Striatopallidal Projection Neurons Research Articles

Cell-type- and region-specific modulation of cocaine seeking by micro-RNA-1 in striatal projection neurons

The persistent and experience-dependent nature of drug addiction may result in part from epigenetic alterations, including non-coding micro-RNAs (miRNAs), which are both critical for neuronal function and modulated by cocaine in the striatum. Two major striatal cell populations, the striato-nigral and striato-pallidal projection neurons, express, respectively, the D1 (D1-SPNs) and D2 (D2-SPNs) dopamine receptor, and display distinct but complementary functions in drug-evoked responses. However, a cell-type-specific role for miRNAs action has yet to be clarified. Here, we evaluated the expression of a subset of miRNAs proposed to modulate cocaine effects in the nucleus accumbens (NAc) and dorsal striatum (DS) upon sustained cocaine exposure in mice and showed that these selected miRNAs were preferentially upregulated in the NAc. We focused on miR-1 considering the important role of some of its predicted mRNA targets, Fosb and Npas4, in the effects of cocaine. We validated these targets in vitro and in vivo. We explored the potential of miR-1 to regulate cocaine-induced behavior by overexpressing it in specific striatal cell populations. In DS D1-SPNs miR-1 overexpression downregulated Fosb and Npas4 and reduced cocaine-induced CPP reinstatement, but increased cue-induced cocaine seeking. In DS D2-SPNs miR-1 overexpression reduced the motivation to self-administer cocaine. Our results indicate a role of miR1 and its target genes, Fosb and Npas4, in these behaviors and highlight a precise cell-type- and region-specific modulatory role of miR-1, illustrating the importance of cell-specific investigations.

Age- and sex-related changes in cortical and striatal nitric oxide synthase in the Q175 mouse model of Huntington's disease

In Huntington's disease (HD), corticostriatal and striatopallidal projection neurons preferentially degenerate as a result of mutant huntingtin expression. Pathological deficits in nitric oxide (NO) signaling have also been reported in corticostriatal circuits in HD, however, the impact of age and sex on nitrergic transmission is not well characterized. Thus, we utilized NADPH-diaphorase (NADPH-d) histochemistry and qPCR assays to assess neuronal NO synthase (nNOS) activity/expression in aged male and female Q175 heterozygous mice. Compared to age-matched controls, male Q175 mice exhibited reductions in NADPH-d staining in the motor cortex at 21, but not, 16 months of age. Comparisons across genotypes showed that striatal NADPH-d staining was significantly decreased at both 16 and 21 months of age. Comparisons within sexes in 21 month old mice revealed a decrease in striatal NADPH-d staining in males, but no changes were detected in females. Significant correlations between cortical and striatal NADPH-d staining deficits were also observed in males and females at both ages. To directly assess the role of constitutively active NOS isoforms in these changes, nNOS and endothelial NOS (eNOS) mRNA expression levels were examined in R6/2 (3 month old) and Q175 (11.5 month old) mice using qPCR assays. nNOS transcript expression was decreased in the cortex (40%) and striatum (54%) in R6/2 mice. nNOS mRNA down-regulation in striatum of Q175 animals was more modest (19%), and no changes were detected in cortex. eNOS expression was not changed in the cortex or striatum of Q175 mice. The current findings point to age-dependent deficits in nNOS activity in the HD cortex and striatum which appear first in the striatum and are more pronounced in males. Together, these observations and previous studies indicate that decreases in nitrergic transmission progress with age and are likely to contribute to corticostriatal circuit pathophysiology particularly in male patients with HD.

Density of metabotropic glutamate receptors subtype 1 in Parkinson’s disease compared to healthy elderly – a ITMM PET study –

Background: Glutamate receptors are divided into 2 types based on their biological functions and molecular structures: ionotropic and metabotropic types. The metabotropic type is classified into 3 groups. Metabotropic glutamate receptor subtype 1 (mGluR1) is one of the group 1 receptors, localized postsynaptically in the cerebellar Purkinje cells, striatonigral and striatopallidal projection neurons, and striatal interneurons. However, the relationship between the mGluR1 and Parkinson’s disease remains to be determined.

Retinoic Acid Receptor β Controls Development of Striatonigral Projection Neurons through FGF-Dependent and Meis1-Dependent Mechanisms.

The mammalian striatum controls sensorimotor and psychoaffective functions through coordinated activities of its two striatonigral and striatopallidal output pathways. Here we show that retinoic acid receptor β (RARβ) controls development of a subpopulation of GABAergic, Gad65-positive striatonigral projection neurons. In Rarb(-/-) knock-out mice, concomitant reduction of Gad65, dopamine receptor D1 (Drd1), and substance P expression at different phases of prenatal development was associated with reduced number of Drd1-positive cells at birth, in contrast to normal numbers of striatopallidal projection neurons expressing dopamine receptor D2. Fate mapping using BrdU pulse-chase experiments revealed that such deficits may originate from compromised proliferation of late-born striosomal neurons and lead to decreased number of Drd1-positive cells retaining BrdU in postnatal day (P) 0 Rarb(-/-) striatum. Reduced expression of Fgf3 in the subventricular zone of the lateral ganglionic eminence (LGE) at embryonic day 13.5 may underlie such deficits by inducing premature differentiation of neuronal progenitors, as illustrated by reduced expression of the proneural gene Ascl1 (Mash1) and increased expression of Meis1, a marker of postmitotic LGE neurons. In agreement with a critical role of FGF3 in this control, reduced number of Ascl1-expressing neural progenitors, and a concomitant increase of Meis1-expressing cells, were observed in primary cell cultures of Rarb(-/-) LGE. This defect was normalized by addition of fibroblast growth factor (FGF). Such data point to role of Meis1 in striatal development, also supported by reduced neuronal differentiation in the LGE of Meis1(-/-) embryos. Our data unveil a novel mechanism of development of striatonigral projection neurons involving retinoic acid and FGF, two signals required for positioning the boundaries of Meis1-expressing cells.

Plasticity in striatopallidal projection neurons mediates the acquisition of habitual actions

In instrumental conditioning, newly acquired actions are generally goal-directed and are mediated by the relationship between the action and its consequences or outcome. With continued training, however, the performance of such actions can become automatic, reflexive or habitual and under the control of antecedent stimuli rather than their consequences. Recent evidence suggests that habit learning is mediated by plasticity in the dorsolateral striatum (DLS). To date, however, no direct evidence of learning-related plasticity associated with overtraining has been reported in this region, nor is it known whether, or which, specific cell types are involved in this learning process. The striatum is primarily composed of two classes of spiny projection neurons, the striatonigral and striatopallidal spiny projection neurons, which express dopamine D1 and D2 receptors, and control direct and indirect pathways, respectively. Here we found evidence of a post-synaptic depression in DLS striatopallidal projecting neurons in the indirect pathway during habit learning in mice. Moreover, this training-induced depression occluded post-synaptic depression induced by co-activation of D2 receptors and transient receptor potential vanilloid 1 (TRPV1) channels, implying that this pathway is involved in habit learning. This hypothesis was further tested by disrupting this signal pathway by knocking out TRPV1 channels, resulting in compromised habit learning. Our findings suggest that post-synaptic plasticity at D2 neurons in the DLS mediates habit learning and, by implicating an interaction between the D2 receptor and TRPV1 channel activity, provide a potential drug target for influencing habitual action control.

The diversity of GABA(A) receptor subunit distribution in the normal and Huntington's disease human brain.

GABA(A) receptors are assembled into pentameric receptor complexes from a total of 19 different subunits derived from a variety of different subunit classes (α1-6, β1-3, γ1-3, δ, ɛ, θ, and π) which surround a central chloride ion channel. GABA(A) receptor complexes are distributed heterogeneously throughout the brain and spinal cord and are activated by the extensive GABAergic inhibitory system. In this chapter, we describe the heterogeneous distribution of six of the most widely distributed subunits (α1, α2, α3, β2,3, and γ2) throughout the human basal ganglia. This review describes the studies we have carried out on the normal and Huntington's disease human basal ganglia using autoradiographic labeling and immunohistochemistry in the human basal ganglia. GABA(A) receptors are known to react to changing conditions in the brain in neurological disorders, especially in Huntington's disease and display a high degree of plasticity which is thought to compensate for loss of function caused by disease. In Huntington's disease, the variable loss of GABAergic medium spiny striatopallidal projection neurons is associated with a loss of GABA(A) receptor subunits in the striosome and/or the matrix compartments of the striatum. By contrast in the globus pallidus, a loss of the GABAergic striatal projection neurons results in a dramatic upregulation of subunits on the large postsynaptic pallidal neurons; this is thought to be a compensatory plastic mechanism resulting from the loss of striatal GABAergic input. Most interestingly, our studies have revealed that the subventricular zone overlying the caudate nucleus contains a variety of proliferating progenitor stem cells that possess a heterogeneity of GABA(A) receptor subunits which may play a role in human brain repair mechanisms.

Cell type-selective expression of the zinc finger-containing gene Nolz-1/Zfp503 in the developing mouse striatum

The zinc finger-containing gene Nolz-1/Zfp503 is a developmentally regulated striatum-enriched gene. In the present study, we characterized the cell type-selective expression pattern of Nolz-1 protein in the developing mouse striatum. Nolz-1 immunoreactivity was present in Isl-1-positive ventral LGE (vLGE, striatal primordia), but absent in Pax6-positive dorsal LGE (dLGE, non-striatal primordia). In the vLGE, Nolz-1 immunoreactivity was detected in early differentiating TuJ1-positive neurons, but not in Ki67-positive proliferating progenitor cells. Moreover, many Nolz-1-immunoreactive cells co-expressed Foxp1 or Foxp2, markers for striatal projection neurons. To further characterize Nolz-1 expression with respect to D1R-containing striatonigral and D2R-containing striatopallidal projection neurons, we used the Drd1-EGFP and Drd2-EGFP transgenic mice. Nolz-1 and EGFP double labeled neurons were found in the developing striatum of Drd1-EGFP and Drd2-EGFP mice, indicating Nolz-1 expression in both populations of striatal projection neurons. Notably, Nolz-1 protein was not expressed in Nkx2.1-positive interneuron progenitors, Lhx8-positive cholinergic interneuron progenitors, nNOS and calretinin-positive interneurons in E18.5 striatum. In the developing nucleus accumbens and olfactory tubercles of ventral striatum, many Nolz-1-positive cells co-expressed Sox1, an important transcriptional regulator for ventral striatum, suggesting a role of Nolz-1 in regulating development of the ventral striatum. Finally, in contrast to postnatal down-regulation of Nolz-1 in the dorsal striatum, Nolz-1 protein was persistently expressed in the olfactory tubercle from E15.5 to adulthood. Taken together, our study suggests that Nolz-1 serves as a marker for early differentiating striatal projection neurons and that Nolz-1 may regulate development of striatal projection neurons.

Shingosine-1-phosphate regulates PKA/DARPP-32 signaling in striatal medium spiny neurons via GPR6-independent mechanisms

The vast majority of striatal neurons are GABAergic medium-sized spiny neurons. These cells receive glutamatergic input from the cortex, thalamus and limbic areas and dopaminergic input from the mesencephalon. Most relevant evidence indicates that dopamine D1 receptors are located on striatonigral projection neurons,5, 7 and that adenosine A2A receptors4, 12, 14 and most dopamine D2 receptors5, 7, 14 are located on striatopallidal projection neurons (see, however, 1, 13). Here we have utilized regulation of the phosphorylation of dopamine- and cyclic AMP-regulated phosphoprotein of mol. wt 32,000 (DARPP-32) to study the possible interactions among nigrostriatal dopaminergic neurons and the two classes of dopaminoceptive target neurons. We show that, in striatal slices, the D2 receptor agonist, quinpirole, strongly inhibits the phosphorylation of DARPP-32 induced by either the D1 receptor agonist, SKF 81297, or the A2A receptor agonist, CGS 21680. Tetrodotoxin abolished the effect of quinpirole on the D1 agonist-induced but not the A2A agonist-induced phosphorylation of DARPP-32. These data indicate that: (i) adenosine A2A and dopamine D2 receptors interact within the same striatopallidal neurons, and (ii) D2 receptors present on the striatopallidal neurons modulate the effects of D1 receptors on the striatonigral neurons. Thus, a single neurotransmitter is capable of activating distinct classes of receptors on distinct populations of target neurons, which, in turn, interact with each other through intercellular communication.

A conductance-based network model of the basal ganglia for probabilistic action selection

The basal ganglia (BG) have been implicated in the learning of a sequence of action selections through trial-and-error. A reinforcement learning-based approach has proposed that the cortico-BG circuit is involved in three basic stages of learning: evaluation of actions, probabilistic selection of an action, and learning from experience. The striatum and mid-brain dopaminergic neurons have been suggested as neural substrates for the first and third stages. Theoretical studies have pointed out the importance of a probabilistic action selection mechanism for learning and on-line adaptation of the behavior. However, the neural substrate of the action selection is still an open question. Our hypothesis for the issue is that the indirect pathway of the BG selects an action to be executed and the direct pathway determines the timing of its execution. Using a conductance-based network model of spiking neurons, we show that the dynamics in the network of the globus pallidus external and the subthalamic nucleus in the indirect pathway provides binary modulation on the substantia nigra par reticulata, that signals a selected action. Furthermore, binary modulation occurs stochastically, and the selection probability is sensitive to inhibitory input on the globus pallidus. These results suggests that the subthalamopallidal network is capable of probabilistic action selection and the selection probability can be biased by the activities of the striatopallidal projection neurons in the indirect pathway, hence can be optimally tuned by the strength of the cortico-striatal synapses through dopamine-dependent plasticity. We conclude that the indirect pathway of the BG is a neural substrate of the probabilistic action selection.

Cytochrome oxidase activity in the monkey globus pallidus and subthalamic nucleus after ablation of striatal interneurons expressing substance P receptors

To understand functional roles of striatal interneurons in primate basal ganglia circuitry, we ablated interneurons expressing substance P (SP) receptors (SPR) in the putamen with SP-saporin, a SPR selective neurotoxin. The effect of SP-saporin injection into the putamen was evaluated by examining the loss of cholinergic interneurons and NADPHd-positive (nicotinamide adenine dinucleotide phosphate diaphorase positive) interneurons. We then analyzed regional metabolic changes using cytochrome oxidase (CO) histochemistry. CO activity in some regions of the internal and external segments of the globus pallidus (GP) in the lesioned hemisphere was lower than that in the contralateral or surrounding GP regions. CO activity in the subthalamic nucleus, however, showed no significant change. The present findings suggest that striatopallidal projection neurons exert enhanced inhibitory influence on the GP without modulatory control by the striatal SPR-expressing interneurons.

Chemical organization of projection neurons in the rat accumbens nucleus and olfactory tubercle

Projection neurons in the ventral striatum, the accumbens nucleus and olfactory tubercle, were examined by combining the retrograde tracing method and immunocytochemistry with antibodies against C-terminals of the preprodynorphin (PPD), preproenkephalin (PPE), preprotachykinin A (PPTA) and preprotachykinin B (PPTB). When the retrograde tracer was injected into the ventral pallidum, about 60% and 40% of retrogradely labeled neurons in the accumbens nucleus were immunoreactive for PPD and PPE, respectively. In contrast, all accumbens nucleus neurons projecting to the ventral mesencephalic regions including the substantia nigra and ventral tegmental area were immunopositive for PPD but not for PPE. Although no olfactory tubercle neurons projected fibers to the mesencephalic regions, 60% and 40% of olfactory tubercle neurons projecting to the ventrolateral portion of the ventral pallidum were immunoreactive for PPD and PPE, respectively, as were the accumbens nucleus neurons. About 70% of accumbens nucleus and olfactory tubercle neurons projecting to the ventral pallidum and all accumbens nucleus neurons projecting to the ventral mesencephalic regions showed PPTA immunoreactivity. A small population (2–12%) of accumbens neurons projecting to the ventral pallidum and mesencephalic regions displayed immunoreactivity for PPTB. Compared with the dorsal striatopallidal projection neurons that were reported to mostly express PPE, it was characteristic of the ventral striatum that only the smaller population (about 40%) of ventral striatopallidal projection neurons expressed PPE. This suggests that the ventral striatopallidal projection system is less specialized than the dorsal striatopallidal system in terms of peptide production, or that the ventral pallidum should be compared with a combined region of the globus pallidus and entopeduncular nucleus in the dorsal system.

Analysis of mRNA expression using double in situ hybridization labeling with isotopic and nonisotopic probes.

AbstractA large number of in vivo studies in the last decade have confirmed that abused drugs are able to up-regulate gene expression in striatal neurons. For example, acute or chronic exposures to psychostimulants, cocaine and amphetamine, increased basal levels of mRNA and protein products of immediate early genes (c-fos, zif/268, ΔFosB, etc.) in the rodent striatum (1–3). Similarly, acute administration of cocaine and amphetamines induced robust mRNA expression of preprodynorphin (PPD) and substance P (SP) in striatonigral projection neurons and preproenkaphalin (PPE) in striatopallidal projection neurons (4–6). The increased immediate early gene and neuropeptide expression is mediated via selective activation of dopamine receptors (primary D1 subtype) (7,8). Glutamatergic transmission also significantly contributes to the genomic effect of stimulants (9,10). The altered gene expression has been considered to be an essential molecular step for the development of psychoplasticity in the striatum related to addictive properties of drugs of abuse (11,12).KeywordsHybridization SignalStriatal NeuronIsotopic ProbeNitroblue Tetrazolium ChlorideAddictive PropertyThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Differential Changes in Striatal Projection Neurons in R6/2 Transgenic Mice for Huntington's Disease

In early adult-onset Huntington's disease (HD), enkephalinergic striatopallidal projection neurons show preferential loss, reduced preproenkephalin (PPE) expression in surviving striatopallidal neurons, and loss of fibers in their projection target area. We examined PPE and PPT (preprotachykinin) gene expression in striatal projection neurons and in striatal projection fibers immunoreactive for the PPE product enkephalin (ENK) and the PPT product substance P (SP) in a transgenic HD model, the R6/2 mouse, to see if changes occur in these neuron types similar to those seen in early adult-onset HD. The results show that PPE mRNA level, the number of striatal neurons expressing PPE, and the staining intensity of fibers immunoreactive for ENK in the pallidum were all decreased. By contrast, the SP-containing striatal projection systems to the pallidum and substantia nigra were relatively normal in R6/2 mice. The selective reduction in striatal PPE in R6/2 mice is reminiscent of adult-onset HD, but the preservation of the striatonigral projection system is not. Thus, R6/2 mice do not strictly mimic adult-onset HD in their striatal pathology.

Sustained unilateral blockade of rat striatal M 1 muscarinic receptors with m1-toxin1 in vivo

m1-Toxin1 is a trace component of the venom of the green mamba that antagonizes M 1 muscarinic receptors specifically and irreversibly in vitro. It was injected into the right caudatoputamen of Wistar rats to occlude M 1 receptors specifically for several days (‘M 1 knockdown’), and to begin to establish the unknown effects of striatal M 1-neurotransmission on movement. The extent and duration of M 1-blockade were evaluated with receptor binding assays and light microscopic autoradiography, using 1–2 nM [ 3H]pirenzepine to assess unoccupied M 1 receptors. An injection of 27 pmol m1-toxin1 blocked almost all of the M 1 receptors in one caudatoputamen (4 pmol), followed by their slow recovery to supranormal levels (115%) during a week. During maximum M 1-blockade, the binding of 1 nM [ 3H] N-methylscopolamine to striatal membranes was reduced by only half, indicating no blockade of M 4 receptors, which comprise 51% of striatal muscarinic receptors. It is concluded that m1-toxin1 can produce acute, focal, selective, unilateral and long-lasting M 1-blockade for investigations of the effects of M 1-neurotransmission in vivo. Rats with extensive unilateral striatal M 1-blockade appeared normal, and did not show unilateral deficits in spontaneous forearm use, in contrast to rats with unilateral 6-hydroxydopamine lesions of the substantia nigra. Since M 1-blockade should decrease the activity of both striatonigral and striatopallidal projection neurons, and the relative activity of these neurons is believed to control movement, the results suggest that a balanced decrease in the activity of these neurons on one side of the brain does not produce a right/left imbalance in spontaneous movement.

A sequential protocol combining dual neuroanatomical tract-tracing with the visualization of local circuit neurons within the striatum

We describe here an experimental approach designed to aid in the identification of complex brain circuits within the rat corpus striatum. Our aim was to characterize in a single section (i) striatal thalamic afferents, (ii) striatopallidal projection neurons and (iii) striatal local circuit interneurons. To this end, we have combined anterograde tracing using biotinylated dextran amine and retrograde neuroanatomical tracing with Fluoro-Gold. This dual tracing protocol was further implemented with the visualization of different subpopulations of striatal interneurons. The subsequent use of three different peroxidase substrates enabled us to unequivocally detect structures that were labeled within a three-color paradigm.

Biotinylated m4-toxin demonstrates more M4 muscarinic receptor protein on direct than indirect striatal projection neurons

The striatum has nearly equal numbers of striatonigral and striatopallidal projection neurons. All are GABAergic and inhibitory, but they lie in separate neuronal circuits (‘direct’ and ‘indirect’, respectively) that appear to exert opposite effects on movement. Methods are needed to evaluate the function of each circuit. A potential way to control striatonigral neurons selectively is via M4 muscarinic receptors. The striatum has many more M4 receptors than other tissues, they are located on approximately half of all projection neurons, and mRNA for M4 receptors is prevalent only in striatonigral neurons. In order to more rigorously compare the distribution of M4 receptors on rat neurons in these pathways a toxin that binds with very high specificity to M4 receptors (m4-toxin) was biotinylated for use as a selective probe for M4 receptor protein. Pooled biotin–toxin complexes were found to retain high M4-specificity and affinity. Neurons were first labeled by retrograde transport of fluorescent microbeads (FluoSpheres) injected into the substantia nigra and globus pallidus. Coincident labeling of only 4% of the cells confirmed the validity of the retrograde labeling technique. Labeled neurons were probed for M4 receptor protein using biotinylated m4-toxin and fluorescent avidin. M4 receptors were found on 14% of indirect and 86% of direct neurons. It may be concluded that there is a relative abundance of M4 receptors controlling the direct pathway. This work supports the hypothesis that M4-selective drugs will prove useful to control the function of striatonigral neurons in the direct projection pathway.

Increased Response to Intrastriatal L(+)-2-Amino-4-Phosphonobutyrate (L-AP4) in Unilateral 6-Hydroxydopamine-Lesioned Rats

Unilateral intrastriatal injection of the prototype Group III metabotropic glutamate receptor agonist L(+)-2-amino-4-phosphonobutyrate (L-AP4) induces rotation in rats with ipsilateral unilateral 6-hydroxydopamine lesions of the nigrostriatal projection but not in unlesioned control animals [Kearney et al., (1998) Neuroscience 87:881–891]. We studied differences in striatal neuron Fos-like immunoreactivity expression, striatal neuron Fos-like immunoreactivity localization, and regional [14C]-2-deoxyglucose uptake after unilateral intrastriatal injection of L-AP4 between control and unilateral 6-hydroxydopamine lesioned rats. There was a left shift of the dose response curve with more striatal neurons expressing Fos-like immunoreactivity at lower doses of L-AP4 in 6-hydroxydopamine lesioned animals. In unlesioned striata, L-AP4 injections tended to induce Fos-like immunoreactivity in striato-pallidal projection neurons. In lesioned animals, the majority of striatal neurons expressing Fos-like immunoreactivity were striatonigral projection neurons. In both control and lesioned animals, intrastriatal injection of L-AP4 produced widespread decreases in [14C]-2-deoxyglucose uptake in basal ganglia nuclei and related regions, but the magnitude of this effect increased markedly in lesioned animals. Striatal dopamine denervation enhances the response of striatal neurons to intrastriatally injected L-AP4 with an apparent shift in the type of striatal projection neuron responding to L-AP4.

Localization of cannabinoid CB(1) receptor mRNA in neuronal subpopulations of rat striatum: a double-label in situ hybridization study.

Double-label in situ hybridization was used to identify the phenotypes of striatal neurons that express mRNA for cannabinoid CB(1) receptors. Simultaneous detection of multiple mRNAs was performed by combining a (35)S-labeled ribonucleotide probe for CB(1) mRNA with digoxigenin-labeled riboprobes for striatal projection neurons (preprotachykinin A, prodynorphin, and preproenkephalin mRNAs) and interneurons (vesicular acetylcholine transporter (VAChT), choline acetyltransferase (ChAT), somatostatin, and glutamic acid decarboxylase (Mr 67,000; GAD67) mRNAs). To ascertain whether CB(1) mRNA was a marker for striatal efferents, digoxigenin-labeled probes for mRNA markers of both striatonigral (prodynorphin or preprotachykinin A mRNAs), and striatopallidal (proenkephalin mRNAs) projection neurons were combined with the (35)S-labeled probe for CB(1). A mediolateral gradient in CB(1) mRNA expression was observed at rostral and mid-striatal levels; in the same coronal sections the number of silver grains per cell ranged from below the threshold of detectability at the medial and ventral poles to saturation at the dorsolateral boundary bordered by the corpus callosum. At the caudal level examined, CB(1) mRNA was denser in the ventral sector relative to the dorsal sector. Virtually all neurons expressing mRNA markers for striatal projection neurons colocalized CB(1) mRNA. Combining a (35)S-labeled riboprobe for CB(1) with digoxigenin-labeled riboprobes for both preproenkephalin and prodynorphin confirmed localization of CB(1) mRNA to striatonigral and striatopallidal neurons expressing prodynorphin and preproenkephalin mRNAs, respectively. However, CB(1) mRNA-positive cells that failed to coexpress the other markers were also apparent. CB(1) mRNA was localized to putative GABAergic interneurons that express high levels of GAD67 mRNA. These interneurons enable functional interactions between the direct and indirect striatal output pathways. By contrast, aspiny interneurons that express preprosomatostatin mRNA and cholinergic interneurons that coexpress ChAT and VAChT mRNAs were CB(1) mRNA-negative. The present data provide direct evidence that cannabinoid receptors are synthesized in striatonigral neurons that contain dynorphin and substance P and striatopallidal neurons that contain enkephalin. By contrast, local circuit neurons in striatum that contain somatostatin or acetylcholine do not synthesize cannabinoid receptors. Published 2000 Wiley-Liss, Inc.

Activation of dopamine D 2 receptors decreases DARPP-32 phosphorylation in striatonigral and striatopallidal projection neurons via different mechanisms

The vast majority of striatal neurons are GABAergic medium-sized spiny neurons. These cells receive glutamatergic input from the cortex, thalamus and limbic areas and dopaminergic input from the mesencephalon. Most relevant evidence indicates that dopamine D 1 receptors are located on striatonigral projection neurons, [5, 7] and that adenosine A 2A receptors [4, 12, 14] and most dopamine D 2 receptors [5, 7, 14] are located on striatopallidal projection neurons (see, however, [1, 13]). Here we have utilized regulation of the phosphorylation of dopamine- and cyclic AMP-regulated phosphoprotein of mol. wt 32,000 (DARPP-32) to study the possible interactions among nigrostriatal dopaminergic neurons and the two classes of dopaminoceptive target neurons. We show that, in striatal slices, the D 2 receptor agonist, quinpirole, strongly inhibits the phosphorylation of DARPP-32 induced by either the D 1 receptor agonist, SKF 81297, or the A 2A receptor agonist, CGS 21680. Tetrodotoxin abolished the effect of quinpirole on the D 1 agonist-induced but not the A 2A agonist-induced phosphorylation of DARPP-32. These data indicate that: (i) adenosine A 2A and dopamine D 2 receptors interact within the same striatopallidal neurons, and (ii) D 2 receptors present on the striatopallidal neurons modulate the effects of D 1 receptors on the striatonigral neurons. Thus, a single neurotransmitter is capable of activating distinct classes of receptors on distinct populations of target neurons, which, in turn, interact with each other through intercellular communication.

Targeted Expression of a Toxin Gene to D1 Dopamine Receptor Neurons byCre-Mediated Site-Specific Recombination

Idiopathic Parkinson's disease involves the loss of midbrain dopaminergic neurons, resulting in the presynaptic breakdown of dopaminergic transmission in the striatum. Huntington's disease and some neurodegenerative diseases with Parkinsonian features have postsynaptic defects caused by striatal cell death. Mice were generated in which an attenuated form of the diphtheria toxin gene (tox-176) was expressed exclusively in D1 dopamine receptor (D1R)-positive cells with the aim of determining the effect of this mutation on development of the basal ganglia and on the locomotor phenotype. Transgenic mice expressing Cre, a site-specific DNA recombinase, were crossed with a second line in which a transcriptionally silenced tox-176 gene was inserted into the D1R gene locus by homologous recombination. Young doubly transgenic mutant mice expressing the tox-176 gene displayed bradykinesia, dystonia, and had falls caused by myoclonic jerks. The mutant brain had evidence of apoptosis and reactive gliosis and, consistent with the D1R expression pattern, the striatum was reduced in volume, and the Islands of Calleja were absent. In contrast, the cortex was of normal thickness. D1Rs were not detectable in mutants by in situ hybridization or ligand autoradiography, whereas D2 dopamine receptor (D2R) mRNA and protein was present in the striatum. In addition, substance P and dynorphin, neuropeptides known to be expressed in D1R-positive striatonigral projection neurons were not detectable. Enkephalin, a marker found in D2-positive striatopallidal projection neurons was expressed in the mutant brain. The mutant represents a novel neurodegenerative disease model with a dramatic extrapyramidal phenotype.