Tyrosine hydroxylase immunoreactivity and [3H]WIN 35,428 binding to the dopamine transporter in a hamster model of idiopathic paroxysmal dystonia.

Recent pharmacological studies and receptor analyses have suggested that dopamine neurotransmission is enhanced in mutant dystonic hamsters (dt(sz)), a model of idiopathic paroxysmal dystonia which displays attacks of generalized dystonia in response to mild stress. In order to further characterize the nature of dopamine alterations, the present study investigated possible changes in the number of dopaminergic neurons, as defined by tyrosine hydroxylase immunohistochemistry, as well as binding to the dopamine transporter labelled with [3H]WIN 35,428 in dystonic hamsters. No differences in the number of tyrosine hydroxylase-immunoreactive neurons were found within the substantia nigra and ventral tegmental area of mutant hamsters compared to non-dystonic control hamsters. Similarly, under basal conditions, i.e. in the absence of a dystonic episode, no significant changes in [3H]WIN 35,428 binding were detected in dystonic brains. However, in animals killed during the expression of severe dystonia, significant decreases in dopamine transporter binding became evident in the nucleus accumbens and ventral tegmental area in comparison to controls exposed to the same external stimulation. Since stimulation tended to increase [3H]WIN 35,428 binding in control brains, the observed decrease in the ventral tegmental area appeared to be due primarily to the fact that binding was increased less in dystonic brains than in similarly stimulated control animals. This finding could reflect a diminished ability of the dopamine transporter to undergo adaptive changes in response to external stressful stimulation in mutant hamsters. The selective dopamine uptake inhibitor GBR 12909 (20 mg/kg) aggravated dystonia in mutant hamsters, further suggesting that acute alterations in dopamine transporter function during stimulation may be an important component of dystonia in this model.     

Serotonin-1 receptor binding sites in the human basal ganglia are decreased in Huntington's chorea but not in Parkinson's disease: a quantitative in vitro autoradiography study

Serotonin-1 receptors were examined in post-mortem human brains, using quantitative in vitro autoradiography. [3H]Serotonin was used as a ligand. Serotonin-1 receptor subtypes were defined with 8-hydroxy-2-(di-n-propylamino)-tetralin and mesulergine. In the control human basal ganglia, the highest density of serotonin-1 binding sites was observed in both lateral and medial globus pallidus and substantia nigra reticulata. Lower densities were seen in the substantia nigra pars compacta, the nucleus accumbens, caudate and putamen. The majority of these serotonin-1 sites belonged to the serotonin-1D class. No significant alteration of the density and distribution of these sites was observed in Parkinson's disease brains. In contrast, a marked decrease in the density of the receptor binding was seen in the basal ganglia and the substantia nigra from patients dying with Huntington's disease. These results suggest that serotonin-1D receptors are expressed by cells intrinsic to the striatum which degenerate in Huntington's disease and project to the substantia nigra reticulata where these receptors are probably presynaptically localized. These observations in pathological human brains agree with the results of lesion studies in animal models and further support a role for serotoninergic mechanisms in movement control.     

Association of dopamine D1 and D2 receptors with specific cellular elements in the basal ganglia of the cat: the uneven topography of dopamine receptors in the striatum is determined by intrinsic striatal cells, not nigrostriatal axons

To ascertain the cellular associations of the D1 and D2 dopamine receptor subtypes in components of the basal ganglia, cats were prepared with unilateral, axon-sparing, ibotenic acid lesions of the striatum (n = 6) or lesions of the nigrostriatal dopamine system by intranigral infusion of 6-hydroxydopamine (n = 8). After 42 days survival, tissue sections from the brains were processed for quantitative, in vitro receptor autoradiography with [3H]SCH23390 (D1 radioligand) or [3H]spiroperidol (D2 radioligand). Lesion-induced changes in basal ganglia nuclei were assessed by comparing them to the corresponding nuclei on the intact side and in naive brains. Ibotenate lesions cause a decline in specific D1 and D2 receptor-binding in the area of the striatal lesion of 94% and 85%, respectively, and completely eliminate the uneven patterns of high- and low-density binding that are characteristic of the cat's caudate nucleus. The globus pallidus, entopeduncular nucleus and pars reticulata of the substantia nigra also show marked reductions in binding after striatal ibotenate lesions. Thus, after caudate nucleus lesions, D2 binding in the two pallidal segments declines by approximately 50%, but remains unchanged in the substantia nigra. Binding of the D1 radioligand (which is not measurable in the globus pallidus) declines by about 75% in the affected regions of the entopeduncular nucleus and pars reticulata, and by about 30% in the pars compacta. Lesions of the nigral dopamine neurons reduce D2 receptor-binding by 95% in the pars compacta and 40% in the pars reticulata, but have no effect on the concentration of D1 or D2 radioligand-binding in the striatum or pallidum. Moreover, such lesions failed to alter the uneven patterns of binding in the striatum. These data suggest that most, if not all, D1 receptors in the basal ganglia are associated with cells of the striatum and their axons in the entopeduncular nucleus and substantia nigra, and likewise, a large majority of D2 receptors are associated with striatal cells and their axons in pallidal structures. Nearly all D2 receptors in the substantia nigra are associated with dopamine neurons (autoreceptors). Finally, the heterogeneous patterns of D1 and D2 receptors in the striatum are a consequence of intrinsic neuronal distributions.     

Different neuronal location of [3H]SCH 23390 binding sites in pars reticulata and pars compacta of the substantia nigra in the rat

The precise neuronal localization of D1 receptors in the substantia nigra has been studied autoradiographically in the rat by measuring the alterations of [3H]SCH 23390 binding site densities in this brain area after 6-hydroxydopamine (6-OHDA) induced destruction of nigrostriatal dopaminergic neurons and after ibotenate-induced lesion of striatal afferents. 6-OHDA-induced nigral lesion provoked a total loss of [3H]SCH 23390 binding sites in the pars compacta and pars lateralis (but not in the pars reticulata) of the substantia nigra. In contrast, ibotenate-induced striatal lesion caused a large diminution of the [3H]ligand binding site density in the pars reticulata but not in the pars compacta and pars lateralis of the substantia nigra. These results suggest that D1 receptors in the pars compacta or pars lateralis of the substantia nigra are located on the dopaminergic perikarya whereas those D1 receptors present in the pars reticulata of the substantia nigra lie on the terminals of nigral afferents of striatal origin.     

The AS/AGU rat: a spontaneous model of disruption and degeneration in the nigrostriatal dopaminergic system

The AS/AGU rat provides an alternative to experimentally produced laboratory models of basal ganglia disorders. This mutant is characterised by disturbances of movement including clumsy gait, whole body tremor, rigidity and difficulty in initiating movement. From an early age, there is a profound depletion of extracellular dopamine in the dorsal caudate-putamen as measured via in vivo microdialysis; levels are only 10–20% of those found in the parent Albino Swiss (AS) strain. Subsequently a depletion of whole tissue dopamine levels occurs and, later still, loss of dopaminergic cells in the substantia nigra pars compacta. The dysfunction in movement and the nigrostriatal dopaminergic system are clearly linked, since movement can be ameliorated by L -DOPA administration. Furthermore, there are depletions in glucose utilisation in several regions of the basal ganglia circuitry, including the substantia nigra pars compacta, the subthalamic nucleus and the ventrolateral thalamus. The AS/AGU rat represents a unique opportunity to investigate the intrinsic factors controlling the integrity of dopaminergic systems and the recent successful positional cloning of the agu gene will allow the molecular mechanisms underlying this interesting phenotype to be analysed.     

Neurokinin-1 and neurokinin-3 receptors in primate substantia nigra

Striatonigral axons co-release GABA and substance P (SP) at their target sites, but little is known about the action of SP at nigral level. Therefore, we studied immunohistochemically the cellular and subcellular localization of SP and its high affinity receptors neurokinin-1 (NK-1R) and neurokinin-3 (NK-3R) at nigral level in squirrel monkeys. Immunofluorescent studies revealed that, although SP+ fibers arborised more densely in the pars reticulata (SNr) than in the pars compacta (SNc), the two nigral divisions harbored numerous neurons expressing NK-1R and NK-3R. Confocal microscopic analyses showed that numerous SNr neurons and virtually all SNc dopaminergic neurons contained both NK-1R and NK-3R. At the electron microscope level, NK-1R and NK-3R were mainly associated with intracellular sites or located at extrasynaptic position on plasma membrane. A small proportion of SP+ boutons also showed NK-3R immunoreactivity. The distribution of NK-1R and NK-3R in SNr and SNc suggests that SP exerts its effect through postsynaptic receptors, as well as via presynaptic autoreceptors and heteroreceptors. These findings indicate that the excitatory peptide SP can modulate the inhibitory action of GABA at nigral level and suggest that the co-release of these two neuroactive substances should be taken into account when considering the functional organization of the basal ganglia.     

Changes in GABA(B) receptor mRNA expression in the rodent basal ganglia and thalamus following lesion of the nigrostriatal pathway

Loss of striatal dopaminergic innervation in Parkinson's disease (PD) is accompanied by widespread alterations in GABAergic activity within the basal ganglia and thalamus. Accompanying changes in GABA_B receptor binding have been noted in some basal ganglia regions in parkinsonian primates, suggesting that plasticity of this receptor may also occur in PD. However, the molecular mechanisms underlying the changes in receptor binding and the manner and extent to which different GABA_B receptor mRNA subunits and splice-variants are affected remain unknown. This study used in situ hybridisation to examine the full profile of changes in expression of the known rat GABA_B receptor genes and gene variants in the basal ganglia and thalamus of rats, brought about by degeneration of the nigrostriatal tract. All of the GABA_B mRNA species examined showed unique expression patterns throughout the basal ganglia and thalamus. In addition, all exhibited a marked loss of expression (between 46 and 80%) in the substantia nigra pars compacta of animals bearing a complete 6-hydroxydopamine-induced lesion of the nigrostriatal tract, confirming the presence of these variants in dopaminergic neurones in this region. Further analysis of autoradioagrams revealed additional changes only in GABA_B(1a) mRNA in discrete anatomical regions. Expression of the GABA_B(1a) variant was significantly increased in the substantia nigra pars reticulata (33±2%), entopeduncular nucleus (26±1%) and the subthalamic nucleus (16±1%). Since these regions all receive reduced GABAergic innervation following nigrostriatal tract lesioning, it is possible that the increased expression occurs as a compensatory measure. In conclusion, these data demonstrate that GABA_B receptor genes exhibit regional- and subunit/variant-specific plasticity at the molecular level under parkinsonian conditions.     

Functional changes of the basal ganglia circuitry in Parkinson's disease

The basal ganglia circuitry processes the signals that flow from the cortex, allowing the correct execution of voluntary movements. In Parkinson's disease, the degeneration of dopaminergic neurons of the substantia nigra pars compacta triggers a cascade of functional changes affecting the whole basal ganglia network. The most relevant alterations affect the output nuclei of the circuit, the medial globus pallidus and substantia nigra pars reticulata, which become hyperactive. Such hyperactivity is sustained by the enhanced glutamatergic inputs that the output nuclei receive from the subthalamic nucleus. The mechanisms leading to the subthalamic disinhibition are still poorly understood. According to the current model of basal ganglia organization, the phenomenon is due to a decrease in the inhibitory control exerted over the subthalamic nucleus by the lateral globus pallidus. Recent data, however, suggest that additional if not alternative mechanisms may underlie subthalamic hyperactivity. In particular, given the reciprocal innervation of the substantia nigra pars compacta and the subthalamic nucleus, the dopaminergic deficit might influence the subthalamic activity, directly. In addition, the increased excitatory drive to the dopaminergic nigral neurons originating from the hyperactive subthalamic nucleus might sustain the progression of the degenerative process. The identification of the role of the subthalamic nucleus and, more in general, of the glutamatergic mechanisms in the pathophysiology of Parkinson's disease might lead to a new approach in the pharmacological treatment of the disease. Current therapeutic strategies rely on the use of L-DOPA and/or dopamine agonists to correct the dopaminergic deficit. Drugs capable of antagonizing the effects of glutamate might represent, in the next future, a valuable tool for the development of new symptomatic and neuroprotective strategies for therapy of Parkinson's disease.     

Expression of cholecystokinin, somatostatin, thyrotropin-releasing hormone, glutamic acid decarboxylase and tyrosine hydroxylase genes in the central nervous motor systems of the genetically dystonic hamster

In the dt ^sz mutant hamster with idiopathic generalized dystonia, functional abnormalities of several neurotransmitters have been suggested to play a role in the development of symptoms. In the present study, we have used histochemistry with ³⁵S-ATP labeled oligonucleotides to determine whether these abnormalities are associated with modulation in the expression of neurotransmitter genes in motor regions. We examined the expression of genes encoding cholecystokinin (CCK), somatostatin (SRIF), thyrotropin-releasing hormone (TRH), glutamic acid decarboxylase (GAD), tyrosine hydroxylase (TH) and growth-associated protein 43 (GAP43) in the cortex and basal ganglia of dystonic hamsters and of non-dystonic control hamsters of a related inbred line and of a non-related outbred line. The distribution of these mRNAs in normal hamster brain was similar to that in normal rat brain. In all cortical regions studied (frontal, parietal and piriformis), the expression of CCK was similar in dystonic and inbred controls but was significantly greater than in outbred controls. In the anterior thalamus, CCK expression was lower in dystonic hamsters than in both control groups. SRIF expression was significantly decreased in the cortex and striatum of dystonic animals than in inbred and outbred control hamsters. GAD expression was lower in the striatum and substantia nigra, pars reticulata of dystonic than in outbred hamsters, but similar values were found in all groups in the other regions studied. TH was lower in the substantia nigra of dystonic than in inbred controls. No changes were found in GAP43 expression. This study demonstrates that changes in modulation of the expression of some peptides and neurotransmitter enzymes can be found in the dystonic hamster, which is in contrast to other animal models such as the dystonic rat, where no such changes have been found. The present data are consistent with previous findings in dt ^sz hamsters that suggest a dysfunction within the basal ganglia-thalamocortical circuits. Received: 29 June 1998 / Accepted: 17 May 1999     

Decreased adenosine receptor binding in dystonic brains of the dt(sz) mutant

In patients with paroxysmal non-kinesigenic dyskinesias, episodes of dystonia can be provoked by stress and also by methylxanthines (e.g. caffeine), which inhibit adenosine A(1)/A(2A) receptors. In the dt(sz) mutant hamster, a model of this movement disorder, adenosine A(1) receptor antagonists were previously found to worsen dystonia, while adenosine A(1) and A(2A) receptor agonists exerted pronounced beneficial effects. Therefore, in the present study, adenosine receptor A(1) and A(2A) binding was determined by autoradiographic analyses in dt(sz) hamsters under basal conditions, i.e. in the absence of a dystonic attack, and in a group of mutant hamsters which exhibited severe stress-induced dystonic attacks prior to kill. In comparison with non-dystonic control hamsters, [(3)H]DPCPX (8-cyclopentyl-1,3-dipropylxanthine) binding to adenosine A(1) receptors and [(3)H]CGS 21680 (2p-(2carboxyethylphen-ethylamino-5'-N-ethlycarboxamindoadenosine) binding to adenosine A(2A) receptors were significantly lower throughout the brain of dystonic animals. Under normal resting conditions, mutant hamsters showed significant decreases in adenosine A(1) (-12 to-42%) and in A(2A) (-19 to-34%) receptor binding compared with controls. Stressful stimulation increased adenosine A(1) and A(2A) receptor binding in almost all brain regions in both control and dystonic hamsters. The stress-induced increase was more marked in mutant hamsters, leading to a disappearance of differences in most regions compared with stimulated controls, except the striatum. In view of previous findings of striking beneficial effects of adenosine A(1) and A(2A) receptor agonists and of striatal dysfunctions in the dt(sz) mutant, the reduced adenosine receptor binding may be an important factor in the pathogenesis of paroxysmal dystonia.     

A direct projection from superior colliculus to substantia nigra pars compacta in the cat

Dopaminergic neurons exhibit a short-latency, phasic response to unexpected, biologically salient stimuli. The midbrain superior colliculus also is sensitive to such stimuli, exhibits sensory responses with latencies reliably less than those of dopaminergic neurons, and, in rat, has been shown to send direct projections to regions of the substantia nigra and ventral tegmental area containing dopaminergic neurons (e.g. pars compacta). Recent electrophysiological and electrochemical evidence also suggests that tectonigral connections may be critical for relaying short-latency (<100 ms) visual information to midbrain dopaminergic neurons. By investigating the tectonigral projection in the cat, the present study sought to establish whether this pathway is a specialization of the rodent, or whether it may be a more general feature of mammalian neuroanatomy. Anterogradely and retrogradely transported anatomical tracers were injected into the superior colliculus and substantia nigra pars compacta, respectively, of adult cats. In the anterograde experiments, abundant fibers and terminals labeled with either biotinylated dextran amine or Phaseolus vulgaris leucoagglutinin were seen in close association with tyrosine hydroxylase-positive (dopaminergic) somata and processes in substantia nigra pars compacta and the ventral tegmental area. In the retrograde experiments, injections of biotinylated dextran amine into substantia nigra produced significant retrograde labeling of tectonigral neurons of origin in the intermediate and deep layers of the ipsilateral superior colliculus. Approximately half of these biotinylated dextran amine-labeled neurons were, in each case, shown to be immunopositive for the calcium binding proteins, parvalbumin or calbindin. Significantly, virtually no retrogradely labeled neurons were found either in the superficial layers of the superior colliculus or among the large tecto-reticulospinal output neurons. Taken in conjunction with recent data in the rat, the results of this study suggest that the tectonigral projection may be a common feature of mammalian midbrain architecture. As such, it may represent an additional route by which short-latency sensory information can influence basal ganglia function.     

Signalling through phospholipase C beta 4 is not essential for midbrain dopaminergic neuron survival

The most prominent progressive neurodegenerative movement disorder, Parkinson's disease, is attributed to selective loss of dopamine neurons in the substantia nigra pars compacta, resulting in severe deficiency of dopamine. The homeo-domain gene, Pit x 3, is essential for proper development of midbrain dopaminergic neurons in the substantia nigra pars compacta and might be involved in midbrain dopaminergic survival pathways. The mGluR1-signaling downstream-effector phospholipase C beta 4 was identified in a suppression subtractive hybridization screen comparing wild-type and Pit x 3-deficient Aphakia midbrain dopaminergic neurons. Expression pattern analysis revealed that phospholipase C beta 4 was expressed in midbrain dopaminergic neurons of the substantia nigra pars compacta and part of the ventral tegmental area, whereas expression of mGluR1alpha was predominantly observed in the more vulnerable midbrain dopaminergic neurons in the lateral substantia nigra pars compacta. However, clear expression of phospholipase C beta 4 in spared midbrain dopaminergic neurons of Aphakia mice located in the ventral tegmental area, indicated that induction and maintenance of phospholipase C beta 4 expression is Pit x 3-independent in these neurons. Furthermore, we report here a normal distribution of midbrain dopaminergic cell bodies and axonal projection to the striatum in phospholipase C beta 4-/- mice, indicating that signaling of phospholipase C beta 4 is not essential for the survival of midbrain dopaminergic neurons.     

Regional distribution of cholecystokinin binding sites in macaque basal ganglia determined by in vitro receptor autoradiography

Cholecystokinin binding sites were labeled with [3H]cholecystokinin-8, [125I]cholecystokinin-33, and [125I]cholecystokinin-8 in major structures of macaque basal ganglia by in vitro receptor autoradiography. Analysis of autoradiograms revealed areas of heavy cholecystokinin binding in the neostriatum and substantia nigra that were set off, often quite sharply, from the adjacent globus pallidus and subthalamic nucleus where labeling was, by contrast, very light. Heavy label characterized the ventromedial and posterior parts of the caudate nucleus and adjacent putamen, binding was of moderate intensity in central areas of these regions, while, the dorsolateral margin of the head of the caudate and precommissural putamen, the dorsolateral one-third of the body of the caudate, and all but the most medial and ventral portions of the posterior putamen lateral to the pallidum were sparsely labeled. The pattern of cholecystokinin binding within the neostriatum was mottled; patches of reduced label stood out from the background of more prominent binding. However, those patches were only imperfectly correlated with the striosomal organization of both the caudate nucleus and putamen as revealed by acetylcholinesterase staining. Cholecystokinin binding in the substantia nigra was also intricately patterned. Moderately dense, vertically orientated bands of label were found in the dorsal one-third to half of the pars reticulata, providing a marked contrast to the near background levels in the ventral pars reticulata and overlying pars compacta. The present study shows that heavy cholecystokinin binding is confined to particular areas within the primate basal ganglia; the pattern of label within the substantia nigra and neostriatum can be linked to intrinsic and afferent connections of these structures. The confinement of binding sites to the dorsal pars reticulata suggests an association with dendrites of pars compacta neurons which invade this region; this interpretation is consistent with recent evidence of depletion of nigral cholecystokinin binding sites in macaques following chemical lesion of dopaminergic cells of the par compacta. In the neostriatum the distribution of binding shows overlap with its topographically organized corticostriatal innervation; portions of heavily labeled striatum coincide with regions innervated by association cortex of the frontal and temporal lobes, whereas regions of diminished binding correspond to areas innervated mainly by sensory and motor cortex. These latter findings suggest that cholecystokinin may have a particularly strong influence on cognitive aspects of striatal function.     

Time of neuron origin and gradients of neurogenesis in midbrain dopaminergic neurons in the mouse

Previous [³H]thymidine studies in Nisslstained sections in rats established that the substantia nigra pars compacta and the ventral tegmental area originate sequentially according to an anterolateral to posteromedial neurogenetic gradient. We investigated whether that same pattern is found in mice in the dopaminergic neurons in each of these structures. Using tyrosine hydroxylase immunostaining combined with [³H]thymidine autoradiography, the time of origin of dopaminergic midbrain neurons in the retrorubral field, the substantia nigra pars compacta, the ventral tegmental area, and the interfascicular nucleus was determined in postnatal day 20 mice. The dams of the experimental animals were injected with [³H]thymidine on embryonic days (E) 11–E12, E12–E13, E13–E14, and E14–E15. The time of origin profiles for each group indicated significant differences between populations. The retrorubral field and the substantia nigra pars compacta arose nearly simultaneously and contained the highest proportion of neurons, 49 to 37%, generated on or before E11. Progressively fewer early-generated neurons were found in the ventral tegmental area (20%), and the interfascicular nucleus (8.5%). In addition, anterior dorsolateral neurons in the substantia nigra and ventral tegmental area were more likely to be generated early than the posterior ventromedial neurons. These findings indicate that mouse and rat brains have nearly identical developmental patterns in the midbrain, and neurogenetic gradients in dopaminergic neurons are similar to those found in Nissl studies in rats.     

Origin,course and termination of dopaminergic substantia nigra neurons projecting to the amygdaloid complex in the cat

The organization of dopaminergic neurons projecting to the amygdala was examined using retrograde (horseradish peroxidase histochemistry) and anterograde ([³H]leucine autoradiography) transport methods and Falck-Hillarp histofluorescence techniques combined with microspectrofluorometry and radiofrequency lesions. Cell bodies located within the pars lateralis and pars compacta of the substantia nigra were found to project to the lateral and central amygdaloid nuclei, respectively. Both of these areas within the substantia nigra contained dopaminergic perikarya, while the central and lateral amygdaloid nuclei contained fluorescent varicosities with features indicative of dopaminergic neurons. Lesions restricted to the pars lateralis of the substantia nigra resulted in a loss of fluorescence in the lateral amygdaloid nucleus. Autoradiographic experiments revealed that the projections from the pars lateralis did not run with fibers originating from the pars compacta in the nigrostriatal tract but rather had their own course occupying a lateral position adjacent to the cerebral peduncle and joining the ventral amygdalo-fugal bundle.The data indicate that, in the cat, there are two separate dopaminergic projections to the amygdala arising from the substantia nigra.     

Innervation of substantia nigra neurons by cholinergic afferents from pedunculopontine nucleus in the rat: neuroanatomical and electrophysiological evidence

Dopaminergic neurons of the substantia nigra pars compacta are excited by nicotine and acetylcholine, and possess both high-affinity nicotine binding sites and intense acetylcholinesterase activity, consistent with a cholinoceptive role. A probable source of cholinergic afferents is the pedunculopontine nucleus, which forms part of a prominent group of cholinergic perikarya located caudal to the substantia nigra in the tegmentum. Although pedunculopontine efferents, many of them cholinergic, project to the substantia nigra pars compacta, it has not been established whether they terminate in this structure. In the first experiment, which combined retrograde tracing with immunohistochemical visualization of cholinergic neurons, cholinergic cells in and around the pedunculopontine nucleus were found to send projections to the substantia nigra. This projection was almost completely ipsilateral. Subsequent experiments employed anaesthetized rats; kainate was microinfused into tegmental sites in order to stimulate local cholinergic perikarya, and concurrently, extracellular recordings were made of single dopaminergic neurons in the substantia nigra. Consistent with our anatomical findings, unilateral microinfusion of kainic acid in or near the pedunculopontine nucleus increased the firing rate of dopaminergic neurons situated remotely in the ipsilateral substantia nigra. The kainate-induced excitation of nigral dopaminergic neurons was dose-related and was prevented by intravenous administration of the centrally-acting nicotinic cholinergic antagonist mecamylamine. These results suggest that cholinergic perikarya in the vicinity of the pedunculopontine tegmental nucleus innervate dopaminergic neurons in the substantia nigra pars compacta via nicotinic receptors.     

Extracellular amino acid levels in the striatum of the dt(sz) mutant, a model of paroxysmal dystonia

The pathophysiology of idiopathic dystonia is still unknown, but it is regarded as a basal ganglia disorder. Previous studies indicated an involvement of a striatal GABAergic disinhibition and a cortico-striatal glutamatergic overactivity in the manifestation of stress-inducible dystonic episodes in the dt(sz) hamster, a model of idiopathic paroxysmal dystonia. These investigations were carried out postmortem or in anesthetized animals. In the present study, in vivo microdialysis in conscious, freely-moving dt(sz) and non-dystonic control hamsters was used to examine the levels of GABA, aspartate, glutamate, glutamine, glycine and taurine in each animal during following conditions: (1) at baseline in the absence of dystonia, (2) during an episode of paroxysmal dystonia precipitated by stressful stimuli, (3) during a recovery period and (4) at baseline after complete recovery. In comparison to non-dystonic controls, which were treated in the same manner as the dystonic animals, no differences could be detected under basal conditions. The induction of a dystonic episode in mutant hamsters led to higher contents of glycine in these animals in comparison to stressed but non-dystonic controls. Significant changes of glycine levels within the animal groups were not detected. The levels of the excitatory amino acids glutamate, glutamine and aspartate as well as the levels of the inhibitory amino acids GABA and taurine did not differ between the animal groups or between the periods of measurement. The higher levels of glycine might contribute to the manifestation of paroxysmal dystonia in dt(sz) hamsters, although unaltered glutamate, glutamine and aspartate levels do not support the hypothesis of a critical involvement of a cortico-striatal overactivity. It seems that a deficiency of GABAergic interneurons, found by previous immunohistochemical examinations, does not lead to reduced extracellular GABA levels in the striatum.     

Synaptic localization of ionotropic glutamate receptors in the rat substantia nigra

Glutamatergic neurotransmission in the substantia nigra pars compacta and pars reticulata is mediated through N-methyl-D-aspartate and alpha-amino-3-hydroxy-5-methyl-4-isoxaline propionic acid/kainate (AMPA) type receptors as well as other glutamate receptors and is critical for basal ganglia functioning. A major glutamatergic input to the substantia nigra originates in the subthalamic nucleus, and the long-lasting stimulation of the dopaminergic cells of the substantia nigra pars compacta by the subthalamic neurons has been implicated in the pathophysiology of Parkinson's disease. The objectives of the present study were to determine the subcellular and subsynaptic localization of subunits of the N-methyl-D-aspartate and AMPA receptors in the substantia nigra, and also to determine whether co-localization of N-methyl-D-aspartate and AMPA receptor subunits occur at individual synapses. To achieve this, pre-embedding and post-embedding immunocytochemistry was applied to sections of substantia nigra using antibodies that recognize the NR1 and NR2A/B subunits of the N-methyl-D-aspartate receptor, and GluR2/3 subunits of the AMPA receptor.In both regions of the substantia nigra, immunolabelling for each of the subunits was observed in numerous perikarya and proximal dendrites. At the subcellular level, silver-intensified immunogold particles localizing N-methyl-D-aspartate and AMPA receptor subunits were most commonly present within dendrites where they were associated with a variety of intracellular organelles and with the internal surface of the plasma membrane. Post-embedding immunogold labelling revealed immunoparticles labelling for NR1, NR2A/B and GluR2/3 to be enriched at asymmetric synaptic specializations, although a large proportion of asymmetric synapses were immunonegative. Double immunolabelling revealed, in addition to single-labelled synapses, the co-localization of subunits of the N-methyl-D-aspartate receptor and subunits of the AMPA receptor at individual asymmetric synapses. Similarly, double immunolabelling also revealed the co-localization of the NRl and NR2A/B subunits of the N-methyl-D-aspartate receptor at individual asymmetric synapses. Labelling for NR1 and GluR2/3 was, on average, relatively evenly distributed across the width of the synapse with a gradual reduction towards the periphery when analysed in single sections.In summary, the present results demonstrate that AMPA and N-methyl-D-aspartate receptors are selectively localized at a subpopulation of asymmetric synapses in the substantia nigra pars compacta and reticulata and that the two receptor types, at least partially co-localize at individual synapses. It is concluded that glutamatergic transmission in the substantia nigra pars compacta and pars reticulata occurs primarily at asymmetric synapses and, at least in part, is mediated by both N-methyl-D-aspartate and AMPA receptors.     

Bilateral cerebral metabolic effects of pharmacological manipulation of the substantia nigra in the rat: Unilateral intranigral application of the putative excitatory neurotransmitter substance P

The metabolic activity of several anatomically distinct brain areas was investigated by means of the quantitative autoradiographic 2-deoxy-d[1-¹⁴C]glucose method in awake rats following unilateral intranigral application of the putative excitatory neurotransmitter substance P. The primary goal was to determine the metabolic effects of substance P on the substantia nigra and its targets. Intranigral injection of 1 mM substance P (1.5 μl) induced metabolic activation locally in the substantia nigra reticulata by 117% and substantia nigra compacta by 35%, as well as distally in the contralateral substantia nigra reticulata by 22% and contralateral substantia nigra compacta by 21%. All the basal ganglia components, the striatum, pallidum, entopeduncular, subthalamic nucleus and nucleus accumbens displayed bilateral metabolic activations after unilateral intranigral substance P injection. Among the principal reticulata efferent projections, the ventromedial, ventrolateral, parafascicular, mediodorsal and centrolateral thalamic nuclei, as well as the pedunculopontine nucleus displayed bilateral metabolic activations after intranigral substance P application. Moreover, unilateral intranigral substance P injection elicited metabolic activations in the thalamic and cortical components of the reticular, intralaminar, limbic and prefrontal systems, mostly bilateral.

It is suggested that substance P applied into one substantia nigra reticulata activates the compacta nigrostriatal dopaminergic and the reticulata nigrothalamic GABAergic outputs inducing distal metabolic effects, similar to those elicited by unilateral nigral electrical stimulation [Savaki et al. (1983) J. comp. Neurol.213, 46–65] and, opposite to several of those induced by intranigral injection of the inhibitory GABA_A agonist muscimol [Savaki et al. (1992) Neuroscience50, 781–794]. Furthermore, it is suggested that the ipsilateral basal ganglia effects induced by intranigral substance P application are mediated via both the compacta dopaminergic nigrostriatal projection and the reticulata GABAergic nigro-thalamo-cortico-striatal loop, whereas the contralateral basal ganglia and associated thalamocortical effects are due to the activation of the GABAergic reticulata efferents and are mediated via an interthalamic circuitry involving the motor, reticular and intralaminar thalamic nuclei.     

Considerations upon the anatomical model of reward-based learning in the basal ganglia

The nigrostriatal pathway appears to be very important in the reward-based learning. The dopaminergic neurons in the substantia nigra pars compacta (SNC) fire in relation to primary rewards and reward-conditioned stimuli, but not to rewards that are expected. It has been hypothesized that the anatomical framework for the selective response of these neurons is organized in the projections from some paralimbic areas in the frontal lobe to the striosomes of the caudate nucleus, which are also directly connected with the dopaminergic neurons of the SNC. Here, we present two additional pathways that may be related with this neurophysiological finding. We hypothesize that the connections of the paralimbic cortices with the ventral system of the basal ganglia and then with the thalamus and the hypothalamus, and the circuit ventral striatum-substantia nigra pars reticulata-thalamus-striatum could be also involved in the reward-based learning.