NORMAL GENOMIC

• Bonsi P et al.
• Functional and ultrastructural analysis of group I mGluR in striatalfast-spiking interneurons.
• Eur J Neurosci. 2007; 25: 1319-31
• Display abstract

• Striatal parvalbumin-containing fast-spiking (FS) interneurons provide apowerful feedforward GABAergic inhibition on spiny projection neurons,through a widespread arborization and electrical coupling. Modulation ofFS interneuron activity might therefore strongly affect striatal output.Metabotropic glutamate receptors (mGluRs) exert a modulatory action atvarious levels in the striatum. We performed electrophysiologicalrecordings from a rat striatal slice preparation to investigate theeffects of group I mGluR activation on both the intrinsic and synapticproperties of FS interneurons. Bath-application of the group I mGluRagonist, (S)-3,5-dihydroxyphenylglycine (3,5-DHPG), caused adose-dependent depolarizing response. Both(S)-(+)-alpha-amino-4-carboxy-2-methylbenzeneacetic acid (LY367385) and7-(hydroxyimino)cyclopropa[b]chromen-1a-carboxylate ethyl ester (CPCCOEt),selective mGluR1 antagonists, significantly reduced the amplitude of themembrane depolarization caused by 3,5-DHPG application. Conversely, mGluR5antagonists, 2-methyl-6-(phenylethylnyl)pyridine hydrochloride (MPEP) and6-methyl-2-(phenylazo)-3-pyridinol (SIB1757), were unable to affect theresponse to 3,5-DHPG, suggesting that only mGluR1 contributes to the3,5-DHPG-mediated excitatory action on FS interneurons. Furthermore,mGluR1 blockade significantly decreased the amplitude of the glutamatergicpostsynaptic potentials, whereas the mGluR5 antagonist applicationproduced a small nonsignificant inhibitory effect. Surprisingly, ourelectron microscopic data demonstrate that the immunoreactivity for bothmGluR1a and mGluR5 is expressed extrasynaptically on the plasma membraneof parvalbumin-immunoreactive dendrites of FS interneurons. Together,these results suggest that despite a common pattern of distribution,mGluR1 and mGluR5 exert distinct functions in the modulation of FSinterneuron activity.

• Mura A, Linder JC, Young SJ, Groves PM
• Striatal cells containing aromatic L-amino acid decarboxylase: animmunohistochemical comparison with other classes of striatal neurons.
• Neuroscience. 2000; 98: 501-11
• Display abstract

• In a previous study, we described a population of striatal cells in therat brain containing aromatic L-amino acid decarboxylase, the enzymeinvolved in the conversion of L-DOPA into dopamine. We have also presentedevidence that these cells produce dopamine in the presence of exogenousL-DOPA. In this paper, we further characterize these striatal aromaticL-amino acid decarboxylase-containing cells in order to determine whetherthey form a subclass of one of the known categories of striatal neurons orif they represent a novel cell type. Using immunohistochemical methods, wecompared the morphology and distribution of the aromatic L-amino aciddecarboxylase-immunolabeled cells with those of other classes of striatalneurons. Our results show that both the morphology and distribution ofaromatic L-amino acid decarboxylase-immunolabeled cells are verydistinctive and do not resemble those of cells labeled for other striatalneuronal markers. Double-labeling procedures revealed that aromaticL-amino acid decarboxylase cells do not co-localize somatostatin orparvalbumin, and only a very small percentage of them co-localizecalretinin. However, the population of aromatic L-amino acid decarboxylasecells label intensely for GABA.Overall, our results suggest that thesearomatic L-amino acid decarboxylase-containing cells represent a class ofstriatal GABAergic neurons not described previously.

• Dunah AW, Wyszynski M, Martin DM, Sheng M, Standaert DG
• alpha-actinin-2 in rat striatum: localization and interaction with NMDAglutamate receptor subunits.
• Brain Res Mol Brain Res. 2000; 79: 77-87
• Display abstract

• Alpha-actinin (alpha-actinin-2) is a protein which links the NR1 and NR2Bsubunits of N-methyl-D-aspartate (NMDA) glutamate receptors to the actincytoskeleton. Because of the importance of NMDA receptors in modulatingthe function of the striatum, we have examined the localization ofalpha-actinin-2 protein and mRNA in striatal neurons, and its biochemicalinteraction with NMDA receptor subunits present in the rat striatum. Usingan alpha-actinin-2-specific antibody, we found intense immunoreactivity inthe striatal neuropil and within striatal neurons that also expressedparvalbumin, calretinin and calbindin. Conversely, alpha-actinin-2immunoreactivity was not detected in neurons expressing cholineacetyltransferase and neuronal nitric oxide synthase. Dual-label in situhybridization revealed that the highest expression of alpha-actinin-2 mRNAis in substance P-containing striatal projection neurons. Thealpha-actinin-2 mRNA is also present in enkephalinergic projection neuronsand interneurons expressing parvalbumin, choline acetyl transferase andthe 67-kDa isoform of glutamic acid decarboxylase, but was not detected insomatostatin-expressing interneurons. Immunoprecipitation of membraneprotein extracts showed that alpha-actinin-2 is present in heteromericcomplexes of NMDA subunits, but is not associated with AMPA receptors inthe striatum. A subunit-specific anti-NR1 antibody co-precipitated majorfractions of NR2A and NR2B subunits, but only a minor fraction of striatalalpha-actinin-2. Conversely, alpha-actinin-2 antibody immunoprecipitatedonly modest fractions of striatal NR1, NR2A and NR2B subunits. These datademonstrate that alpha-actinin-2 is a very abundant striatal protein, butexhibits cellular specificity in its expression, with very high levels insubstance-P-containing projection neurons, and very low levels insomatostatin and neuronal nitric oxide synthase interneurons. Despite thehigh expression of this protein in the striatum, only a minority of NMDAreceptors are linked to alpha-actinin-2. This interaction may identify asubset of receptors with distinct anatomical and functional properties.

• Hohmann AG, Herkenham M
• Localization of cannabinoid CB(1) receptor mRNA in neuronal subpopulationsof rat striatum: a double-label in situ hybridization study.
• Synapse. 2000; 37: 71-80
• Display abstract

• Double-label in situ hybridization was used to identify the phenotypes ofstriatal 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-labeledriboprobes for striatal projection neurons (preprotachykinin A,prodynorphin, and preproenkephalin mRNAs) and interneurons (vesicularacetylcholine 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) mRNAexpression was observed at rostral and mid-striatal levels; in the samecoronal sections the number of silver grains per cell ranged from belowthe threshold of detectability at the medial and ventral poles tosaturation at the dorsolateral boundary bordered by the corpus callosum.At the caudal level examined, CB(1) mRNA was denser in the ventral sectorrelative to the dorsal sector. Virtually all neurons expressing mRNAmarkers for striatal projection neurons colocalized CB(1) mRNA. Combininga (35)S-labeled riboprobe for CB(1) with digoxigenin-labeled riboprobesfor both preproenkephalin and prodynorphin confirmed localization of CB(1)mRNA to striatonigral and striatopallidal neurons expressing prodynorphinand preproenkephalin mRNAs, respectively. However, CB(1) mRNA-positivecells that failed to coexpress the other markers were also apparent. CB(1)mRNA was localized to putative GABAergic interneurons that express highlevels of GAD67 mRNA. These interneurons enable functional interactionsbetween the direct and indirect striatal output pathways. By contrast,aspiny interneurons that express preprosomatostatin mRNA and cholinergicinterneurons that coexpress ChAT and VAChT mRNAs were CB(1) mRNA-negative.The present data provide direct evidence that cannabinoid receptors aresynthesized in striatonigral neurons that contain dynorphin and substanceP and striatopallidal neurons that contain enkephalin. By contrast, localcircuit neurons in striatum that contain somatostatin or acetylcholine donot synthesize cannabinoid receptors. Published 2000 Wiley-Liss, Inc.

• Yung KK, Ng TK, Wong CK
• Subpopulations of neurons in the rat neostriatum display GABABR1 receptorimmunoreactivity.
• Brain Res. 1999; 830: 345-52
• Display abstract

• Immunoreactivity for gamma aminobutyric acid BR1 receptor (GABABR1) wasdetected in the neuropilar elements as well as in the perikarya of neuronsin the neostriatum. Many of the GABABR1-immunoreactive perikarya weremedium-sized with a thin rim of cytoplasm. They resembled the morphologyof medium spiny neurons, the projection neurons of the neostriatum. Inaddition, some GABABR1-immunoreactive neurons were densely labeled andwere of medium to large in size. These neurons were characterized bydouble immunofluorescence using their neurochemicals as markers. Over 90%of the parvalbumin- and choline acetyltransferase-immunoreactive neuronsand about 80% of the nitric oxide synthase-immunoreactive neuronsdisplayed GABABR1 immunoreactivity. The present results show for the firsttime that the major four subpopulations of striatal neurons expressGABABR1 receptor and may have a functional implication in the GABAneurotransmission in the microcircuitry of the neostriatum.

• Sidibe M, Smith Y
• Thalamic inputs to striatal interneurons in monkeys: synaptic organizationand co-localization of calcium binding proteins.
• Neuroscience. 1999; 89: 1189-208
• Display abstract

• Recent studies indicate that extrinsic inputs from sensorimotor regions ofthe cerebral cortex and the centromedian intralaminar thalamic nucleusterminate preferentially upon specific subpopulations of striatal outputneurons in monkeys. The objective of the present study was to verifywhether this specificity of innervation also characterizes the synapticinteractions between thalamic inputs from the centromedian nucleus and thefour major populations of striatal interneurons. This was achieved bydouble labelling techniques at the electron microscope level, combiningthe anterograde transport of biotinylated-dextran amine with theimmunostaining for specific markers of striatal interneurons(somatostatin, parvalbumin, choline acetyltransferase and calretinin).Injections of biotinylated-dextran amine in the centromedian nucleus ledto dense bands of anterograde labelling which, in double immunostainedsections, largely overlapped with the four populations of interneurons inthe post-commissural region of the putamen. In the electron microscope,biotinylated-dextran amine-containing terminals formed asymmetricaxo-dendritic synapses with somatostatin-, parvalbumin-, and cholineacetyltransferase-containing elements. However, synapses betweenanterogradely labelled terminals and calretinin-positive neurons were notfound. In sections processed to localize biotinylated-dextran amine andparvalbumin or calretinin, double-labelled terminals (biotinylated-dextranamine/parvalbumin and biotinylated-dextran amine/calretinin),morphologically similar to thalamostriatal boutons, were found in thestriatum indicating that calcium binding proteins may be expressed bythalamostriatal neurons. To test this possibility, we combined theretrograde transport of lectin-conjugated horseradish peroxidase from theputamen with parvalbumin and calretinin immunostaining and found that,indeed, most of the retrogradely labelled cells in the centromediannucleus displayed parvalbumin and calretinin immunoreactivity. Moreover,co-localization studies revealed that calretinin and parvalbumin co-existin single neurons of the centromedian nucleus. In conclusion, striatalinterneurons immunoreactive for somatostatin, parvalbumin and cholineacetyltransferase, but not those containing calretinin, receive stronginputs from the centromedian nucleus in monkeys. Moreover, our findingsindicate that parvalbumin and calretinin co-exist in individualthalamostriatal neurons. In combination with our previous data, theseresults suggest that thalamic information may be conveyed to striatalprojection neurons both, directly via excitatory synaptic inputs, orindirectly via striatal interneurons. The relative importance of thosedirect and indirect thalamic influences upon the activity of striataloutput neurons remains to be established.

• Chen Q, Veenman CL, Reiner A
• Cellular expression of ionotropic glutamate receptor subunits on specificstriatal neuron types and its implication for striatal vulnerability inglutamate receptor-mediated excitotoxicity.
• Neuroscience. 1996; 73: 715-31
• Display abstract

• Glutamate receptors are composed of subtype-specific subunits. Variationin the precise subunit composition of a receptor may result in significantfunctional differences. Thus, a precise knowledge of subunit compositionon striatal neurons is a prerequisite for understanding the selectivevulnerability of striatal neurons to excitatory amino acids. In thepresent study, we used an immunohistochemical double-labelling approach tolocalize ionotropic glutamate receptor subunits (NMDAR1, GluR1, GluR2/3,GluR4 and GluR5/6/7) on specific striatal neuron populations. Our resultsshowed that striatal cholinergic and somatostatin interneurons were notlabelled for the alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate,receptor subunits GluR1, GluR2/3 and GluR4. Most cholinergic andsomatostatin interneurons (83.3% to 100%), however, were double-labelledfor the N-methyl-D-aspartate receptor subunit NR1 and kainic acid receptorsubunits GluR5/6/7. All parvalbumin interneurons were labelled for GluR1and GluR4, and 96% GluR1 positive and 95% GluR4 positive neurons were alsodouble-labelled as parvalbumin interneurons. About half of all parvalbumininterneurons co-localized with GluR2/3, and over 97% were labelled for NR1and GluR5/6/7. Among striatal projection neurons, enkephalin-positive(mainly striatopallidal) neurons, striatonigral neurons (mainly substanceP-positive) and calbindin-positive matrix neurons were not immunostainedfor GluR1 or GluR4. In contrast, 95% to 100% of each of these types ofprojection neurons were double-labelled for NR1, GluR2/3 and GluR5/6/7.Our results demonstrate that striatal neuron types differ in theirexpression of ionotropic glutamate receptor subunits and subtypes. Theclear difference between striatal interneurons and projection neurons inionotropic glutamate receptor subtypes/subunits supports the idea thatdifferential glutamate receptor expression mechanism may account for theselective vulnerability of striatal projection neurons to excitotoxicity,and that glutamate receptor-mediated excitotoxicity may be involved in thestriatal neurodegenerative diseases.

• Testa CM, Standaert DG, Landwehrmeyer GB, Penney JB Jr, Young AB
• Differential expression of mGluR5 metabotropic glutamate receptor mRNA byrat striatal neurons.
• J Comp Neurol. 1995; 354: 241-52
• Display abstract

• Metabotropic glutamate receptors (mGluRs) mediate the effects of glutamateneurotransmission on intracellular second messenger systems. Among theseven distinct mGluR receptor isoforms currently identified, the mGluR5isoform is expressed particularly prominently in the striatum, where itmay contribute to neuronal plasticity, motor behaviors, and excitotoxicinjury. mGluR5 mRNA expression in striatal enkephalinergic,somatostatinergic, and cholinergic neurons was examined using double labelin situ hybridization techniques. mGluR5 expression is abundant in a largenumber of medium-sized striatal cells but is absent in a significantminority of neurons. Double label in situ hybridization with 35S-dATP- anddigoxygenin-dUTP-tailed oligonucleotide probes demonstrated that mGluR5message is highly expressed by enkephalinergic striatal neurons but is notdetectable in cholinergic or somatostatin interneurons. In addition, somenonenkephalin, presumably substance P, neurons were also strongly labeledfor mGluR5. The differential expression of mGluR5 in striatal projectionneurons vs. interneurons may contribute to the selective vulnerability ofthese neurons to disease processes.

• Tallaksen-Greene SJ, Albin RL
• Localization of AMPA-selective excitatory amino acid receptor subunits inidentified populations of striatal neurons.
• Neuroscience. 1994; 61: 509-19
• Display abstract

• Two-color immunofluorescence histochemistry and immunohistochemistry incombination with retrograde tract-tracing techniques were used to examinethe relationship of alpha-amino-3-hydroxy-5-methyl-4-isoxazole proprionicacid (AMPA)-selective glutamate receptor subunits (GluR1, GluR2/3/4c andGluR4) to identified populations of striatal projection neurons andinterneurons. The majority of striatonigral and striatopallidal neuronswere double-labeled for GluR2/3/4c. These findings were confirmed usingcalbindin to label matrix projection neurons. In contrast, immunostainingof the GluR1 subunit was not observed to co-localize with any striatalprojection neurons. Striatal interneurons immunostained for parvalbuminwere also labeled by antibodies directed against the GluR1 subunit.Approximately 50% of parvalbumin neurons also contained GluR2/3/4c.Somatostatin immunoreactivity did not co-localize with either the GluR1 orGluR2/3/4c subunits. GluR4-immunoreactive neurons were not observed instriatum. This study demonstrates that AMPA-selective glutamate receptorsare differentially localized on subpopulations of striatal neurons andinterneurons. These findings suggest that discrete striatal neuronpopulations may express different AMPA receptor subunit combinations whichmay account for their functional specificity.

• Figueredo-Cardenas G, Anderson KD, Chen Q, Veenman CL, Reiner A
• Relative survival of striatal projection neurons and interneurons afterintrastriatal injection of quinolinic acid in rats.
• Exp Neurol. 1994; 129: 37-56
• Display abstract

• An excitotoxic process mediated by the NMDA type glutamate receptor may beinvolved in striatal neuron death in Huntington's disease (HD). To explorethis possibility, we have injected an NMDA-receptor-specific excitotoxin,quinolinic acid (QA), into the striatum in adult rats and 2-4 monthspostlesion explored the relative patterns of survival for the variousdifferent types of striatal projection neurons and interneurons and forthe striatal efferent fibers in the different striatal projection areas.The perikarya of specific types of striatal neurons were identified byneurotransmitter immunohistochemical labeling or by retrograde labelingfrom striatal target areas, while the striatal efferent fiber plexuseswere identified by neurotransmitter immunohistochemical labeling. Thepattern of survival for the perikarya of each neuron type as a function ofdistance from the center of the injection site was determined, and therelative survival of each type was compared. For the fibers in targetareas, computer-assisted image analysis was used to determine the degreeof fiber loss for each projection target. In the study of perikaryalvulnerability, we found that the somatostatin-neuropeptide Y (SS/NPY)interneurons were the most vulnerable to QA and the cholinergic neuronswere invulnerable to QA. The perikarya of all projection neuron types(striatopallidal, striatonigral, and striato-entopeduncular) were lessvulnerable than the SS/NPY interneurons and more vulnerable than thecholinergic interneurons. Among projection neuron perikarya, there wasevidence of differential vulnerability, with striatonigral neuronsappearing to be the most vulnerable. Examination of immunolabeled striatalfibers in the striatal target areas indicated that striato-entopeduncularfibers better survived intrastriatal QA than did striatopallidal orstriatonigral fibers. The apparent order of vulnerability observed in thisstudy among projection neurons and/or their efferent fiber plexuses andthe invulnerability observed in this study of cholinergic interneurons issimilar to that observed in HD. The vulnerability of the SS/NPYinterneurons to QA is, however, in stark contrast to their invulnerabilityin HD. The results thus suggest that although the excitotoxin hypothesisof striatal neuron death in HD has merit, QA injections into adult ratstriatum do not strictly mimic the outcome in HD. This suggests thateither adult rats are not a completely suitable subject for mimicking HDor the HD excitotoxic process does not involve a freely circulatingexcitotoxin such as QA.