Single-axon tracing and three-dimensional reconstruction of centre median-parafascicular thalamic neurons in primates

The axonal projections from the centre median (CM)/parafascicular (Pf) thalamic complex in squirrel monkeys were studied after microiontophoretic injections of biotinylated dextran amine under electrophysiological guidance. A total of 29 axons connected to their parent cell body were entirely reconstructed from serial sections with a camera lucida. Our investigation shows that the CM and Pf nuclei in primates comprise three types of projection neurons: (1) neurons that innervate densely and focally the striatum; (2) neurons that arborize diffusely in the cerebral cortex; and (3) neurons that innervate both striatum and cerebral cortex. Striatal innervation of CM origin consists of dense clusters of axon terminals exhibiting pedunculated varicosities and forming oblique bands in the dorsolateral sector of putamen (sensorimotor striatal territory). The same type of striatal innervation occurs in the head of caudate nucleus (associative striatal territory) in cases of Pf-labeled neurons. The CM neurons that target cerebral cortex arborize principally in motor and premotor areas, whereas Pf neurons innervate chiefly prefrontal areas. Cortical innervation from both nuclei is much more profuse in layers V and VI than in layer I. Our three-dimensional reconstruction studies show that dendritic and axonal arborizations of CM neurons extend essentially along the sagittal plane. These results revealed that, in contrast to rodents where virtually all Pf neurons project to both striatum and cortex, the primate CM/Pf complex harbors several types of highly patterned projection neurons. As such, this complex might be considered as an integral part of the widely distributed basal ganglia neuronal system.     

Efferent connections of the centromedian and parafascicular thalamic nuclei in the squirrel monkey: a PHA-L study of subcortical projections.

The subcortical projections of the centromedian (CM) and the parafascicular (Pf) thalamic nuclei were examined in the squirrel monkey (Saimiri sciureus) by using the lectin Phaseolus vulgaris-leucoagglutinin (PHA-L) as an anterograde tracer. Both CM and Pf project massively to the striatum where they arborize in a complementary fashion. On the one hand, CM innervates most of the putamen caudal to the anterior commissure, a dorsolateral rim of the putamen rostral to the anterior commissure, discrete areas of the head of the caudate nucleus close to the internal capsule, and a lateral sector of the body of the caudate nucleus. On the other hand, Pf provides a heavy input to the head, body, and tail of the caudate nucleus, and to the rostral putamen, excluding the areas innervated by CM. In addition, Pf projects more discretely to the nucleus accumbens and the olfactory tubercle. Therefore, the projections from both CM and Pf cover the entire striatum, with those from CM arborizing into the "sensorimotor" striatal territory and the ones from Pf innervating the "associative-limbic" striatal territory. Furthermore, CM and Pf project to extrastriatal subcortical structures, such as the globus pallidus, the subthalamic nucleus, and the substantia nigra, where they also terminate in a complementary fashion. Topographically and cytologically, Pf is closely related to the subparafascicular nucleus (sPf). The Pf-sPf complex projects to the hypothalamus, the substantia innominata, the peripeduncular nucleus, and the amygdala. It also gives rise to descending efferents arborizing in various brainstem structures, including the inferior olivary complex. Additional studies with retrograde double-labeling methods show that distinct cell groups within CM project to the motor cortex and the striatum. Likewise, separate neuronal populations within the CM-Pf-sPf complex give rise to striatal and brainstem projections, the former arising from CM and Pf and the latter mainly from sPf. The complementary nature of CM and Pf projections to the striatum and other basal ganglia components suggests that this thalamic complex participates in a highly ordered manner in the parallel processing of the information that flows through the basal ganglia.     

Amygdaloid projections to the frontal cortex and the striatum in the rat.

Projections from the basolateral nucleus of the amygdala (BLA) to the frontal cortex and the striatum were studied by using Phaseolus vulgaris-leucoagglutinin (PHA-L) anterograde tracing technique in the rat. PHA-L injections into the rostral part of the BLA resulted in a dense labeling of fibers with boutons in the dorsal bank of the rhinal fissure and in the lateral and the medial agranular cortex. PHA-L injections into the caudal part of the BLA produced a dense labeling of fibers in the medial surface of the frontal cortex. In most of the cortical regions, labeled fibers were predominantly distributed in two bands: one in the deep part of layers I and II and the other, heavier band, in layers V and VI. PHA-L injections into the rostral BLA resulted in a dense labeling of fibers with boutons in the olfactory tubercle, the rostral and caudolateral portion of the nucleus accumbens, and a large region of the caudate-putamen. The labeled area of the caudate-putamen included the rostroventral area, the central area, and the area caudal to the anterior commissure and dorsal and lateral to the globus pallidus. PHA-L injections into the caudal BLA produced fiber labeling in the most rostromedial area of the caudate-putamen facing the lateral ventricle, the medial portion of the nucleus accumbens, and the lateral septum. In the rostroventral striatum, PHA-L-labeled fibers selectively innervated the matrix compartment that contains abundant somatostatin-immunoreactive fibers. Compartmental segregation was less clear in the caudodorsolateral caudate-putamen and in the nucleus accumbens. Electron microscopy revealed that PHA-L-labeled boutons in the striatum contained abundant, small, round vesicles. These boutons formed asymmetrical synapses with dendritic spines of striatal neurons.     

Differential synaptology of vGluT2-containing thalamostriatal afferents between the patch and matrix compartments in rats

The striatum is divided into two compartments named the patch (or striosome) and the matrix. Although these two compartments can be differentiated by their neurochemical content or afferent and efferent projections, the synaptology of inputs to these striatal regions remains poorly characterized. By using the vesicular glutamate transporters vGluT1 and vGluT2, as markers of corticostriatal and thalamostriatal projections, respectively, we demonstrate a differential pattern of synaptic connections of these two pathways between the patch and the matrix compartments. We also demonstrate that the majority of vGluT2-immunolabeled axon terminals form axospinous synapses, suggesting that thalamic afferents, like corticostriatal inputs, terminate preferentially onto spines in the striatum. Within both compartments, more than 90% of vGluT1-containing terminals formed axospinous synapses, whereas 87% of vGluT2-positive terminals within the patch innervated dendritic spines, but only 55% did so in the matrix. To characterize further the source of thalamic inputs that could account for the increase in axodendritic synapses in the matrix, we undertook an electron microscopic analysis of the synaptology of thalamostriatal afferents to the matrix compartments from specific intralaminar, midline, relay, and associative thalamic nuclei in rats. Approximately 95% of PHA-L-labeled terminals from the central lateral, midline, mediodorsal, lateral dorsal, anteroventral, and ventral anterior/ventral lateral nuclei formed axospinous synapses, a pattern reminiscent of corticostriatal afferents but strikingly different from thalamostriatal projections arising from the parafascicular nucleus (PF), which terminated onto dendritic shafts. These findings provide the first evidence for a differential pattern of synaptic organization of thalamostriatal glutamatergic inputs to the patch and matrix compartments. Furthermore, they demonstrate that the PF is the sole source of significant axodendritic thalamic inputs to striatal projection neurons. These observations pave the way for understanding differential regulatory mechanisms of striatal outflow from the patch and matrix compartments by thalamostriatal afferents.     

The centre me´dian and parafascicular thalamic nuclei project respectively to the sensorimotor and associative-limbic striatal territories in the squirrel monkey

The striatal projections of the centre me´dian (CM) and parafascicular (Pf) thalamic nuclei were examined in the squirrel monkey (Saimiri sciureus) by using the lectin wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP) as an anterograde tracer. CM was found to project massively to the putamen, where terminal fields appeared principally in the form of oblique bands, and more diffusely to the dorsolateral border of the caudate nucleus. Striatal inputs from Pf were found more rostrally, especially in the ventromedial portion of the putamen, the entire ventromedial half of the caudate nucleus, and the ventral striatum including the nucleus accumbens and the olfactory tubercle. Pf terminal fields in the rostral striatum often displayed a patchy organization. Both CM and Pf projections were found to terminate in the matrix compartment of the striatum as defined by acetylcholinesterase staining. These results suggest that CM is more specifically involved in sensorimotor and Pf in associative and limbic aspects of basal ganglia function in primates.     

The centre médian and parafascicular thalamic nuclei project respectively to the sensorimotor and associative-limbic striatal territories in the squirrel monkey.

The striatal projections of the centre médian (CM) and parafascicular (Pf) thalamic nuclei were examined in the squirrel monkey (Saimiri sciureus) by using the lectin wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP) as an anterograde tracer. CM was found to project massively to the putamen, where terminal fields appeared principally in the form of oblique bands, and more diffusely to the dorsolateral border of the caudate nucleus. Striatal inputs from Pf were found more rostrally, especially in the ventromedial portion of the putamen, the entire ventromedial half of the caudate nucleus, and the ventral striatum including the nucleus accumbens and the olfactory tubercle. Pf terminal fields in the rostral striatum often displayed a patchy organization. Both CM and Pf projections were found to terminate in the matrix compartment of the striatum as defined by acetylcholinesterase staining. These results suggest that CM is more specifically involved in sensorimotor and Pf in associative and limbic aspects of basal ganglia function in primates.     

The primate centromedian-parafascicular complex: anatomical organization with a note on neuromodulation

In addition to the cerebral cortex, the striatum receives excitatory input from the thalamus. The centromedian (centre median, CM) and parafascicular (Pf) nuclei are an important source of thalamostriatal projections. Anterograde tract-tracing indicates the CM-Pf complex provides dense afferents to the matrix compartment of the striatum. Whereas CM projects to the entire sensorimotor territory of the striatum, the Pf provides complementary input to the entire associative sector. The Pf also provides lighter input to the nucleus accumbens. Both CM and Pf provide light to moderately dense inputs to other components of the basal ganglia in a largely complementary manner, covering motor or associative-limbic territories of the subthalamic nucleus, globus pallidus and ventral midbrain. In turn, the CM and Pf receive mainly segregated input from parallel motor and associative-limbic circuits of the basal ganglia. The CM and Pf may therefore be considered important participants in parallel processing of motor and associative-limbic information in the basal ganglia. Connections of the CM and Pf with other thalamic nuclei suggest they also participate in integrative functions within the thalamus. In addition, inputs from the brainstem reticular core, reciprocal connections with the cerebral cortex and reticular thalamic nucleus suggest a role in state-dependant information processing. Consideration of the differential connections of the CM and Pf, and better understanding of their role in pathophysiology, may eventually lead to development of an important new target for relief of a variety of neurological and psychiatric disorders.     

Nigral and pallidal inputs to functionally segregated thalamostriatal neurons in the centromedian/parafascicular intralaminar nuclear complex in monkey

In primates, thalamostriatal projections from the centromedian (CM) and parafascicular (Pf) nuclei are strong and organized according to a strict pattern of functional connectivity with various regions of the striatal complex. In turn, the CM/Pf complex receives a substantial innervation from the internal globus pallidus (GPi). In this study, we demonstrate that the substantia nigra pars reticulata (SNr) also provides a massive input to Pf in monkeys. These pallidothalamic and nigrothalamic projections provide routes whereby information can flow in functional loops between the basal ganglia and the intralaminar nuclear group. To understand better the anatomical organization and the degree of functional specificity of these loops, we combined retrograde and anterograde labeling methods from functionally defined regions of the striatum and GPi/SNr to determine the relationships between thalamostriatal neurons and basal ganglia afferents. Together with previous studies, our data suggest the existence of tightly connected functional circuits between the basal ganglia and the CM/Pf in primates: 1) A "sensorimotor" circuit links together the medial two-thirds of CM, the postcommissural putamen, and the ventrolateral part of the caudal GPi; 2) a "limbic" circuit involves the rostral one-third of Pf, the ventral striatum, and the rostromedial pole of GPi; and 3) an "associative"circuit exists between the caudal two-thirds of Pf, the caudate nucleus, and the SNr. An additional "associative" circuit that involves the caudate-receiving territory of GPi (dorsal one-third), the dorsolateral Pf (Pfdl), and the precommissural putamen was also disclosed. In conclusion, findings of this study provide additional evidence for the high degree of functional specificity of the thalamostriatal system through which CM/Pf may provide attention-specific sensory information important for conditional responses to the primate striatum.     

Differential synaptic innervation of striatofugal neurones projecting to the internal or external segments of the globus pallidus by thalamic afferents in the squirrel monkey

It is well established that the centromedian nucleus (CM) is the major source of thalamic afferents to the sensorimotor territory of the striatum in monkeys. However, the projection sites of striatal neurones contacted by thalamic afferents still remain to be determined. We therefore carried out an anatomical study aimed at elucidating the hodology of striatal neurones that receive input from the CM in squirrel monkeys. Our approach was to combine the anterograde transport of Phaseolus vulgaris-leucoagglutinin (PHA-L) or biocytin from the CM with the retrograde transport of biotinylated dextran-amine (bio-dex) or PHA-L from the internal (GPi) or external (GPe) segments of the globus pallidus. Following CM injections, rich plexuses of anterogradely labelled, thin varicose fibres aggregated in the form of bands that were confined to the postcommissural region of the putamen. On the other hand, injections into the GPe or GPi led to profuse retrograde labelling of a multitude of medium-sized spiny neurones. In cases where the injections involved the caudoventral two-thirds of the GPe or GPi, the retrogradely labelled striatopallidal cells and the anterogradely labelled thalamostriatal fibres occurred in the sensorimotor territory of the putamen. After injections into either pallidal segments, clusters of retrogradely labelled cells were in register with bands of anterogradely labelled thalamic fibres. However, electron microscopic analysis of striatal regions containing both anterogradely labelled thalamic afferents and retrogradely labelled cells revealed that terminals from the CM frequently form asymmetric synapses with dendritic shafts and spines of striato-GPi cells but rarely with those of striato-GPe cells. In conclusion, our findings demonstrate that thalamic afferents from the CM innervate preferentially striatopallidal neurones projecting to the GPi in monkeys. These results indicate that the striatopallidal neurones contributing to the “direct” and “indirect” output pathways are differentially innervated by thalamic afferents in primates. © 1996 Wiley-Liss, Inc.     

Rat intralaminar thalamic nuclei projections to the globus pallidus: a biotinylated dextran amine anterograde tracing study

The topographical organization and ultrastructural features of the intralaminar thalamic nuclei (ITN) projections to the globus pallidus (GP) were studied using the biotinylated dextran amine (BDA) anterograde tracing method in the rat. To assess the functional association of BDA injection sites in the ITN, the known topographical organization of the ITN-neostriatal (Str) projections and calcium binding protein (CaBP) immunostaining patterns of the Str and GP were used. BDA injection in the lateral part of the lateral parafascicular nucleus and the caudal part of the central lateral nucleus labeled fibers and boutons mainly in the dorsolateral sensorimotor territory of the Str and the middle territories of the GP. BDA injection in the medial part of the lateral parafascicular nucleus and the central lateral nucleus labeled mainly the middle association territory of the Str and the border and the caudomedial territories of the GP. BDA injection in the medial parafascicular nucleus and the central medial nucleus labeled mainly the medial limbic territory of the Str. The medial parafascicular nucleus projected to the medial-most region of the GP, while the central medial nucleus projection to the GP was very sparse. Electron microscopic observations indicated that BDA-labeled boutons form asymmetric synapses mainly on 0.5-2.0 microm diameter dendritic shafts in the GP. The boutons were small but had a relatively long active zone. The present observations together with the known topographical organization of striatopallidal projections indicated that the ITN-GP projections were topographically organized in parallel to the ITN-Str projections. Thus, each part of the ITN projecting to the sensorimotor, the association, and the limbic territories of the Str also projects to the corresponding functional territories of the GP.     

Synaptic organization of projections from the amygdala to visual cortical areas TE and V1 in the macaque monkey

The primate amygdaloid complex projects to a number of visual cortices, including area V1, primary visual cortex, and area TE, a higher-order unimodal visual area involved in object recognition. We investigated the synaptic organization of these projections by injecting anterograde tracers into the amygdaloid complex of Macaca fascicularis monkeys and examining labeled boutons in areas TE and V1 using the electron microscope. The 256 boutons examined in area TE formed 263 synapses. Two hundred twenty-three (84%) of these were asymmetric synapses onto dendritic spines and 40 (15%) were asymmetric synapses onto dendritic shafts. Nine boutons (3.5%) formed double asymmetric synapses, generally on dendritic spines, and 2 (1%) of the boutons did not form a synapse. The 200 boutons examined in area V1 formed 211 synapses. One hundred eighty-nine (90%) were asymmetric synapses onto dendritic spines and 22 (10%) were asymmetric synapses onto dendritic shafts. Eleven boutons (5.5%) formed double synapses, usually with dendritic spines. We conclude from these observations that the amygdaloid complex provides an excitatory input to areas TE and V1 that primarily influences spiny, probably pyramidal, neurons in these cortices.     

Striatal projections of the vagal-responsive region of the thalamic parafascicular nucleus in macaque monkeys

We recently reported that the thalamic parafascicular nucleus (Pf) in monkeys is strongly activated by vagus nerve afferents. The main forebrain target of Pf is the striatum, but the specific striatal regions receiving visceral input via this pathway are unknown. We examined the projections of this region by injecting anterograde tracers into the vagus evoked potential (VEP) focus in Pf of macaque monkeys. The VEP was strongest lateral and anterior to the habenulointerpeduncular tract, but it was distributed across the entire horizontal extent of the ventral half of Pf. All injections produced labeled terminals in the caudate (Cd), especially the Cd tail and the adjacent ventral posterior Pu. Terminations occurred throughout the Cd head and body but spared the most anterior and dorsolateral parts. Injections in more anterior and lateral portions of Pf produced progressively more terminations in Pu, mainly in the precommissural region and the medial aspect of posterior Pu. Dual injections of different tracers revealed overlapping projections with interdigitated strands of striatal terminations from separate regions of Pf as well as the posteromedial to anterolateral topographic gradient of increasing Pf projections to Pu. An injection in the most anteromedial portion of Pf produced strong labeling in the ventral striatum. Thus, Pf transmits viscerosensory information to the "associative" and "limbic" territories of the striatum. These findings suggest the broad involvement of homeostatic afferent activity in striatal function and perhaps a role for the striatum in autonomic function.     

Restoration of thalamostriatal projections in rat neostriatal grafts: an electron microscopic analysis

The thalamostriatal projections to rat neostriatal grafts were studied by using the Phaseolus vulgaris-leucoagglutinin (PHA-L) axonal tracing technique. Two to 6 months after implantation of striatal primordia into adult neostriata, PHA-L was injected into two different portions of the intralaminar nuclear complex of the thalamus. In the host neostriatum, labeled fibers from the parafascicular nucleus (PF) arborized in a large region in the neostriatum, but avoided small patchlike areas. Most of the fibers from PF had irregular curved trajectories with short side branches that formed boutons. Labeled fibers from the centromedial and paracentral nuclei (CeM-PC) projected to a similarly large area within the neostriatum but did not show any nonuniformity. CeM-PC axons had relatively straight trajectories and formed boutons en passant. Both sets of thalamostriatal projection fibers were found in the grafts. Some of the labeled fibers in the grafts formed dense, focal arborizations. Compared to the host neostriatum, the distribution of postsynaptic elements in the grafts was altered dramatically. In the host neostriatum, 89% of the terminals from PF terminated onto dendritic shafts; 93% of the CeM-PC terminals contacted dendritic spines. However, only 47% of the PF terminals in the grafts contacted dendritic shafts; 53% of them terminated on dendritic spines. In grafts, 81% of the terminals from CeM-PC region contacted dendritic spines; 19% of them made synapses on dendritic shafts. The shift of postsynaptic elements in the grafts suggests a loss of pathway specificity in the induction of dendritic spines on neostriatal neurons in grafts.     

Fine structure of the medullary lateral line area of Chelon labrosus (order perciformes), a nonelectroreceptive teleost

The ultrastructure and synaptic organization of the nucleus medialis and cerebellar crest of the teleost Chelon labrosus have been investigated. The nucleus medialis receives projections from the anterior and posterior lateral line nerves. This nucleus consists of oval neurons and large crest cells (“Purkinje-like” cells) whose apical dendrites branch in the overlying molecular layer, the cerebellar crest. In the dorsal region of the nucleus medialis, the perikarya and smooth primary dendrites of the crest cells are interspersed among myelinated fibers and nerve boutons. The ventral layer of the nucleus medialis contains crest cell perikarya and dendrites as well as oval neurons. The cerebellar crest lacks neuronal bodies, but the apical dendrites of crest cells receive synapses from unmyelinated and myelinated fibers. In the cerebellar crest, two types of terminals are presynaptic to the crest cell dendrites: boutons with spherical vesicles that from asymmetric synapses with dendritic spines and boutons containing pleomorphic vesicles that from symmetric synapses with dendritic spines and boutons containing pleomorphic vesicles that from symmetric synapses directly on the dendritic shaft. Most axon terminals found on the somata and primary dedrites of crest cells in the nucleus medialis have pleomorphic vesicles and form symmetric contacts, though asymmetric with spherical vesicles and mixed synapses can be observed; these mixed synapses exhibit gap junctions and contain spherical vesicles. Unlike crest cells, the oval neuron perikarya receive three types of contacts (symmetric, asymmetric, and mixed). The origins and functions of these different bouton types in the nucleus medialis are discussed. © 1995 Willy-Liss, Inc.     

The thalamostriatal systems: anatomical and functional organization in normal and parkinsonian states

Although we have gained significant knowledge in the anatomy and microcircuitry of the thalamostriatal system over the last decades, the exact function(s) of these complex networks remain(s) poorly understood. It is now clear that the thalamostriatal system is not a unique entity, but consists of multiple neural systems that originate from a wide variety of thalamic nuclei and terminate in functionally segregated striatal territories. The primary source of thalamostriatal projections is the caudal intralaminar nuclear group which, in primates, comprises the centromedian and parafascicular nuclei (CM/Pf). These two nuclei provide massive, functionally organized glutamatergic inputs to the whole striatal complex. There are several anatomical and physiological features that distinguish this system from other thalamostriatal projections. Although all glutamatergic thalamostriatal neurons express vGluT2 and release glutamate as neurotransmitter, CM/Pf neurons target preferentially the dendritic shafts of striatal projection neurons, whereas all other thalamic inputs are almost exclusively confined to the head of dendritic spines. This anatomic arrangement suggests that transmission of input from sources other than CM/Pf to the striatal neurons is likely regulated by dopaminergic afferents in the same manner as cortical inputs, while the CM/Pf axo-dendritic synapses do not display any particular relationships with dopaminergic terminals. A better understanding of the role of these systems in the functional circuitry of the basal ganglia relies on future research of the physiology and pathophysiology of these networks in normal and pathological basal ganglia conditions. Although much remains to be known about the role of these systems, recent electrophysiological studies from awake monkeys have provided convincing evidence that the CM/Pf-striatal system is the entrance for attention-related stimuli to the basal ganglia circuits. However, the processing and transmission of this information likely involves intrinsic GABAergic and cholinergic striatal networks, thereby setting the stage for complex physiological responses of striatal output neurons to CM/Pf activation. Finally, another exciting development that will surely generate significant interest towards the thalamostriatal systems in years to come is the possibility that CM/Pf may be a potential surgical target for movement disorders, most particularly Tourette syndrome and Parkinson's disease. Although the available clinical evidence is encouraging, these procedures remain empirical at this stage because of the limited understanding of the thalamostriatal systems.     

GABAergic neural elements in the rat basilar pons: electron microscopic immunochemistry

Previous light microscopic immunoperoxidase studies of glutamic acid decarboxylase (GAD)-immunoreactive neural elements in the rat basilar pontine nuclei revealed immunocytochemical reaction product in neuronal somata and axon terminals. In the present study, pre-embedding immunoperoxidase labeling of GAD or gamma-aminobutyric acid (GABA) and postembedding immunogold labeling of GABA allowed the ultrastructural visualization of these neural elements in the basilar pontine nuclei of colchicine-treated animals. At the electron microscopic level, immunolabeled neuronal somata exhibited smoothly contoured nuclei, whereas some dendrites also contained reaction product after immunocytochemical treatment and were postsynaptic to both immunoreactive and nonimmunoreactive axon terminals. Synaptic boutons immunoreactive for GAD or GABA exhibited cross-sectional areas that ranged from 0.1 to 3.8 microns 2 and generally appeared round or elongated in most sections. The majority (95%) of immunolabeled boutons contained pleomorphic synaptic vesicles and formed symmetric synapses at their postsynaptic loci; however, boutons exhibiting round vesicles and boutons forming asymmetric synapses (5%) were also immunopositive. Small (less than 1.5 microns 2) GAD- or GABA-labeled axon terminals formed synaptic contact mainly with small dendritic profiles, dendritic spines, and neuronal somata, whereas large labeled boutons (greater than 1.5 microns 2) formed synapses with all sizes of dendritic profiles. Occasionally, a single immunolabeled bouton formed synaptic contact with two separate postsynaptic dendrites. It is suggested that the immunolabeled neuronal somata and dendrites observed in the rat basilar pontine nuclei represent a population of pontine local circuit neurons; however, it is known that GABAergic cell groups extrinsic to the pontine gray provide afferent projections to the basilar pons, and therefore at least some immunoreactive axon terminals present in the pontine nuclei are derived from these extrinsic sources. The ultrastructural observation of GABAergic neural elements in the rat basilar pontine nuclei confirms previous light microscopic findings and provides an anatomical substrate through which GABAergic neurons, whether arising from an intrinsic or extrinsic source, might exert an inhibitory influence on target cells within the pontine nuclei.     

Synaptic microcircuitry of tyrosine hydroxylase-containing neurons and terminals in the striatum of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated monkeys

A population of tyrosine hydroxylase (TH)-containing neurons that is up-regulated after lesion of the nigrostriatal dopaminergic pathway has been described in the primate striatum. The goal of this study was to examine the morphology, synaptology, and chemical phenotype of these neurons and TH-immunoreactive (-ir) terminals in the striatum of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated rhesus monkeys. TH-ir perikarya were small (10-12 microm), displayed nuclear invaginations, and received very few synaptic inputs. On the other hand, TH-containing dendrites were typically large in diameter (>1.0 microm) and received scarce synaptic innervation from putative excitatory and inhibitory terminals forming asymmetric and symmetric synapses, respectively. More than 70% of TH-positive intrastriatal cell bodies were found in the caudate nucleus and the precommissural putamen, considered as the associative functional territories of the primate striatum. Under 10% of these cells displayed calretinin immunoreactivity. TH-ir terminals rarely formed clear synaptic contacts, except for a few that established asymmetric axodendritic synapses. Almost two-thirds of TH-containing boutons displayed gamma-aminobutyric acid (GABA) immunoreactivity in the striatum of parkinsonian monkeys, whereas under 5% did so in the normal striatum. These findings provide strong support for the existence of a population of putative catecholaminergic interneurons in the associative territory of the striatum in parkinsonian monkeys. Their sparse synaptic innervation raises interesting issues regarding synaptic and nonsynaptic mechanisms involved in the regulation and integration of these neurons in the striatal microcircuitry. Finally, the coexpression of GABA in TH-positive terminals in the striatum of dopamine-depleted monkeys suggests dramatic neurochemical changes in the catecholaminergic modulation of striatal activity in Parkinson's disease.     

Synaptic relationships between axon terminals from the mediodorsal thalamic nucleus and gamma-aminobutyric acidergic cortical cells in the prelimbic cortex of the rat

Although the reciprocal interconnections between the prefrontal cortex and the mediodorsal nucleus of the thalamus (MD) are well known, the involvement of inhibitory cortical interneurons in the neural circuit has not been fully defined. To address this issue, we conducted three combined neuroanatomical studies on the rat brain. First, the frequency and the spatial distribution of synapses made by reconstructed dendrites of nonpyramidal neurons were identified by impregnation of cortical cells with the Golgi method and identification of thalamocortical terminals by degeneration following thalamic lesions. Terminals from MD were found to make synaptic contacts with small dendritic shafts or spines of Golgi-impregnated nonpyramidal cells with very sparse dendritic spines. Second, a combined study that used anterograde transport of Phaseolus vulgaris leucoagglutinin (PHA-L) and postembedding gamma-aminobutyric acid (GABA) immunocytochemistry indicated that PHA-L-labeled terminals from MD made synaptic junctions with GABA-immunoreactive dendritic shafts and spines. Nonlabeled dendritic spines were found to receive both axonal inputs from MD with PHA-L labelings and from GABAergic cells. In addition, synapses were found between dendritic shafts and axon terminals that were both immunoreactive for GABA. Third, synaptic connections between corticothalamic neurons that project to MD and GABAergic terminals were investigated by using wheat germ agglutinin conjugated to horseradish peroxidase and postembedding GABA immunocytochemistry. GABAergic terminals in the prelimbic cortex made symmetrical synaptic contacts with retrogradely labeled corticothalamic neurons to MD. All of the synapses were found on cell somata and thick dendritic trunks. These results provide the first demonstration of synaptic contacts in the prelimbic cortex not only between thalamocortical terminals from MD and GABAergic interneurons but also between GABAergic terminals and corticothalamic neurons that project to MD. The anatomical findings indicate that GABAergic interneurons have a modulatory influence on excitatory reverberation between MD and the prefrontal cortex.     

The striatum and the globus pallidus send convergent synaptic inputs onto single cells in the entopeduncular nucleus of the rat: a double anterograde labelling study combined with postembedding immunocytochemistry for GABA.

The entopeduncular nucleus is one of the major output stations of the basal ganglia. In order to better understand the role of this structure in information flow through the basal ganglia, experiments have been performed in the rat to examine the chemical nature, morphology, and synaptology of the projections from the globus pallidus and striatum to the entopeduncular nucleus. In order to examine the morphology and synaptology of pallidoentopeduncular terminals and striatoentopeduncular terminals, rats were subjected to a double anterograde labelling study. The globus pallidus was injected with Phaseolus vulgaris-leucoagglutinin (PHA-L), and on the same side of the brain, the striatum was injected with biocytin. The entopeduncular nuclei of these animals were then examined for anterogradely labelled pallidal and striatal terminals. Rich plexuses of PHA-L-labelled pallidal terminals and biocytin-labelled striatal terminals were identified throughout the entopeduncular nucleus. At the electron microscopic level, the pallidal boutons were classified as two types. The majority (Type 1), were large boutons that formed symmetrical synapses with the dendrites and perikarya of neurones in the entopeduncular nucleus. Type 2 PHA-L-labelled terminals were much rarer, slightly smaller, and formed asymmetrical synapses. It is suggested that the Type 2 boutons are not derived from the globus pallidus but from the subthalamic nucleus. The biocytin-labelled terminals from the striatum had the typical morphological features of striatal terminals and formed symmetrical synapses. The distribution of the postsynaptic targets of the pallidal terminals and the striatal terminals differed in that the pallidal terminals preferentially made synaptic contact with the more proximal regions of the neurones in the entopeduncular nucleus, whereas the striatal terminals were located more distally on the dendritic trees. Examination in the electron microscope of areas where there was an overlap of the two sets of anterogradely labelled boutons revealed that terminals from the globus pallidus and the striatum made convergent synaptic contact with the perikarya and dendrites of individual neurones in the entopeduncular nucleus. In order to examine the chemical nature of the input to the entopeduncular nucleus from the globus pallidus and the striatum, ultrathin sections were immunostained by the postembedding method to reveal endogenous GABA. Three classes of GABA-containing terminals were identified; two of them formed symmetrical synapses and one rare type formed asymmetrical synapses. The combination of the GABA immunocytochemistry and anterograde labelling revealed that both the striatal and pallidal afferents that make symmetrical synapses with neurones in the entopeduncular nucleus, including those involved in convergent inputs, are GABAergic.(ABSTRACT TRUNCATED AT 400 WORDS)     

Intralaminar and tectal projections to the subthalamus in the rat

Projections from the posterior intralaminar thalamic nuclei and the superior colliculus (SC) to the subthalamic nucleus (STN) and the zona incerta (ZI) have been described in the primate and rodent. The aims of this study was to investigate several questions on these projections, using modern neurotracing techniques in rats, to advance our understanding of the role of STN and ZI. We examined whether projection patterns to the subthlamus can be used to identify homologues of the primate centromedian (CM) and the parafascicular nucleus (Pf) in the rodent, the topography of the projection including what percent of intralaminar neurons participate in the projections, and electron microscopic examination of intralaminar synaptic boutons in STN. The aim on the SC‐subthalamic projection was to examine whether STN is the main target of the projection. This study revealed: (i) the areas similar to primate CM and Pf could be recognized in the rat; (ii) the Pf‐like area sends a very heavy topographically organized projection to STN but very sparse projection to ZI, which suggested that Pf might control basal ganglia function through STN; (iii) the projection from the CM‐like area to the subthalamus was very sparse; (iv) Pf boutons and randomly sampled asymmetrical synapses had similar distributions on the dendrites of STN neurons; and (v) the lateral part of the deep layers of SC sends a very heavy projection to ZI and moderate to sparse projection to limited parts of STN, suggesting that SC is involved in a limited control of basal ganglia function.