of Glu metabolism.

gamma-Aminobutyric acid transporters in the cat periaqueductal gray: a light and electron microscopic immunocytochemical study.

The gamma-aminobutyric acid (GABA) plasma membrane transporters (GATs) mediate GABA uptake into presynaptic axon terminals and glial processes, thus contributing to the regulation of the magnitude and duration of the action of GABA at the synaptic cleft. The aim of the present study was to investigate the expression of three high-affinity GABA transporters (GAT-1, GAT-2, and GAT-3) in the periaqueductal gray matter (PAG) of adult cats by using immunocytochemistry with affinity-purified antibodies. Light microscopic observations revealed GAT-1 immunoreactivity in punctate structures, particularly dense in the lateral portion of the dorsolateral PAG column. Weak GAT-2-immunopositive puncta were homogeneously distributed in the PAG. GAT-3 immunoreactivity was detected in each column of the PAG but was more intense in the dorsolateral PAG column and around the aqueduct. Electron microscopic studies showed GAT-1 immunoreactivity in distal astroglial processes, in unmyelinated and small myelinated axons, and in axon terminals making symmetric synapses on both PAG neurons and dendrites. GAT-2 immunoreactivity was present mostly in the form of patches of different sizes in the cytoplasm of neuronal elements like the perikarya and dendrites of PAG neurons, in myelinated and unmyelinated axons, and in the axon terminals forming both symmetric and asymmetric synapses. Labeling was also observed in nonneuronal elements. Astrocytic cell bodies and their distal processes as well as the ependymal cells lining the wall of the aqueduct showed patches of GAT-2 immunoreactivity. Electron microscopic observation revealed GAT-3 immunoreactivity exclusively in distal astrocytic processes adjacent to the somata of PAG neurons and in axon terminals making both symmetric and asymmetric synapses. The present results suggest that three types of termination systems of GABAergic transmission are present in the cat periaqueductal gray matter.

Neuronal, glial, and epithelial localization of gamma-aminobutyric acid transporter 2, a high-affinity gamma-aminobutyric acid plasma membrane transporter, in the cerebral cortex and neighboring structures.

Neuronal and glial high-affinity Na+/Cl(-)-dependent plasma membrane gamma-aminobutyric acid (GABA) transporters (GATs) contribute to regulating neuronal function. We investigated in the cerebral cortex and neighboring regions of adult rats the distribution and cellular localization of the GABA transporter GAT-2 by immunocytochemistry with affinity-purified polyclonal antibodies that react monospecifically with a protein of 82 kDa. Conventional and confocal laser-scanning light microscopic studies revealed intense GAT-2 immunoreactivity (ir) in the leptomeninges, choroid plexus, and ependyma. Weak GAT-2 immunoreactivity also was observed in the cortical parenchyma, where it was localized to puncta of different sizes scattered throughout the radial extension of the neocortex and to few cell bodies. In sections double-labeled with GAT-2 and glial fibrillary acidic protein (GFAP) antibodies, some GAT-2-positive profiles also were GFAP positive. Ultrastructural studies showed GAT-2 immunoreactivity mostly in patches of varying sizes scattered in the cytoplasm of neuronal and nonneuronal elements: GAT-2-positive neuronal elements included perikarya, dendrites, and axon terminals forming both symmetric and asymmetric synapses; nonneuronal elements expressing GAT-2 were cells forming the pia and arachnoid mater; astrocytic processes, including glia limitans and perivascular end feet; ependymal cells; and epithelial cells of the choroid plexuses. The widespread cellular expression of GAT-2 suggests that it may have several functional roles in the overall regulation of GABA levels in the brain.

Neuronal and glial localization of NR1 and NR2A/B subunits of the NMDA receptor in the human cerebral cortex.

N-Methyl-D-aspartate (NMDA) receptors play a critical role in many cortical functions and are implicated in several neuropsychiatric diseases. In this study, the cellular expression of the NMDAR1 (NR1) and NMDAR2A and B (NR2A and B) subunits was investigated in the human cerebral cortex by immunocytochemistry with antibodies that recognize the NR1 or the NR2A and B subunits of the NMDA receptor. In frontal (areas 10 and 46) and temporal (area 21) association cortices and the cingulofrontal transition cortex (area 32), NR1 and NR2A/B immunoreactivity (ir) were similar and were localized to numerous neurons in all cortical layers. NR1- and NR2A/B-positive neurons were mostly pyramidal cells, but some nonpyramidal neurons were also labeled. Electron-microscopic observations showed that NR1 and NR2A/B ir were similar. In all cases, labeling of dendrites and dendritic spines was intense. In addition, both NR1 and NR2A/B were consistently found in the axoplasm of some axon terminals and in distal astrocytic processes. This investigation revealed that numerous NMDA receptors are localized to dendritic spines, and that they are also localized to axon terminals and astrocytic processes. These findings suggest that the effects of cortical NMDA activation in the human cortex do not depend exclusively on the opening of NMDA channels located at postsynaptic sites, and that the localization of NMDA receptors is similar in a variety of mammalian species.

GABA transporter-1 (GAT-1) immunoreactivity in the cat periaqueductal gray matter.

Using light- and electron-microscopic immunocytochemistry, we investigated the regional distribution and the ultrastructural localization of GAT-1, a prominent GABA transporter, in the cat PAG. Light microscopic observations indicate that GAT-1-immunoreactive elements are particularly dense in PAG-DL and form a pair of longitudinal columns extending in the intermediate region of this structure. At electron-microscopic level, GAT-1 immunoreactivity was present in axon terminals forming symmetric synapses and in the distal processes of astroglial cells. These data further confirm the existence of longitudinal columns within PAG. They also indicate that GAT-1 could influence the action of GABA on its receptors, probably regulating the magnitude and duration of GABA's synaptic action on PAG neurons, and suggest that astrocytes may play an important role in this process.

Immunocytochemical localization of substance P receptor in rat periaqueductal gray matter: a light and electron microscopic study.

The distribution of substance P receptor (SPR) protein in the rat periaqueductal gray matter (PAG) was investigated with a polyclonal antibody in the four subdivisions obtained by cytochrome-oxidase histochemistry (Co-hi). At light microscopic analysis, immunoreactivity appeared particularly dense in the dorsal subdivision of the PAG, was less intense in the other subdivisions, and formed several longitudinally organized columns. SPR-like immunoreactivity (SP(R-i)) was localized mostly to cell bodies and dendrites of small and medium-sized neurons, which constituted about 6% of the total neuronal population of the PAG. At the electron microscopic level, SP(R-i) could be observed on postsynaptic as well as on nonsynaptic regions of both cell bodies and dendrites. A small proportion of axons (4.2%) and axon terminals (5.3%) showed SP(R-i), the majority of labeled axon terminals, amounting to about 70% of synapsing elements, formed asymmetric synapses with dendrites. Rare astroglial processes displaying SP(R-i) were also observed scattered throughout the neuropil of all PAG subdivisions. Our observations suggest that 1) also in the PAG, SP may act in a diffuse, nonsynaptic manner, probably on targets that are distant from its sites of release; and 2) SP may modulate excitatory neurotransmission acting presynaptically on those labeled axons that form asymmetric synapses.

Glutamate-positive neurons and terminals in the cat periaqueductal gray matter (PAG): a light and electron microscopic immunocytochemical study.

The morphology, distribution, proportion, size, and synaptic organization of periaqueductal gray matter neurons labeled with immunocytochemical techniques by an anti-glutamate (Glu) polyclonal serum were investigated in six adult cats (PAG-GLU 1-6). At the light microscopic level, numerous Glu-positive neurons were found throughout each subdivision of the periaqueductal gray matter. Their proportion and size, calculated in semi-thin sections (1-microm-thick), varied slightly among the subdivisions of the periaqueductal gray matter. The morphology of Glu-positive neurons was similar to that of the multipolar, triangular, and fusiform cells described in previous Golgi studies. Numerous puncta, interpreted as dendrites, axons, and axon terminals were also present in all subdivisions without preferential distribution. At the electron microscopic level, all synaptic contacts made by Glu-positive axon terminals were of the asymmetric type, but not all presynaptic elements making asymmetric synapses were labeled. The vast majority of postsynaptic elements contacted by Glu-positive axon terminals were labeled and unlabeled dendrites. The present results describe for the first time the presence of both Glu-positive neurons and terminals in the feline periaqueductal gray matter and provide further evidence that Glu is the probable neurotransmitter of numerous excitatory neurons of this structure.

Laminar pattern of termination of the ipsilateral cortical projection from SII to SI in cats.

The present light and electron microscopic experiments were carried out on the first somatic sensory area (SI) of cats to determine the laminar distribution of axon terminals from the ipsilateral second somatic sensory area (SII) and to identify the types of synapses between these terminals and the neuronal elements of SI. Phaseolus vulgaris-leucoagglutinin (PHA-L) was iontophoretically injected into multiple sites and at different cortical depths of the forepaw representation zone of SII. Fixed brain blocks containing the injected SII and ipsilateral SI were cut into slices and processed immunocytochemically to stain PHA-L-filled fibers and terminals. Light microscopic examination of SI revealed patches of anterograde labeling in the forepaw representation zone, concentrated mainly in supragranular layers. In these layers, thin immunolabeled fibers branched extensively and formed a dense plexus that was more prominent in layers II and I. Conversely, the infragranular layers contained fragments of vertically oriented thick fibers that rarely emitted axon collaterals. PHA-L-labeled axons had numerous swellings along their course, interpreted as boutons en passant, and stalked boutons. Of 19,661 labeled terminals (17,833 beads and 1,828 stalked boutons), 84.74% were observed in supragranular layers, with the highest concentration in layer II (33.15%) and lower in layers I (26.27%) and III (25.30%). The proportion of terminals was lower in layers IV (6.49%) and V (5.45%) and lowest in layer VI (3.32%). These counts also showed that boutons en passant were the majority (90.70%) and stalked boutons, the minority (9.30%). The ratio of these two types of presynaptic specializations was similar (9:1) in all six layers. Electron microscopic examination of the labeled regions of SI showed that both axon swellings and stalked boutons formed synapses of the asymmetric type with SI neuronal elements. The majority (85.37%) of a sample of 130 labeled terminals synapsed on SI neurons in layers I-III. The identified postsynaptic profiles were dendritic spines (61.11%) or medium-sized and small dendrites (38.89%). These results are discussed in relation to those of a companion study on the laminar pattern of the projection from SI to SII of cats (P. Barbaresi, A. Minelli, and T. Manzoni, 1994, J. Comp. Neurol. 343:582-596). Based on the anatomical organization of these reciprocal connections, there seems to be no clear hierarchicalal relationship between SI and SII in cats.

Commissural connections of the cat periaqueductal gray matter studied with anterograde and retrograde tract-tracing techniques.

The commissural connections of the periaqueductal gray matter were investigated by light and electron microscopy by using the anterograde tracer Phaseolus vulgaris leucoagglutinin and the retrograde tracer horseradish peroxidase. In the first group of seven animals (1-7), single injections of Phaseolus vulgaris leucoagglutinin were performed iontophoretically (4.5 microA for 30 min) into various subdivisions of the periaqueductal gray matter. On light microscopic examination, injection sites were characterized by several immunolabeled neurons of different sizes and morphology, with the cytoplasm, nucleus and neuronal processes intensely stained. Many labeled fibers turned from injection sites toward all contralateral periaqueductal gray matter subdivisions, but anterograde labeling was densest in the regions homotopic to those injected. Commissural fibers bore along their course many en passant boutons of different sizes and morphology, and gave off spine-like processes, at the end of which one terminal bouton was observed. Labeled fibers branched into numerous collaterals which ended in a terminal array of 10-20 en passant and en grappe boutons. At the electron microscopic level, commissural axons were observed in close proximity to the cytoplasmic membranes of cells. Axon terminals formed symmetric or asymmetric synapses mainly on dendritic shafts of neurons and rarely on vesicle-containing profiles. Horseradish peroxidase experiments were carried out in four cats (1-4). The tracer was injected iontophoretically into different regions of the periaqueductal gray matter of three cats (1-3). Retrogradely labeled neurons giving rise to commissural connections had a morphology similar to that of polygonal, triangular and fusiform cells described in previous Golgi studies. The perikaryal cross-sectional area of commissural neurons was smaller than that of neurons projecting outside the periaqueductal gray matter (mean value of commissural neurons 149.77 microns 2 vs 261.19 microns 2 for projecting neurons), which were retrogradely labeled by pressure-injecting horseradish peroxidase into several targets of periaqueductal gray matter (4). Moreover, since the distribution of sizes of the two populations of the periaqueductal gray matter overlapped in the range of 90-300 microns 2, a considerable number of projecting neurons were as small as commissural neurons. The present results suggest that commissural fibers could reciprocally connect zones of the periaqueductal gray matter with similar functions, and originate from small and medium-sized neurons, some of which are also projecting neurons.

Topographical relations between ipsilateral cortical afferents and callosal neurons in the second somatic sensory area of cats.

Experiments were carried out on the second somatic sensory area (SII) of cats to study 1) the laminar distribution of axon terminals from the ipsilateral first somatic sensory cortex (SI); and 2) the topographical relations between their terminal field and the callosal neurons projecting to the contralateral homotopic cortex. To label simultaneously in SII both ipsilateral cortical afferents and callosal cells, cats were given iontophoretic injections of Phaseolus vulgaris-leucoagglutinin (PHA-L) in the forepaw zone of ipsilateral SI, and pressure injections of horseradish peroxidase (HRP) in the same zone of contralateral SII. The possibility that ipsilateral cortical axon terminals synapse callosal neurons was investigated with the electron microscope by combining lesion-induced degeneration with retrograde HRP labelling. Fibers and terminations immunolabelled with PHA-L from ipsilateral SI were distributed in SII in a typical patchy pattern and were mostly concentrated in supragranular layers. Labelled fibers formed a very dense plexus in layer III and ramified densely also in layers I and II. Labelled axon terminals were both en passant and single-stalked boutons. Counts of 8,303 PHA-L-labelled terminals of either type showed that 82.40% were in supragranular layers. The highest concentration was in layer III (43.99%), followed by layers II (30.32%) and I (8.09%). The remaining terminals were distributed among layers IV (6.96%), V (4.93%), and VI (5.68%). The same region of SII containing anterogradely labelled axons and terminals also contained numerous neurons retrogradely labelled with HRP from contralateral SII. Callosal projection neurons were pyramidal, dwelt mainly in layer III, and were distributed tangentially in periodic patches. Patches of anterograde and retrograde labelling either interdigitated or overlapped both areally and laminarly. In the zones of overlap, numerous PHA-L-labelled axon terminals were seen in close apposition to HRP-labelled pyramidal cell dendrites. Combined HRP-electron microscopic degeneration experiments showed that in SII axon terminals from ipsilateral SI form asymmetric synapses with HRP-labelled dendrites and dendritic spines pertaining to callosal projection neurons. These results are discussed in relation to the layering and function of the SI to SII projection, and to the evidence that SII neurons projecting to the homotopic area of the contralateral hemisphere have direct access to the sensory information transmitted from ipsilateral SI.

Thalamic connections of the second somatic sensory area in cats studied with anterograde and retrograde tract-tracing techniques.

The thalamic connections of the second somatosensory area in the anterior ectosylvian gyrus of cats have been investigated using the retrograde tracer horseradish peroxidase and the anterograde tracer Phaseolus vulgaris leucoagglutinin. Horseradish peroxidase was injected iontophoretically in several somatotopic zones of the second somatosensory area map of six cats. Sites of horseradish peroxidase delivery were identified preliminarily by recording with microelectrodes the responses of neurons to skin stimulation. Phaseolus vulgaris leucoagglutinin was iontophoretically injected within the ventrobasal complex (one cat) or in the posterior complex (one cat). Horseradish peroxidase injections into cytoarchitectonic area SII retrogradely labeled neurons in the ipsilateral ventrobasal complex and in the posterior complex. Counts of labeled neurons from the ipsilateral thalamus showed that the overwhelming majority of horseradish peroxidase-labeled neurons were in the ventrobasal complex (96.3-96.9%) and few were in the posterior complex (3.1-3.7%). Neurons labeled in the ventrobasal complex were observed throughout the anteroposterior extent of the nucleus, while their mediolateral distribution varied with the site of horseradish peroxidase delivery in the body map of the second somatosensory area, which indicates that the projections from the ventrobasal complex to the second somatosensory area are somatotopically organized. In the cat in which the horseradish peroxidase injection involved both the second somatosensory area proper and the second somatosensory area medial, which lies in the lower bank of suprasylvian sulcus, labeled neurons were almost as numerous in the ventrobasal complex as in the posterior complex. Phaseolus vulgaris leucoagglutinin injected in the ventrobasal complex anterogradely labeled thalamocortical fibers in the ipsilateral anterior ectosylvian gyrus. In this case, patches of labeled fibers and terminals were distributed exclusively within the cytoarchitectonic borders of the second somatosensory area proper. Labeled terminals were numerous in layer IV and lower layer III, but terminal boutons and fibers with axonal swellings, probably forming synapses en passant, were frequently observed also in layers VI and I. Injection of Phaseolus vulgaris leucoagglutinin in the posterior complex labeled thalamocortical fibers in two distinct regions in the ipsilateral anterior ectosylvian gyrus, one lying laterally and the other medially, which correspond, respectively, to the fourth somatosensory area and the second somatosensory area medial. In both areas the densest plexus of labeled fibers and axon terminals was in layer IV and lower layer III, but numerous labeled fibers and terminals were also observed in layer I. In this case, only rare fragments of labeled fibers were present in second somatosensory area proper, but no labeled terminals could be observed.

Organization of claustro-cortical projections to the primary somatosensory area of primates.

Different fluorescent dyes were injected in the face (S1fa) and hand (S1hn) representations of the primary somatosensory cortex, involving both areas 3b and 1. Claustral neurons labeled by either S1fa or S1hn were divided in two populations. One population was located in the dorsal part of the nucleus: neurons labeled from S1fa were placed laterally to those labeled from S1hn. The second population was located more ventrally, with a rostro-caudal distribution of S1fa vs S1hn neurons. These findings demonstrate the existence of ordered and possibly multiple somatosensory representations in the monkey claustrum.

Matching of receptive fields in the association projections from SI to SII of cats.

Anatomical and electrophysiological experiments were performed on cats to investigate the pattern of divergence and convergence in the association projections from the first (SI) to the second (SII) somatic sensory cortex and to ascertain whether diverging and converging fibre components from SI have receptive fields (RFs) matching those of target neurons in SII. In the first group of six cats, a single deposit of horseradish peroxidase (HRP) was iontophoretically placed (2-4 microA for 20 minutes) into an electrophysiologically identified site of the SII map: the digit (3 cats), forepaw (2 cats), and arm (1 cat) zones. The forelimb representation in ipsilateral SI was subsequently explored with microelectrodes and RFs from small clusters of neurons systematically mapped. Planar maps of this area were reconstructed with the aid of a computer from serial sections, to correlate on the tangential plane the topographical distribution of retrogradely labelled association neurons with the physiological map of the forelimb. Since diverging projections were observed from a zone of SI to multiple zones of SII, double-labelling experiments were carried out in a second group of three cats, in which two retrograde fluorescent dyes (diamidino yellow and fast blue) were injected by pressure into two different sites of the SII map, to ascertain whether SI sends diverging projections by branching axons. HRP injections in SII retrogradely labelled a discrete number of association neurons in SI. Their distribution area was several tens of times wider than that covered by the injection site. This suggests that a remarkable amount of divergence and convergence exists in the association projections from SI to SII. Despite the substantial difference in the extent of the injected and labelled areas, RFs of afferent and target neurons corresponded closely. Injections covering a small region within a single digit zone of SII labelled neurons throughout the entire representation of the same digit in SI, while neurons labelled in somatotopically inappropriate zones were rare. RFs mapped from several sites of the labelled region in SI were individually smaller than the RF mapped from the injection site in SII, but the overall size of afferent RFs encompassed that of target neurons. Divergence and convergence in the SI projections to SII zones representing more proximal portions of the forelimb may be even greater since HRP injections in the forepaw and arm zones of SII labelled a number of neurons also in the digit zone of SI, providing the RFs mapped from the injection sites were sufficiently wide to include the digits.(ABSTRACT TRUNCATED AT 400 WORDS)

Callosal connections of the somatic sensory areas II and IV in the cat.

The homotopic and heterotopic callosal connections in the forelimb representations of the second (SII) and fourth (SIV) somatic sensory areas of cats were investigated by means of the axonal transport of horseradish peroxidase (HRP) in conjunction with microelectrode recording. The tracer was injected in the electrophysiologically identified hand and/or digit zone of SII (six cats) or SIV (four cats). The homotopic area in the contralateral hemisphere was explored with microelectrodes in five animals (three injected in SII and two in SIV) to map neuronal receptive fields. The aim was to correlate in the same experimental case the topography of labelled callosal neurons with the physiological map of the forelimb. Labelled cells and recording sites were plotted on planar maps reconstructed with the aid of a computer from serial coronal sections from the anterior ectosylvian gyrus. After SII injections, labelled callosal neurons were observed throughout the forelimb representation in the contralateral area, but in the tangential plane their distribution was uneven. Each somatotopic zone composing the forelimb map, that is, the arm, hand, and digit zones, contained several subzones in which callosal neurons were either dense or rare. Microelectrode explorations showed that receptive fields mapped from callosal and relatively acallosal subzones representing the same body part were similar in extent and location. After SIV injections, labelled callosal neurons were observed throughout the forelimb and proximal body representation of the contralateral area. Although slight regional variations in the density of labelled cells were apparent, no subzones bare of callosal labelling were observed in SIV. In both SII and SIV, callosal neurons were concentrated mainly in layer III, but a significant number was also evident in the infragranular layers. After HRP injections in the digit zone of SII or SIV, labelled cell bodies were also observed in heterotopic areas of the contralateral hemisphere. Most of these neurons were clustered in the medial bank of the coronal sulcus and in two other heterotopic cortical regions lying, respectively, in the anterior suprasylvian sulcus and in the lateral branch of the ansate sulcus. Some callosal cells interconnecting SII and SIV were also labelled. The results show that the distal forelimb zones in SII and SIV are callosally connected with the respective homotopic zones and with several somatosensory fields located heterotopically in the contralateral hemisphere.

Glutamate decarboxylase-immunoreactive neurons and terminals in the periaqueductal gray of the rat.

The periaqueductal gray of 5 rats was processed for immunocytochemistry using an antiserum to glutamate decarboxylase. In both colchicine-pretreated (4 rats) and untreated (1 rat) animals, glutamate decarboxylase-positive cell bodies were present in all periaqueductal gray subdivisions, especially in the dorsal and ventrolateral subdivision. The perikaryal cross-sectional area of labelled neurons was smaller than that of periaqueductal gray projecting neurons retrogradely labelled with horseradish peroxidase in separate experiments. The morphology of glutamate decarboxylase-containing neurons resembled that of small polygonal, triangular and fusiform cells described in previous Golgi studies. Glutamate decarboxylase immunoreactivity was also observed in a large number of terminal-like structures, most of which were distributed close to the somata and dendrites of both glutamate decarboxylate-positive and -negative neurons. At all rostrocaudal levels the highest concentration of these elements was observed around the aqueduct. These results suggest that two sub-populations of neurons are present in the periaqueductal gray of rats, one consisting of small-sized glutamate decarboxylase-positive neurons (intrinsic neurons) and the other of large-sized glutamate decarboxylase-negative neurons (projecting neurons). Intrinsic circuits could be present between glutamate decarboxylase-positive and -negative neurons and between glutamate decarboxylase-positive neurons.