5-HT1D receptors in the guinea pig brain: pre- and postsynaptic localizations in the striatonigral pathway

The caudate-putamen, globus pallidus and substantia nigra pars reticulata of the guinea pig contain high densities of the 5-HT1D receptor subtype. The cellular localization of these sites in the striatonigral pathway was investigated using receptor autoradiography and selective neurotoxin lesions. In guinea pigs with unilateral 6-hydroxydopamine lesions of the nigral dopaminergic cells, no significant decrease was observed in any of the components of the striatonigral pathway. In contrast, when quinolinic acid was injected in the caudate-putamen, marked reductions in [3H]5-HT binding were seen in the caudate-putamen, the globus pallidus and the substantia nigra pars reticulata, on the side ipsilateral to the lesion. These data, which are comparable to previous results in human pathologies where similar cell populations are known to degenerate (Parkinson disease and Huntington's chorea), indicate a presynaptic localization of 5-HT1D receptors on the terminals of the striatal neurons projecting to the pars reticulata of the substantia nigra. In addition, these receptors could be located on the cell bodies or dendrites of these neurons in the striatum, postsynaptically to serotoninergic fibers.     

Substantia nigra opiate receptors on basal ganglia efferents

Many behavioral effects of opiate narcotics and peptides have been linked to effects on dopamine neurons originating in the substantia nigra pars compacta and ventral tegmental area. Selective brain lesions were combined with quantitative autoradiography to determine whether opiate receptors are on dopaminergic somata and/or processes in the substantia nigra pars compacta and ventral tegmental area. 6-Hydroxydopamine lesions that eliminated dopamine neurons produced little change in the pattern or density of [³H]-naloxone binding in the substantia nigra pars compacta or ventral tegmental area. Radiofrequency lesions of the internal capsule or globus pallidus and kainic acid lesions of the striatum markedly decreased [³H]-naloxone binding in the pars compacta and pars reticulata. These results are consistent with a dense distribution of opiate receptors on pallido-nigral and/or striato-nigral fibers and strengthen the likelihood that local effects of opiates on dopamine function in the nigrostriatal pathway are mediated indirectly by actions on nondopaminergic processes.     

Changes in [3H]zolpidem and [3H]Ro 15-1788 binding in rat globus pallidus and substantia nigra pars reticulata following a nigrostriatal tract lesion

Changes in GABA(A) receptor alpha(1) subunit gene expression occur in the globus pallidus and substantia nigra pars reticulata following lesions of the nigrostriatal tract. To determine whether these changes are translated at the protein level, we performed quantitative autoradiography with the alpha(1) selective ligand, [3H]zolpidem, and the non-selective benzodiazepine site ligand, [3H]Ro 15-1788. Binding of both [3H]zolpidem and [3H]Ro 15-1788 was significantly increased in the substantia nigra pars reticulata (13. 5+/-4.1 and 26.3+/-2.9%, respectively) and significantly reduced in the globus pallidus (20.9+/-0.8 and 18.3+/-1.3%, respectively). These changes in alpha(1) subunit protein expression may help to compensate for the pathological changes in GABAergic activity that occur after striatal dopamine depletion.     

Visualization of a dopamine D1 receptor mRNA in human and rat brain.

Using 32P-labeled oligonucleotides derived from the coding region of human dopamine D1 receptor mRNA we have localized in the human and rat brain the cells containing the mRNAs coding for this receptor. Dopamine D1 receptor mRNA in human brain was found to be contained in the neurons of the caudate and putamen nuclei as well as in the nucleus accumbens, some cortical regions and some nuclei of the amygdala. In the rat brain, cells containing D1 receptor mRNA were enriched in caudate-putamen and accumbens nuclei, olfactory tubercle, islands of Calleja, some cortical areas and in several thalamic nuclei. Moreover, in both species, it was absent from the neurons of the substantia nigra both pars compacta and pars reticulata and ventral tegmental area as well as from the globus pallidus pars lateralis and medialis in human and globus pallidus and entopeduncular nucleus in rat. In general, a good agreement was found with the distribution of binding sites labeled with the D1 antagonist SCH 23390. The main exception was the absence of D1 receptor mRNA in globus pallidus and substantia nigra, regions where high densities of receptor sites are found. These data support the notion that sites in these two regions are localized to projections from striatal neurons and that dopaminergic neurons do not express this receptor.     

Non-dopaminergic nigrostriatal pathway

The nigrostriatal projection was studied with a retrograde tracing method (Evans blue, EB) combined with a technique for dopamine histofluorescence. The study, realized in control rats and in animals with 6-hydroxydopamine-induced lesions of the dopaminergic pathway, yielded the following results.

(1) In 3 control rats injected with 0.2 μl of a 10% solution of EB in thecenter of the caudate-putamen 1 mm anterior to the globus pallidus, 96% of all substantia nigra neurons retrogradely labelled with the dye contained dopamine fluorescence. The remaining ones (average 350 per brain) were devoid of dopamine fluorescence and predominantly found in the posterior 75% of the substantia nuigra. These last cells were confined to the upper-half of the pars reticulata.

(2) In a series of 6 animals, the cytotoxic agent 6-hydroxydopamine was injected in various locations in the vicinity of either the substantia nigra ir the nigrostriatal tract 12–15 days prior to the injections of 0.2 μl of EB in the same striatal locations as in the controls. Despite a reduction of up to 85% in the number of dopaminergic cell bodies, the substantia nigra of these rats contained the same average number of EB-labelled neurons devoid of dopamine fluorescence.

(3) Eight rats received smaller injections (0.1 μl) of EB in various striatal sites and in tqo further cases such injections were placed in the globus pallidus to determine more accurately the anatomical location of the dopamine-negative nigral neurons retrogradely labelled with the dye. Following the striatal injections, these cells were found mostly in the upper-half of the pars reticulata and were arranged in longitudinally oriented clusters whose mediolateral location depended on the striatal injection site.

Following the pallidal injections, retrogradely labelled neruons devoid of dopamine fluorescence were found in greater numbers and were located in all areas of the pars reticulata. The possibility of retrograde labelling of some nigrothalamic neurons was not entirely ruled out in these two cases.

(4) Finally 6 rats received 0.1 μl injections of EB in various parts of the parietal cortex. In these cases the substantia nigra did not contain any EB-positivedopamine-negative neurons.

These results are interpreted as evidence in support of the existence of a topographically organized non-dopaminergic nigrostriatal projection.

Evidence for the presynaptic localization of opiate binding sites on striatal efferent fibers

Using quantitative receptor autoradiography, [3H]D-Ala-D-Leu-enkephalin (DADL) and [3H]naloxone binding were studied in rat striatum and striatal projection areas (globus pallidus (GP) and substantia nigra pars reticulata (SNr] after unilateral striatal kainic acid lesions. [3H]DADL and [3H]naloxone binding were each examined by two methods. Initially, [3H]DADL binding was performed in 50 mM Tris-HCl (pH 7.4), 30 mM NaCl, 3 mM manganese acetate and 2 microM GTP; [3H]naloxone binding was carried out in 50 mM Tris-HCl (pH 7.4) and 100 mM NaCl. Subsequent studies were carried out in 150 mM Tris-HCl (pH 7.4) and either [3H]DADL plus 500 nM morphiceptin (to block [3H]DADL binding to mu receptors) or [3H]naloxone plus 10 nM delta receptor peptide (to block [3H]naloxone binding to delta receptors). At one and eight weeks in the lesioned striatum, [3H]DADL binding was reduced by 70% and 82%, respectively, when compared to the control side. [3H]Naloxone binding was reduced by 35% and 20%. In GP and SNr, [3H]DADL binding was reduced by 31% and 41%, respectively, at one week and 27% and 26% at eight weeks. [3H]Naloxone binding was reduced 19% in GP at eight weeks. A parsimonious explanation of these results is that opiate binding sites are located on presynaptic terminals of striatal efferent fibers to globus pallidus and substantia nigra pars reticulata as well as on local striatal axon collaterals. Since opiate peptides have recently been found to coexist with GABA in some striatal neurons, opiate peptides may play a role in striatal function by controlling GABA release from striatal efferent fibers. It is possible that pallidal and nigral opiate binding could be utilized as a marker for striatal terminals.     

Lesion Study of the Distribution of Serotonin 5-HT4 Receptors in Rat Basal Ganglia and Hippocampus

The regional distribution of 5-hydroxytryptamine (5-HT₄) receptors labelled with [³H]GR113808 was examined in rat basal ganglia and hippocampus after specific lesions. Lesion of serotonin neurons induced by injections of 5,7-dihydroxytryptamine into the dorsal and medial raphe nuclei resulted in increased 5-HT₄ receptor binding in most regions examined, compared with controls. More precisely, there was a 78% increase in the rostral but no change in the caudal part of caudate-putamen, and 83% and 54% increases in the shell and core of the nucleus accumbens respectively. In the substantia nigra, the increase in 5-HT₄ binding was larger (72%) than that in the globus pallidus (32%). In the hippocampus, 63%, 30% and 28% increases were measured in CA2, CA1 and CA3 respectively. Following lesion of dopamine neurons by intranigral injection of 6-hydroxydopamine, increased 5-HT₄ receptor binding was observed in the caudal (59%), but not the rostral part of caudate-putamen, as well as in the globus pallidus (93%). Since no decreases in 5-HT₄ receptor density were detected after the dopamine lesion, it was concluded that these receptors are not expressed in dopamine neurons. Kainic acid lesions of the caudate-putamen were associated with dramatic local decreases in 5-HT₄ receptor binding on the injected side (-89%), which suggested that striatal neurons express 5-HT₄ receptors. Corresponding decreases of 72 and 20% in receptor density were detected in globus pallidus and substantia nigra, consistent with a presumed localization of 5-HT₄ receptors on striatal GABA neurons projecting to these regions. In the substantia nigra, the decrease in [³H]GR113808 binding was localized to the pars lateralis, indicating that striatal neurons belonging to the cortico-striato-nigrotectal pathway, and containing GABA and dynorphin, express 5-HT₄ receptors.     

Changes in [3H]muscimol binding in substantia nigra, entopeduncular nucleus, globus pallidus, and thalamus after striatal lesions as demonstrated by quantitative receptor autoradiography

A quantitative autoradiographic technique for measuring the binding of [3H]muscimol to central nervous system GABA receptors is described using tritium-sensitive film. [3H]Muscimol binding was studied in primary and secondary striatal projection areas of rat brain following kainic acid lesions of the striatum. Seven days after the lesion, binding affinities in the striatum and its projection areas were not altered significantly. There was a loss of [3H]muscimol receptors in the striatum. Receptors increased in numbers in the ipsilateral globus pallidus (19%), entopeduncular nucleus (22%), and substantia nigra pars reticulata (38%). [3H]Muscimol binding was decreased in the ipsilateral anteroventrolateral and ventromedial (8%) thalamic nuclei. [3H]Muscimol binding in other brain areas (layer IV of the cerebral cortex, central gray, superior colliculus, and stratum moleculare of hippocampus) was not affected. The findings suggest that a loss of striatal innervation resulted in increased numbers of GABA receptors in striatal projection sites. It is further suggested that loss of inhibitory striatal inputs to neurons in the entopeduncular nucleus and substantia nigra pars reticulata may activate GABAergic projections to thalamus and thus result in decreased numbers of thalamic GABA receptors.     

Cannabinoid receptor and WIN-55,212-2-stimulated [35S]GTPγS binding and cannabinoid receptor mRNA levels in the basal ganglia and the cerebellum of adult male rats chronically exposed to Δ9-tetrahydrocannabinol

The inhibition of motor behavior in rodents caused by the exposure to plant or synthetic cannabinoids has been reported to develop tolerance after repeated exposure. This tolerance seems to have a pharmacodynamic basis, since downregulation of cannabinoid receptors in motor areas, basal ganglia and cerebellum, has been demonstrated in cannabinoid-tolerant rats. The present study was designed to further explore this previous evidence by analyzing simultaneously in several motor areas of delta 9-tetrahydrocannabinol- (delta 9-THC)-tolerant rats: 1. Cannabinoid receptor binding, by using [3H]WIN-55,212-2 autoradiography; 2. Cannabinoid receptor activation of signal transduction mechanisms, by using WIN-55,212-2-stimulated [35S]-guanylyl-5'-O-(gamma-thio)-triphosphate ([35S]-GTP gamma S) autoradiography; 3. Cannabinoid receptor mRNA expression, quantitated by in situ hybridization. Results were as follows. As expected, the exposure to delta 9-THC for 5 d resulted in a decrease of cannabinoid receptor binding in the molecular layer of the cerebellum, medial, and lateral caudate-putamen and, in particular, entopeduncular nucleus. We also found decreased cannabinoid receptor binding in the superficial and deep layers of the cerebral cortex, two regions used as a reference to test the specificity of changes observed in motor areas. There were only two brain regions, the globus pallidus and the substantia nigra, where the specific binding for cannabinoid receptors was unaltered after 5 d of a daily delta 9-THC administration. However, in the substantia nigra, the magnitude of WIN-55,212-2-stimulated [35S]-GTP gamma S binding was lesser in delta 9-THC-tolerant rats than controls, thus suggesting a possible specific change at the level of receptor coupling to GTP-binding proteins. This was not seen neither in the globus pallidus nor in the lateral caudate-putamen, where agonist stimulation produced similar [35S]-GTP gamma S binding levels in delta 9-THC-tolerant rats and controls. Finally, animals chronically exposed to delta 9-THC also exhibited a decrease in the levels of cannabinoid receptor mRNA in the medial and lateral caudate-putamen, but there were no changes in the cerebellum (granular layer) and cerebral cortex. In summary, the chronic exposure to delta 9-THC resulted in a decrease in cannabinoid receptor binding and mRNA levels in the caudate-putamen, where cell bodies of cannabinoid receptor-containing neurons in the basal ganglia are located. However, this decrease particularly affected the receptor binding levels in those neurons projecting to the entopeduncular nucleus, but not in those projecting to the globus pallidus and substantia nigra, although, in this last region, a specific decrease in the efficiency of receptor activation of signal transduction mechanisms was seen in delta 9-THC-tolerant rats. The chronic exposure to delta 9-THC also resulted in decreased cannabinoid receptor binding in the cerebellum, although without affecting mRNA expression.     

Comparative cellular distribution of GABAA and GABAB receptors in the human basal ganglia: immunohistochemical colocalization of the alpha 1 subunit of the GABAA receptor, and the GABABR1 and GABABR2 receptor subunits

The GABA(B) receptor is a G-protein linked metabotropic receptor that is comprised of two major subunits, GABA(B)R1 and GABA(B)R2. In this study, the cellular distribution of the GABA(B)R1 and GABA(B)R2 subunits was investigated in the normal human basal ganglia using single and double immunohistochemical labeling techniques on fixed human brain tissue. The results showed that the GABA(B) receptor subunits GABA(B)R1 and GABA(B)R2 were both found on the same neurons and followed the same distribution patterns. In the striatum, these subunits were found on the five major types of interneurons based on morphology and neurochemical labeling (types 1, 2, 3, 5, 6) and showed weak labeling on the projection neurons (type 4). In the globus pallidus, intense GABA(B)R1 and GABA(B)R2 subunit labeling was found in large pallidal neurons, and in the substantia nigra, both pars compacta and pars reticulata neurons were labeled for both receptor subunits. Studies investigating the colocalization of the GABA(A) alpha(1) subunit and GABA(B) receptor subunits showed that the GABA(A) receptor alpha(1) subunit and the GABA(B)R1 subunit were found together on GABAergic striatal interneurons (type 1 parvalbumin, type 2 calretinin, and type 3 GAD neurons) and on neurons in the globus pallidus and substantia nigra pars reticulata. GABA(B)R1 and GABA(B)R2 were found on substantia nigra pars compacta neurons but the GABA(A) receptor alpha(1) subunit was absent from these neurons. The results of this study provide the morphological basis for GABAergic transmission within the human basal ganglia and provides evidence that GABA acts through both GABA(A) and GABA(B) receptors. That is, GABA acts through GABA(B) receptors, which are located on most of the cell types of the striatum, globus pallidus, and substantia nigra. GABA also acts through GABA(A) receptors containing the alpha(1) subunit on specific striatal GABAergic interneurons and on output neurons of the globus pallidus and substantia nigra pars reticulata.     

Presynaptic and postsynaptic D1 dopamine receptors in the nigrostriatal system of the rat brain: a quantitative autoradiographic study using the selective D1 antagonist [3H]SCH 23390

Ibotenic acid lesions of the caudate-putamen in rat brain resulted in dramatic reductions in [3H]SCH 23390 binding in both the ipsilateral caudate-putamen and substantia nigra reticulata as assessed by quantitative autoradiography. Nigral ibotenic acid and 6-hydroxydopamine lesions did not significantly alter the binding in either structure. This indicates that D1 receptors in the caudate-putamen are postsynaptic on striatal neurons, while those in the substantia nigra reticulata are presynaptic on nerve terminals originating in the caudate-putamen.     

Changes in [H]zolpidem and [H]Ro 15-1788 binding in rat globus pallidus and substantia nigra pars reticulata following a nigrostriatal tract lesion

Changes in GABA_A receptor α₁ subunit gene expression occur in the globus pallidus and substantia nigra pars reticulata following lesions of the nigrostriatal tract. To determine whether these changes are translated at the protein level, we performed quantitative autoradiography with the α₁ selective ligand, [³H]zolpidem, and the non-selective benzodiazepine site ligand, [³H]Ro 15-1788. Binding of both [³H]zolpidem and [³H]Ro 15-1788 was significantly increased in the substantia nigra pars reticulata (13.5±4.1 and 26.3±2.9%, respectively) and significantly reduced in the globus pallidus (20.9±0.8 and 18.3±1.3%, respectively). These changes in α₁ subunit protein expression may help to compensate for the pathological changes in GABAergic activity that occur after striatal dopamine depletion.     

Cannabinoid modulation of the dopaminergic circuitry: implications for limbic and striatal output

Cannabinoid modulation of dopaminergic transmission is suggested by the ability of delta9-tetrahydrocanabinoid to affect motor and motivated behaviors in a manner similar to that produced by pharmacological manipulation of the nigrostriatal and mesocorticolimbic dopamine systems. These behavioral effects as well as analogous effects of endocannabinoids are largely mediated through the cannabinoid type 1 receptor (CB1R). This receptor is located within the substantia nigra and ventral tegmental area, which respectively house the somata of nigrostriatal and mesocorticolimbic dopaminergic neurons. The CB1R is also abundantly expressed in brain regions targeted by the efferent terminals of these dopaminergic neurons. In this review we present the accumulating anatomical and electrophysiological evidence indicating that in each of these systems cannabinoids modulate dopamine transmission largely if not exclusively through indirect mechanisms. The summarized mechanisms include presynaptic release of amino acid transmitters onto midbrain dopamine neurons and onto both cortical and striatal neurons that express dopamine D1-like or D2-like receptors functionally affiliated with the CB1 receptor. The review concludes with a consideration of the psychiatric and neurological implications of cannabinoid modulation of dopamine transmission within these networks.     

Striatonigral projection neurons contain D1 dopamine receptor-activated c-fos

Dopamine receptor agonists which stimulate the D1 receptor have been shown to activate c-fos in the striatum ipsilateral to a 6-hydroxydopamine (6-OHDA) lesion of the nigrostriatal pathway. In the present study, striatal neurons ipsilateral to a 6-OHDA lesion of the medial forebrain bundle were retrogradely labelled by injection of the fluorescent tracer Fluoro-Gold into the substantia nigra pars reticulata. Five days later, c-fos was induced in the 6-OHDA-denervated striatum by injection of the selective D1 agonist SKF 38393. C-fos-positive nuclei were frequently found in medium-sized striatal cell bodies labelled with Fluoro-Gold. These results indicate that D1 agonists activate c-fos in medium-sized neurons that project to the substantia nigra pars reticulata.     

Stability of substantia nigra pars reticulata neuronal discharge rates during dopamine receptor blockade and its possible mechanisms

It is hypothesized that substantia nigra pars reticulata neurons become overactive during a deficit of dopamine transmission. In this study, we examined how acute dopamine receptor blockade (SCH23390 and eticlopride) affects impulse activity of substantia nigra pars reticulata neurons and their response to iontophoretic gamma-amino-n-butyric acid in awake, unrestrained rats. No changes in discharge rate were found during complete dopamine receptor blockade, but these neurons showed a diminished response to gamma-amino-n-butyric acid, suggesting gamma-amino-n-butyric acid receptor hyposensitivity. This may result from tonic increase in gamma-amino-n-butyric acid input from the striatum and globus pallidus, which are activated during dopamine receptor blockade. As substantia nigra pars reticulata neurons are autoactive and resistant to tonic increases in gamma-amino-n-butyric acid input, changes in their responsiveness to phasic gamma-amino-n-butyric acid inputs, not tonic increase discharge rate, may underlie movement disturbance following dopamine deficit.     

利用神经通路示踪方法证实大鼠纹状体神经元的连接

Using neural pathway tracing and immunohistochemical technique, the striato-direct pathway (BDA3 kDa injected into the rat lateral globus pallidus) and striato-indirect pathway (BDA3 kDa injected into the substantia nigra pars reticulata) neurons were specifically labeled, and then subjected to double-labeled immunohistochemistry for mu-OPIOID Receptor (specifically-labeled striatal patch compartment), D1, and D2, respectively. The experimental findings showed that there are no statistically significant differences in the soma diameter and the number of primary dendrites between the striato-direct (substantia nigra pars reticularis) and indirect (globus pallidum externum) neurons labeled retrograde by BDA3 kDa. In addition, these two kinds of projection neurons revealed no obvious coexistence. This evidence indicates that as a highly sensitive neural pathway tracer, BDA could yield reliably and exquisitely detailed labeling of target neurons and synaptic structures. The variance of the morphologic structures and the localization of neurons were not statistically significant between the striato-substantia nigra pars reticularis and the globus pallidum externum projection neurons. Mesencephalic and thalamic neurons correlated with striatal neurons in morphology. Especially the latter which make typical excitatory synaptic contacts with striato-direct and -indirect neurons. Thus, this evidence suggests that thalamic neurons may extensively excite striatal neurons.     

Expression of D4 dopamine receptors in striatonigral and striatopallidal neurons in the rat striatum

Recent studies have reported the regional distribution of D(4) dopamine receptors in the rat striatum at the cellular and subcellular levels. However, the precise identity of the striatal neurons that express these receptors remains unknown. We have studied the expression of D(4) receptors in the striatal interneurons as well as in the output regions of the striatum using immunohistochemistry. Furthermore, we have evaluated the contribution of the striatum to D(4) receptor immunoreactivity in these areas by means of ibotenic acid lesion of the striatum. D(4) receptors were observed in the substantia nigra pars reticulata (SNr), the entopeduncular nucleus (EP) and the globus pallidus (GP), and they were found, using electron microscopy, to be located presynaptically. D(4) immunoreactivity in the striatal output nuclei was observed to dramatically decrease following lesion of the striatum with ibotenic acid. Striatal interneurons were not found to express D(4) receptors. These results demonstrate that D(4) receptors are located almost exclusively in striatal projection neurons, in both striatonigral and striatopallidal neurons.     

Fatty acid incorporation depicts brain activity in a rat model of Parkinson's disease

Following pulse labeling with [ ]arachidonic acid ([ ]AA), its incorporation pattern in brain reflects regional changes in neurotransmitter signal transduction using phospholipase A₂, that is, functional activity. In a rat model of Parkinson's disease, unilateral 6-hydroxydopamine lesion in the substantia nigra, [ ]AA acid incorporation from blood was increased in cerebral cortex, caudate putamen, globus pallidus, entopeduncular nucleus, subthalamic nucleus and substantia nigra pars reticulata ipsilateral to the lesion. This increased [ ]AA incorporation likely reflects disinhibition of basal ganglia and cortical circuits secondary to absent inhibitory nigrostriatal dopaminergic input.     

Excitatory amino acid receptor regulation after subthalamic nucleus lesions in the rat

The subthalamic nucleus plays a pivotal role in the regulation of basal ganglia output. Recent electrophysiologic, lesion and immunocytochemical studies suggest that the subthalamic nucleus uses an excitatory amino acid as a neurotransmitter. After complete ablation of the subthalamic nucleus, we have examined the NMDA, AMPA, kainate and metabotropic subtypes of excitatory amino acid receptors in two major subthalamic projection areas (globus pallidus and substantia nigra pars reticulata) with quantitative autoradiography. Two weeks after ablation, binding sites for [³H]AMPA and [³H]kainate increased in substantia nigra pars reticulata ipsilateral to the lesion. In globus pallidus on the lesioned side, [³H]glutamate binding to the NMDA recognition site decreased. The results suggest that glutamate receptors regulate after interruption of subthalamic nucleus output.