Ultrastructure of cholinergic synaptic terminals in the thalamic anteroventral,ventroposterior, and dorsal lateral geniculate nuclei of the rat

The principal relay nuclei of the thalamus receive their cholinergic innervation from two midbrain cholinergic groups: the pedunculopontine tegmental nucleus and the laterodorsal tegmental nucleus. The different thalamic nuclei exhibit populations of cholinergic axons which vary in density and morphology when examined at the light microscopic level. However, the ultrastructure of the cholinergic terminals in different thalamic nuclei has not been described. This study was undertaken to confirm that synaptic contacts are formed by cholinergic axons in several principal thalamic relay nuclei, to describe their ultrastructural morphology, and to identify the types of postsynaptic elements contacted by cholinergic synaptic terminals. The thalamic nuclei examined in this study are the dorsal lateral geniculate nucleus, ventroposteromedial nucleus, ventroposterolateral nucleus, and anteroventral nucleus. Our results confirm that cholinergic axons form synaptic terminals in these thalamic nuclei. Cholinergic synaptic terminals contact structures outside the characteristic synaptic glomeruli, are never postsynaptic, and have morphologies and postsynaptic targets which differ among the thalamic nuclei. In the ventroposterior nuclei, cholinergic terminals form asymmetric synaptic contacts onto larger dendrites in the extraglomerular neuropil. In the anteroventral nucleus, cholinergic terminals form both symmetric and asymmetric synaptic contacts onto dendrites and somata. Cholinergic terminals in the anteroventral nucleus are larger than those in other nuclei. In the dorsal lateral geniculate nucleus, cholinergic terminals contact both somata and dendrites in the extraglomerular neuropil, but the synaptic contacts in this nucleus are symmetric in morphology.     

Light and electron microscopic evidence for a GABAergic projection from the caudal basal forebrain to the thalamic reticular nucleus in rats.

Neurons in the magnocellular nucleus of the caudal basal forebrain extend an axonal projection which arborizes within the reticular nucleus of the thalamus. The present study addresses the ultrastructure and neurochemistry of this projection in rats. Many labeled terminals are apparent within the thalamic reticular nucleus following Phaseolus vulgaris leucoagglutinin injections into the caudal basal nucleus; anterogradely labeled axon terminals most commonly contact both somata and dendrites of reticular nucleus neurons with symmetric membrane specializations. Thus, the majority of the labeled terminals examined contrast with choline acetyltransferase positive terminals which have been previously identified as contacting dendrites and forming asymmetric synapses within this nucleus. Many of the neurons within the caudal basal nucleus which are retrogradely labeled following tracer injections into the thalamic reticular nucleus are gamma-aminobutyric acid (GABA) immunoreactive. In addition, following injections of Phaseolus vulgaris leucoagglutinin or fluoro-ruby into the caudal basal forebrain, some of the labeled axonal swellings and boutons within the thalamic reticular nucleus also contain glutamic acid decarboxylase. These results indicate that a significant component of the projection is GABAergic. These anatomical observations suggest that the projection from the caudal basal nucleus onto the thalamic reticular nucleus could facilitate the relay of information through the dorsal thalamus by inhibiting reticular nucleus neurons, and thus, in turn, disinhibiting thalamic relay neurons.     

Morphology and connectivity of parabrachial and cortical inputs to gustatory thalamus in rats

The ventroposterior medialis parvocellularis (VPMpc) nucleus of the thalamus, the thalamic relay nucleus for gustatory sensation, receives primary input from the parabrachial nucleus, and projects to the insular cortex. To reveal the unique properties of the gustatory thalamus in comparison with archetypical sensory relay nuclei, this study examines the morphology of synaptic circuitry in the VPMpc, focusing on parabrachiothalamic driver input and corticothalamic feedback. Anterogradely visualized parabrachiothalamic fibers in the VPMpc bear large swellings. At electron microscope resolution, parabrachiothalamic axons are myelinated and make large boutons, forming multiple asymmetric, adherent, and perforated synapses onto large‐caliber dendrites and dendrite initial segments. Labeled boutons contain dense‐core vesicles, and they resemble a population of terminals within the VPMpc containing calcitonin gene‐related peptide. As is typical of primary inputs to other thalamic nuclei, parabrachiothalamic terminals are over five times larger than other inputs, while constituting only 2% of all synapses. Glomeruli and triadic arrangements, characteristic features of other sensory thalamic nuclei, are not encountered. As revealed by anterograde tracer injections into the insular cortex, corticothalamic projections in the VPMpc form a dense network of fine fibers bearing small boutons. Corticothalamic terminals within the VPMpc were also observed to synapse on cells that were retrogradely filled from the same injections. The results constitute an initial survey describing unique anatomical properties of the rodent gustatory thalamus. J. Comp. Neurol. 523:139–161, 2015. © 2014 Wiley Periodicals, Inc.     

Synaptic organization of cholinergic neurons in the monkey neostriatum

Cholinergic neurons in the monkey neostriatum were examined at the light and electron microscopic level by immunohistochemical methods in order to localize choline acetyltransferase (ChAT), the synthesizing enzyme for acetylcholine. At the light microscopic level a sparse distribution of cholinergic neurons was identified throughout the caudate nucleus. Neurons had large (25-30 microns) somata, eccentric invaginated nuclei, primary dendrites of unequal diameters, and varicosities on distal dendritic branches. Ultrastructural study showed that the cholinergic cells had a cytoplasm abundant in organelles. Within dendritic branches, mitochondria and cisternae were localized primarily to varicosities. Synaptic inputs were distributed mostly to the dendrites and at least four types that formed symmetric or asymmetric synapses were observed. Immunoreactive fibers were relatively numerous within the neuropil and exhibited small diameters (0.1-0.15) micron) and swellings at frequent intervals. Cholinergic boutons that formed synapses were compared to unlabeled terminals making asymmetric synapses with dendritic spines. Results showed that ChAT-positive axons had significantly smaller cross-sectional areas, shorter synaptic junctions, and a higher density and surface area of mitochondria than the unlabeled boutons. Cholinergic axons formed symmetric synapses mostly with dendritic spines (53%) and the shafts of unlabeled primary and distal dendrites (37%). A relatively small proportion of the boutons contacted axon initial segments (1%) and cell bodies (9%) that included medium-sized neurons with unindented (spiny) and indented (aspiny) nuclei. The majority of dendritic spines contacted by cholinergic axons were also postsynaptic to unlabeled boutons forming asymmetric synapses. The results suggest that cholinergic neurons in the primary neostriatum belong to a single morphological class corresponding to the large aspiny (type II) interneuron identified in previous Golgi studies. Present results along with earlier Golgi-electron microscopic observations from this laboratory suggest that neostriatal cholinergic cells integrate many sources of intrinsic and extrinsic inputs. The observed convergence of ChAT-immunoreactive boutons and unlabeled axons onto the same dendritic spines suggests that intrinsic cholinergic axons modulate extrinsic inputs onto neostriatal spiny neurons at postsynaptic sites close to the site of afferent input.     

A description of the GABAergic neurons and axon terminals in the motor nuclei of the cat thalamus

The GABA neurons and their processes in the cat motor thalamic nuclei were identified and studied with glutamic acid decarboxylase (GAD) immunocytochemistry at both the light and electron microscopic levels. The three nuclei that comprise the motor thalamus, ventral anterior (VA), ventral medial (VM), and ventral lateral (VL), each displayed a characteristic distribution pattern of GAD-positive structures that was consistent with their afferent and intrinsic neuronal organization. All three thalamic nuclei displayed a population of small, GAD-positive cells the dendrites of which contained synaptic vesicles and participated in complex synaptic arrays such as serial synapses, triads, and glomeruli. Based on their ultrastructural features, these GAD-containing cells were identified as local circuit neurons. In contrast, the larger, GAD-negative cells were presumed to be the thalamocortical projection neurons. The axons of GAD-positive local circuit neurons could not be identified in these preparations. The number of GAD-positive dendrites in the neuropil was different for the three thalamic nuclei. In the VA and VM, the GAD-positive dendrites were numerous and formed symmetric synapses with dendrites of GAD-negative cells, mainly in association with corticothalamic boutons. Within VL, the GAD-containing dendrites were more numerous than in VA and VM and formed synapses at influential locations on presumed thalamocortical projection neurons, such as bases of primary dendrites, and bifurcation sites of primary and secondary dendrites. The VA and anterolateral VM nuclei that receive inhibitory GABAergic afferents from the entopeduncular nucleus and substantia nigra contained the highest concentration of large GAD-positive axon terminals. These boutons contained pleomorphic vesicles and numerous mitochondria and formed symmetric synapses and multiple puncta adherentes with dendrites and somata of presumed thalamocortical projection neurons. The size, ultrastructural features, and distribution of these GAD-positive boutons were similar to those features described for basal ganglia terminals in the motor thalamus of the cat. In addition, similar large-size GAD-positive boutons were observed in the medial VM, which receives basal ganglia afferents exclusively from the substantia nigra. The concentration of these terminals in medial VM along the dendrites of thalamocortical projection neurons was much less than that in VA and anterolateral VM. The VL nucleus which lacks basal ganglia input did not contain any large GAD-positive boutons.(ABSTRACT TRUNCATED AT 400 WORDS)     

Organization of connections between the thalamic reticular and the anterior thalamic nuclei in the rat

The thalamic reticular nucleus (TRN) receives topographically organized input from specific sensory nuclei such as the lateral geniculate nucleus. The present study shows this in the rat. However, the pattern of thalamic connections to the limbic reticular sector is unknown. Injecting biocytin into the ventral parts of anteroventral and anteromedial nuclei labeled neurons and axons in the rostral TRN. Filled axon collaterals and their terminals occupied a rectangular sheet in a plane close to the horizontal, and were confined to the inner zone (the medial portion) of the limbic TRN. Retrogradely filled cells were in the middle of the rostral pole in the same horizontal plane, receiving synapses from surrounding labeled boutons. In electron micrographs, thalamic terminals were found to contain round, densely packed synaptic vesicles and formed asymmetrical synapses onto reticular somata and dendritic profiles. Displacing the injection site along the dorso-ventral and rostro-caudal axs in the anterior nuclei produced corresponding shifts of antero- and retrograde labeling within the inner reticular zone. Projections from the dorsal portions of the anterior nuclei did not follow this pattern. Axons from the anterodorsal nucleus occupied the rostralmost tip of both inner and outer zones of the dorsal limbic sector. In accordance with earlier reports, the limbie sector was found to represent several dorsal thalamic nuclei parallel to each other medio-laterally. A topography is described for the limbic reticulo-thalamic connections, suggesting that the rostral TRN is able to influence circumscribed areas of the limbic thalamus. © 1995 Wiley-Liss, Inc.     

GABAergic circuitry in the dorsal division of the cat medial geniculate nucleus

Gamma-aminobutyric acid (GABA) is the main inhibitory neurotransmitter of the thalamus. We used postembedding immunocytochemistry to examine the synaptic organization of GABA-positive profiles in the dorsal superficial subdivision of the cat medial geniculate nucleus (MGN). Three groups of GABA-positive profiles participate in synapses: axon terminals, dendrites, and presynaptic dendrites. The presynaptic GABA-positive terminals target mainly GABA-negative dendrites. The GABA-positive postsynaptic profiles receive input primarily from GABA-negative axons. The results indicate that the synaptic organization of GABA-positive profiles in the dorsal superficial subdivision of the MGN nucleus is very similar to that in other thalamic nuclei.     

Parvalbumin immunoreactivity in the thalamus of guinea pig: Light and electron microscopic correlation with gamma-aminobutyric acid immunoreactivity

The relationship of the calcium binding protein parvalbumin (PV) with gamma-aminobutyric acidergic (GABAergic) neurons differs within different thalamic nuclei and animal species. In this study, the distribution of PV and GABA throughout the thalamus of the guinea pig was investigated at the light microscopic level by using immunoperoxidase methods. Intense PV labelling was found in all the GABAergic neurons of the reticular nucleus and in scattered GABAergic neurons in the anteroventral nucleus, whereas GABAergic interneurons in the ventrobasal and lateral geniculate nuclei were not PV labelled. At the electron microscopic level, preembedding immunuperoxidase for PV was combined with postembedding immunogold for GABA. In the ventrobasal nucleus, four types of profiles were recognized: 1) terminals with flattened vesicles and forming symmetric synapses, which were labelled with both PV and GABA and could therefore be identified as afferents from the reticular nucleus; 2) boutons morphologically similar to presynaptic dendrites of interneurons, labelled only with GABA; 3) large terminals with round vesicles and asymmetric synapses, labelled only with PV, which contacted GABAergic presynaptic dendrites in glomerular arrangements and resembled ascending excitatory afferents; and 4) terminals unlabelled by either antiserum. In the ventrobasal nucleus of the guinea pig a double immunocytochemical labelling permits therefore the differentiation of two populations of GABAergic vesicle-containing profiles, i. e., the terminals originating from reticular nucleus (that are double labelled) and the presynaptic dendrites originating from interneurons (that are GABA-labelled only). The possibility to differentiate GABAergic inputs from the reticular nucleus and from interneurons can shed light to the functional interpretation of synaptic circuits in thalamic sensory nuclei. © 1994 Wiley-Liss, Inc.     

Synaptic distribution of afferents from reticular nucleus in ventroposterior nucleus of cat thalamus

This study was aimed at determining the synaptic circuitry that contributes to the alterations in thalamic function that accompany changes in behavioral states. The somatosensory sector of the thalamic reticular nucleus (RTN) was identified by microelectrode recording in cats and injected with Phaseolus vulgaris-leucoagglutinin (PHA-L). The axons of labeled RTN cells gave rise to collaterals within the RTN and continued into the dorsal thalamus where they terminated predominately in the ventral posterior lateral nucleus (VPL). After small injections in the upper limb representation of RTN, most labeled terminations in VPL were confined to its medial part, suggesting the presence of a topographic organization in the projection. Terminations were concentrated in localized, focal aggregations of boutons. Combined electron microscopic immunocytochemistry, using immunogold labeling for γ-aminobutyric acid (GABA), showed that the PHA-L labeled boutons were GABA-positive terminals that ended in symmetrical synapses. Eighty-two percent of these synapses were on dendrites of relay neurons, 8.5% on dendrites of interneurons, and 9.3% on somata. The terminals of RTN axons form the majority of axon terminals ending in symmetrical synapses in VPL. Their concentration on relay neurons probably underlies the capacity of the RTN projection to reduce background activity of VPL relay neurons in the awake state and to maintain oscillatory behavior of these neurons in drowsiness and early phases of Sleep. © 1995 Wiley-Liss, Inc.     

Projection and innervation patterns of individual thalamic reticular axons in the thalamus of the adult rat: A three-dimensional,graphic, and morphometric analysis

The γ-aminobutyric acid-ergic thalamic reticular nucleus (Rt), which carries matching topographical maps of both the thalamus and cortex and in which constituent cells can synaptically communicate between each other, is the major extrinsic source of thalamic inhibitions and disinhibitions. Whether all the Rt axonal projections into the thalamus are similarly organized and have common projection and innervation patterns are questions of great interest to further our knowledge of the functioning of the Rt. The present study provides architectural and morphometric data of individual, anterogradely labeled axonal arbors that arose from distinct parts of the Rt. One hundred twenty-seven Rt neurons from all regions of Rt were marked juxtacellularly with biocytin or Neurobiotin in urethane-anesthetized adult rats. Eighteen two-dimensional and 14 three-dimensional reconstructions of single tracer-filled Rt neurons were made from serial, frontal, horizontal, or sagittal sections. Both the somatodendritic and axonal fields of tracer-filled Rt cells were mapped in three dimensions and illustrated to provide a complementary stereotaxic reference for future studies. Most marked units projected to a single nucleus of the anterior, dorsal, intralaminar, posterior, or ventral thalamus. Axons emerging from cells in distinct sectors of the Rt projected to distinct nuclei. Within a sector, neurons with separate dendritic fields innervated separate regions either in a single nucleus or into different but functionally related thalamic nuclei. Neurons with an overlap of their dendritic fields gave rise either to overlapping axonal arborizations or, more rarely, to distinct axonal arbors within two different thalamic nuclei implicated in the same function. In rare instances, an Rt axon could project within these two nuclei. Thalamic reticular axons commonly displayed a single well-circumscribed arbor containing a total of about 4,000 ± 1,000 boutons. Every arbor was composed of a dense central core, which encompassed a thalamic volume of 5–63 × 10⁶ μm³ and was made up of patches of maximal innervation density (10 ± 4 boutons/tissue cube of 25 μm each side), surrounded by a sparse component. The metric relationships between the Rt axonal arbors and the dendrites of their target thalamocortical neurons were determined. Both the size and maximal innervation density of the axonal patches were found to fit in with the somatodendritic architecture of the target cells. The Rt axonal projections of adult rats are thus characterized by their (1) well-focused terminal field with a patchy distribution of boutons and (2) parallel organization with a certain degree of divergence. The role of the Rt-mediated thalamic inhibition and disinhibition may be to contrast significant with nonrelevant ongoing thalamocortical information. J. Comp. Neurol. 391:180–203, 1998. © 1998 Wiley-Liss, Inc.     

Noradrenergic innervation of the substantia innominata: a light and electron microscopic analysis of dopamine beta-hydroxylase immunoreactive elements in the rat

Noradrenergic input to the rat substantia innominata (SI) was analyzed in this study by immunocytochemical localization of dopamine beta-hydroxylase (DBH), the synthetic enzyme for noradrenaline. DBH immunoreactive (DBH+) axons ramified extensively within the SI and appeared to be contiguous with the DBH+ terminal fields within the bed nucleus of stria terminalis and the amygdaloid complex. DBH+ axons in the SI exhibited many large boutons en passant and boutons terminaux. These DBH+ boutons appeared much larger than those in the cerebral cortex, hippocampus, and thalamus. Electron microscopic analysis revealed that DBH+ boutons formed asymmetrical synapses with mainly dendrites, but also somata and spines of SI neurons. Dendrites which were postsynaptic to DBH+ boutons also formed synapses with many other unlabeled axon terminals. Since previous studies have shown that dendrites of SI cholinergic neurons formed few synapses, the present result suggests that the noradrenergic influence of SI cholinergic neurons may be mediated mainly by polysynaptic pathways.     

Cholinergic innervation of the human thalamus: dual origin and differential nuclear distribution.

The cholinergic innervation of the human thalamus was studied with antibodies against the enzyme choline acetyltransferase (ChAT) and nerve growth factor receptor (NGFr). Acetylcholinesterase histochemistry was used to delineate nuclear boundaries. All thalamic nuclei displayed ChAT-positive axons and varicosities. Only the medial habenula contained ChAT-positive perikarya. Some intralaminar nuclei (central medial, central lateral, and paracentral), the reticular nucleus, midline nuclei (paraventricular and reuniens), some nuclei associated with the limbic system (anterodorsal nucleus and medially situated patches in the mediodorsal nucleus) and the lateral geniculate nucleus displayed the highest density of ChAT-positive axonal varicosities. The remaining sensory relay nuclei and the nuclei interconnected with the motor and association cortex displayed a lower level of innervation. Immunoreactivity for NGFr was observed in cholinergic neurons of the basal forebrain but not in cholinergic neurons of the upper brainstem. The contribution of basal forebrain afferents to the cholinergic innervation of the human thalamus was therefore studied with the aid of NGFr-immunoreactive axonal staining. The anterior intralaminar nuclei, the reticular nucleus, and medially situated patches in the mediodorsal nucleus displayed a substantial number of NGFr-positive varicose axons, presumably originating in the basal forebrain. Rare NGFr-positive axonal profiles were also seen in many of the other thalamic nuclei. These observations suggest that thalamic nuclei affiliated with limbic structures and with the ascending reticular activating system are likely to be under particularly intense cholinergic influence. While the vast majority of thalamic cholinergic input seems to come from the upper brainstem, the intralaminar and reticular nuclei, and especially medially situated patches within the mediodorsal nucleus also appear to receive substantial cholinergic innervation from the basal forebrain.     

Mesencephalic and diencephalic afferent connections to the thalamic nucleus rotundus in the lizard,Psammodromus algirus

The present work is an analysis of the afferent projections to the thalamic nucleus rotundus in a lizard, both at the light- and electron-microscopic level, using biotinylated dextran amine (BDA) as a neuroanatomical tracer. This study has confirmed previously reported afferent projections to nucleus rotundus in reptiles and has also identified a number of new cellular aggregates projecting to this dorsal thalamic nucleus. After BDA injections into nucleus rotundus, retrogradely labelled neurons were observed consistently within the following neuronal groups in the midbrain and the diencephalon: (i) the stratum griseum centrale of the optic tectum; (ii) the nucleus subpretectalis in the pretectum; (iii) the nucleus ansa lenticularis posterior, the posterior nucleus of the ventral supraoptic commissure, and the posteroventral nucleus, in the dorsal thalamus and (iv) the lateral suprachiasmatic nucleus and part of the reticular complex in the ventral thalamus. Tectal axons entering nucleus rotundus were fine and varicose and formed exclusively asymmetric synaptic contacts, mainly on small dendritic profiles. Rotundal neurons had symmetric synapses made by large boutons probably of nontectal origin. After comparing our results with those in other reptiles, birds and mammals, we propose that the sauropsidian nucleus rotundus forms part of a visual tectofugal pathway that conveys mesencephalic visual information to the striatum and dorsal ventricular ridge, and is similar to the mammalian colliculo-posterior/intralaminar-striatoamygdaloid pathway, the function of which may be to participate in visually guided behaviour.     

Some aspects of the organization of the thalamic reticular complex.

Anatomical methods which depend upon the anterograde axonal transport of isotopically labeled neuronal proteins or the retrograde axonal transport of the enzyme, horseradish peroxidase, have been used to elucidate the relationships between the reticular complex and the dorsal thalamus and cerebral cortex. Injections of tritiated amino acids in the dorsal thalamus or cerebral cortex in rats, cats and monkeys, show that as the bundles of thalamo-cortical and cortico-thalamic fibers joining a particular dorsal thalamic nucleus to its associated area of the cerebral cortex traverse the reticular complex, they each give rise to a dense zone of terminals occupying a sector of the reticular complex which is relatively constant for that dorsal thalamic nucleus and cortical area. However, because of the wide extent of the dendritic fields of the reticular cells and the degree of overlap between the sectors of the complex subtended by adjacent dorsal thalamic nuclei and adjacent cortical areas, it is likely that the reticular complex samples thalamo-cortical and cortico-thalamic activity in a somewhat unspecific manner. Fibers passing to the reticular complex from the intralaminar nuclei of the thalamus appear to be associated with the projection from the intralaminar nuclei to the striatum. Injections of tritiated amino acids in the reticular complex itself and injections of horseradish peroxidase in various other parts of the brain show that the only efferent pathway from the reticular complex terminates in the nuclei of the dorsal thalamus. The reticular complex does not appear to send fibers to other components of the ventral thalamus nor to the cerebral cortex.     

Organization of projections from the anterior pole of the nucleus reticularis thalami (NRT) to subdivisions of the motor thalamus: light and electron microscopic studies in the rhesus monkey.

Projections to the motor-related thalamic nuclei from the anterior pole of the reticular thalamic nucleus (NRT) were studied after injections of biotinylated dextran amine and wheat germ agglutinin conjugated horseradish peroxidase at light and electron microscopic levels, respectively. Each injection resulted in anterograde labeling in the three subdivisions of the ventral anterior nucleus (pars parvicellularis, VApc; pars densicellularis, VAdc; and pars magnocellularis, VAmc) and in the ventral lateral nucleus (VL). NRT fibers had beaded shapes and coursed in a posterior direction giving rise to relatively diffuse terminal plexuses. The average size of the beads (0.7 microm2) and their density per 100 microm of fiber length (23.7-25.7) were similar between the nuclei studied. At the electron microscopic level, anterogradely labeled boutons displayed positive immunoreactivity for gamma-aminobutyric acid (GABA), contained pleomorphic synaptic vesicles, and formed relatively long (approximately 0.4 microm) symmetric synaptic contacts. Usually, a single terminal formed synapses on more than one postsynaptic structure. Synaptic contacts were on projection and local circuit neurons and targeted mainly their distal dendrites. In the VAmc, synapses on local circuit neurons composed 48% of the total sample, in the VAdc/VApc and in the VL the proportion was higher, 65% and 62%, respectively. The results suggest that the input from the anterior pole of the monkey reticular nucleus to the motor-related thalamic nuclei is organized differently from what is known on the organization of connections of NRT with sensory thalamic nuclei in other species in that the terminal fields of individual fibers are diffuse rather than focal and that at least 50% of synapses are established on GABAergic local circuit neurons.     

Differential Expression of the GABAA Receptor Complex in the Dorsal Thalamus and Reticular Nucleus: An Immunohistochemical Study in the Adult and Developing Rat

The distribution of the GABAA receptor/benzodiazepine receptor/chloride channel complex was investigated in the thalamus of the rat by means of immunohistochemistry in adulthood, as well as during embryonic and postnatal development, using a monoclonal antibody. In adults, the immunoreactivity for the GABAA receptor complex was intensely expressed by neuronal processes throughout the dorsal thalamus. Neuronal perikaryal membranes were frequently outlined by punctate immunostaining; cell bodies, intrathalamic fibre bundles and the internal capsule did not display immunoreactivity for the GABAA receptor. Regional differences in the expression of the receptor were consistently observed: the immunostaining was much lighter in the thalamic reticular nucleus than in the dorsal thalamic nuclei and, among the latter, the anteroventral nucleus and the ventral nuclear complex displayed the most intense immunopositivity. Immunostaining for the GABAA receptor was already expressed in embryos at E14, and was homogeneously distributed throughout the neuropil of the dorsal and ventral thalamic primordia. During the first two postnatal weeks, a regional differentiation of the immunopositivity was appreciable in the thalamus, with a progressive reduction in the reticular nucleus and a parallel increase in the dorsal thalamic structures. Immunoreactive neuronal perikarya were not observed in the thalamus at any developmental stage. The expression of the GABAA receptor complex appeared to have reached a mature configuration by the end of the third postnatal week. These findings indicate that in adults the GABAA receptor is differentially expressed by thalamic nuclear structures, including the reticular nucleus. Furthermore, the maturation of the receptor in the thalamus undergoes a rearrangement during the first postnatal weeks that results in a considerable regression within the reticular nucleus.     

Cholinergic innervation of the mediodorsal thalamic nucleus in the monkey: Ultrastructural evidence supportive of functional diversity

The ultrastructural organization of association nuclei in the primate thalamus is largely unexplored. In the present study we have combined electron microscopy with immunocytochemistry for the acetylcholine synthesizing enzyme choline acetyltransferase (ChAT) to assess the cholinergic synaptic organization of the mediodorsal (MD) nucleus in macaque monkeys. The cholinergic innervation of the MD nucleus showed striking regional variations with the greatest density of immunoreactive axons and varicosities found within the parvicellular division. Electron microscopic examination revealed that these ChAT immunoreactive (ChAT-IR) axons were primarily small and unmyelinated. The majority of immunoreactive synaptic profiles were found within the extraglomerular neuropil (80.5%), with the remainder present in glomerular regions. Within the glomerular and extra-glomerular neuropil ChAT-IR profiles made contact with both conventional, presumably relay cell dendrites (CD), as well as with synaptic vesicle containing dendrites (SVCD) of local circuit neurons. In the glomeruli the frequency of synapses was approximately equal for CDs and SVCDs while in the extraglomerular areas 75% of the synaptic contacts were with CDs. ChAT-IR synaptic profiles had a diversity of junctional complex morphologies. Within glomeruli they made symmetric synapses with CDs and predominantly asymmetric with SVCDs. The majority of extraglomerular contacts (60%) were classified as asymmetric and these as well as the smaller number of symmetric synapses contacted both CDs and SVCDs. In accord with results of physiological studies, these anatomical data indicate that cholinergic input to thalamic nuclei influences relay cell activity both directly and indirectly via local circuit neurons. © 1993 Wiley-Liss, Inc.     

Selective GABAergic innervation of thalamic nuclei from zona incerta

Thalamocortical circuits that govern cortical rhythms and ultimately effect sensory transmission consist of three major interconnected elements: excitatory thalamocortical and corticothalamic neurons and GABAergic cells in the reticular thalamic nucleus. Based on the present results, a fourth component has to be added to this scheme. GABAergic fibres from an extrareticular diencephalic source were found to selectively innervate relay cells located mainly in higher-order thalamic nuclei. The origin of this pathway was localized to zona incerta (ZI), known to receive collaterals from corticothalamic fibres. First-order nuclei were innervated only in zones showing a high density of calbindin-positive neurons. The large GABA-immunoreactive incertal terminals established multiple contacts preferentially on the proximal dendrites of relay cells via symmetrical synapses with multiple release sites. The distribution, ultrastructural characteristics and postsynaptic target selection of extrareticular terminals were similar to type II muscarinic acetylcholine receptor-positive boutons, which constituted up to 49% of all GABAergic terminals in the posterior nucleus. This suggests that a significant proportion of the GABAergic input into certain thalamic territories involved in higher-order functions may have extrareticular origin. Unlike the reticular nucleus, ZI receives peripheral and layer V cortical input but no thalamic feedback; it projects to brainstem centres and has extensive intranuclear recurrent collaterals. This indicates that ZI exerts a conceptually new type of inhibitory control over the thalamus. The proximally situated, multiple active zones of ZI terminals indicate a powerful influence on the firing properties of thalamic neurons, which is conveyed to multiple cortical areas via relay cells which have widespread projections to neocortex.     

Differential gene expression for glutamic acid decarboxylase and type II calcium-calmodulin-dependent protein kinase in basal ganglia, thalamus, and hypothalamus of the monkey.

In situ hybridization histochemistry, using cRNA probes, revealed a complementarity in the distributions of cells in the basal ganglia, basal nucleus of Meynert, thalamus, hypothalamus, and rostral part of the midbrain that showed gene expression for glutamic acid decarboxylase (GAD) or the alpha-subunit of type II calcium-calmodulin-dependent protein kinase (CAM II kinase-alpha). Cells in certain nuclei such as the thalamic reticular nucleus, globus pallidus, and pars reticulata of the substantia nigra show GAD gene expression only; others in nuclei such as the basal nucleus of Meynert, medial mamillary nuclei, and ventromedial hypothalamic nuclei show CAM II kinase-alpha gene expression only. A few nuclei, for example, the pars compacta of the substantia nigra and the greater part of the subthalamic nucleus, display gene expression for neither GAD nor CAM II kinase-alpha. In other nuclei, notably those of the dorsal thalamus, and possibly in the striatum, GAD- and CAM II kinase-expressing cells appear to form two separate populations that, in most thalamic nuclei, together account for the total cell population. In situ hybridization reveals large amounts of CAM II kinase-alpha mRNA in the neuropil of most nuclei containing CAM II kinase-alpha-positive cells, suggesting its association with dendritic polyribosomes. The message may thus be translated at those sites, close to the synapses with which the protein is associated. The in situ hybridization results, coupled with those from immunocytochemical staining for CAM II kinase-alpha protein, indicate that CAM II kinase-alpha is commonly found in certain non-GABAergic afferent fiber systems but is not necessarily present in the postsynaptic cells on which they terminate. It appears to be absent from most GABAergic fiber systems but can be present in the cells on which they terminate. This suggests that the kinase may be differentially engaged in pre- and postsynaptic functions at certain synapses.