Rat Neostriatum Research Articles

Evidence on extracellular dopamine level in rat striatum: implications for the validity of quantitative microdialysis

Microdialysis zero-net-flux (ZNF) method is commonly used to monitor drug-induced changes in neurotransmitter baseline and release/uptake processes. Recent studies in this field suggest that microdialysis ZNF method seriously underestimates the resting concentration of extracellular dopamine in the rat neostriatum because probe implantation preferentially damages nearby dopamine release sites and that dopamine uptake inhibition increases the relative recovery of dopamine by microdialysis. This study assessed the validity of these claims by examining current data on extracellular dopamine levels at rest and after drug application obtained by voltammetry, a technique thought to induce less tissue disruption than microdialysis. To obtain the extracellular baseline value for dopamine from the evoked overflow data, we modified the existing dopamine kinetic model to suit both the resting and stimulated circumstances. It was found that dopamine uptake inhibition did in fact decrease the microdialysis relative recovery of dopamine, implying that the average basal extracellular dopamine level is within the range of 7-20 nm in rat striatum. This study concludes that the microdialysis ZNF method indeed underestimates the extracellular dopamine concentration, although not by as much as had been thought. Chronic microdialysis damages both neurotransmitter release and uptake, but it does so in a somewhat relative and proportional way for both processes. Thus the validity of the microdialysis ZNF method is not seriously undermined.

Presynaptic localization of an AMPA‐type glutamate receptor in corticostriatal and thalamostriatal axon terminals

The neostriatum is known to receive glutamatergic projections from the cerebral cortex and thalamic nuclei. Vesicular glutamate transporters 1 and 2 (VGluT1 and VGluT2) are located on axon terminals of corticostriatal and thalamostriatal afferents, respectively, whereas VGluT3 is found in axon terminals of cholinergic interneurons in the neostriatum. In the present study, the postsynaptic localization of ionotropic glutamate receptors was examined in rat neostriatum by the postembedding immunogold method for double labelling of VGluT and glutamate receptors. Immunoreactive gold particles for AMPA receptor subunits GluR1 and GluR2/3 were frequently found not only on postsynaptic but also on presynaptic profiles immunopositive for VGluT1 and VGluT2 in the neostriatum, and GluR4-immunoreactive particles were observed on postsynaptic and presynaptic profiles positive for VGluT1. Quantitative analysis revealed that 27-45% of GluR1-, GluR2-, GluR2/3- and GluR4-immunopositive particles found in VGluT1- or VGluT2-positive synaptic structures in the neostriatum were associated with the presynaptic profiles of VGluT-positive axons. In contrast, VGluT-positive presynaptic profiles in the neostriatum showed almost no immunoreactivity for NMDA receptor subunits NR1 or NR2A/B. Furthermore, almost no GluR2/3-immunopositive particles were observed in presynaptic profiles of VGluT3-positive (cholinergic) terminals that made asymmetric synapses in the neostriatum, or in those of VGluT1- or VGluT2-positive terminals in the neocortex. The present results indicate that AMPA receptor subunits but not NMDA receptor subunits are located on axon terminals of corticostriatal and thalamostriatal afferents, and suggest that glutamate released from these axon terminals controls the activity of the terminals through the presynaptic AMPA autoreceptors.

Adenovirally expressed noggin and brain-derived neurotrophic factor cooperate to induce new medium spiny neurons from resident progenitor cells in the adult striatal ventricular zone.

Neurogenesis from endogenous progenitor cells in the adult forebrain ventricular wall may be induced by the local viral overexpression of cognate neuronal differentiation agents, in particular BDNF. Here, we show that the overexpression of noggin, by acting to inhibit glial differentiation by subependymal progenitor cells, can potentiate adenoviral BDNF-mediated recruitment of new neurons to the adult rat neostriatum. The new neurons survive at least 2 months after their genesis in the subependymal zone and are recruited primarily as GABAergic DARPP-32+ medium spiny neurons in the caudate-putamen. The new medium spiny neurons successfully project to the globus pallidus, their usual developmental target, extending processes over several millimeters of the normal adult striatum. Thus, concurrent suppression of subependymal glial differentiation and promotion of neuronal differentiation can mobilize endogenous subependymal progenitor cells to achieve substantial neuronal addition to otherwise non-neurogenic regions of the adult brain.

Differential effects of somatostatin on exploratory behavior after unilateral injections into rat neostriatum

The effects of somatostatin (SRIF) microinjected unilaterally (left or right) at a dose of 10, 50, and 100 ng into the neostriatum of male Wistar rats on exploratory behavior were studied. Unilateral injections of SRIF suppressed dose-related the exploratory activity as decreased the number of horizontal and vertical movements compared to the respective controls. The effect was more pronounced when SRIF was microinjected into the right neostriatum as compared to the left neostriatum. These findings suggest some asymmetric effects of SRIF, depending on the dose and the microinjected hemisphere.

Differential expression of NMDA and AMPA receptor subunits in DARPP-32-containing neurons of the cerebral cortex, hippocampus and neostriatum of rats

Dopamine and cyclic adenosine 3′,5′-monophosphate-regulated phosphoprotein, 32 kDa (DARPP-32) is a key element of dopamine/D1/DARPP-32/protein phosphatase-1 (PP-1) signaling cascades of mammalian brain. We are interested in the expression patterns of N-methyl- d-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptors in DARPP-32-containing neurons, which may constitute morphological basis for interaction between dopamine and ionotropic glutamate receptors in dopaminoceptive cells. Double immunofluorescence was performed to visualize neurons showing coexpression of DARPP-32 with NMDA or AMPA receptor subunits (i.e., NR1, NR2a/b, glutamate receptor subunit 1 [GluR1], GluR2/3, and GluR4) in the forebrains of rats. Distribution of DARPP-32-positive neurons completely or partially overlapped with that of NMDA receptor- or AMPA receptor-immunoreactive ones in the frontal and parietal cortex, hippocampus and neostriatum, and neurons double-labeled with DARPP-32/NR1, DARPP-32/NR2a/b, DARPP-32/GluR1, DARPP-32/GluR2/3, or DARPP-32/GluR4 immunoreactivity were numerously observed. Semiquantification analysis indicated that most of DARPP-32-containing neurons (86–98%) expressed NR1, NR2a/b and GluR2/3, while less of them (14–90%) expressed GluR1 and GluR4. Although high rates (90–98%) of DARPP-32-positive cells expressed NMDA receptors in all regions above, variant percentages of them expressing AMPA receptor subunits were observed among the cortex (54–90%), hippocampus (59–97%) and neostriatum (14–97%). The study presents differential expression patterns of NMDA and AMPA receptors in DARPP-32-postive neurons in these forebrain regions. Taken together with previous reports, the present data suggest that interaction between dopamine and glutamate receptors may occur in the dopaminoceptive neurons with distinct receptor compositions and may be involved in modulating neuronal properties and excitotoxicity in mammalian forebrain.

Erratum: Aznavour N, Mechawar N, Watkins KC, Descarries L. 2003. Fine structural features of the acetylcholine innervation in the developing neostriatum of rat, J Comp Neurol 460:280–291.

AbstractThe original article to which this Erratum refers was published in Journal of Comparative Neurology(2003)460;280–291

Fine structural features of the acetylcholine innervation in the developing neostriatum of rat.

The acetylcholine (ACh) innervation in the developing neostriatum of rat was examined by means of light and electron microscopic immunocytochemistry with a highly sensitive antibody against choline acetyltransferase (ChAT). ChAT-immunoreactive cell bodies and their emerging processes, located at birth in the lateral part of neostriatum, progressively pervaded the whole region, to give rise to an extremely dense axonal network. As visualized and measured in single thin sections, at postnatal (P) ages P8, P16, and P32, the intrinsic and relational features of ChAT-immunostained profiles of axon varicosities in the lateral and medial parts of neostriatum were similar to those previously described in the adult. At the three postnatal ages, the immunoreactive profiles were comparable in shape, size, and vesicular content, and displayed one or more mitochondria with increasing frequency (from 10% at P8 to 29% at P32). The proportion which showed a synaptic junction was low at the three ages (8-16%), indicating an average synaptic incidence of 22% for whole varicosities after stereological extrapolation. The observed junctions were relatively small, mostly symmetrical, and made with dendritic spines or branches. The frequency of synapses on spines versus branches increased with age, from 20% at P8 to almost 60% at P32. Thus, the relational features of the neostriatal ACh innervation were similar to adult as soon as it appeared, as previously observed to be the case in the developing cerebral cortex. The diffuse mode of transmission may therefore be characteristic of both ACh interneurons (neostriatum) and projection neurons (cerebral cortex) in the CNS, and could be determining their functional properties during development as well as at maturity.

Differential expression of N-methyl- d-aspartate receptor subunit messenger ribonucleic acids and immunoreactivity in the rat neostriatum during postnatal development

The present study was performed to investigate the patterns of gene expression of N-methyl- d-aspartate (NMDA) receptors (NRs) in the rat neostriatum during postnatal development. Reverse transcriptase–polymerase chain reactions (RT–PCR) indicated that levels of NR1, NR2A and NR2D mRNAs reached peak levels between postnatal days 7 (PND 7) and PND 14. The levels of NR2B and NR2C mRNAs were low at PND 1 and their levels increased at PND 7 and remained high in adults. Immunofluorescence combined with image analysis revealed that the levels of NR1 immunoreactivity rose to its maximum at PND 14. In contrast, NR1 immunoreactivity rose progressively in perikarya of striatal neurons. Levels of NR2A immunoreactivity in the neostriatum were highest in adults. However, levels of NR2A immunoreactivity were higher in striatal neurons at PND 1 and PND 7. Levels of NR2B immunoreactivity were highest at PND 7. In the perikarya of striatal neurons however, the highest levels of NR2B immunoreactivity were detected at PND 14 and adult striatal neurons. In addition, double immunofluorescence revealed that the levels of NR1 immunoreactivity increased but the levels of NR2A immunoreactivity were the same in parvalbumin (PV)-positive striatal interneurons of PND 14 and adult rats. NR2B immunoreactivity was not detected in PV-positive neurons of PND 14 rats, but intense NR2B labeling was seen in PV-positive neurons of adult rats. Last but not least, in choline acetyltransferase (ChAT)-positive striatal interneurons of PND 14 and adult rats, levels of NR1 and NR2A immunoreactivity was seen to increase. Level of NR2B immunoreactivity remained the same in ChAT-positive neurons of PND 14 and adult rats. The present results indicate that there are differential patterns of expression of NR mRNAs and immunoreactivity in the neostriatum during different stages of postnatal development. Different combinations of NR have been found in different subpopulations of striatal neurons at different developmental stages. NR1, NR2A and NR2B are the major NMDA receptor subunits expressed during development. The change in patterns of expression of NR may be related to the functional maturation of neurons in the neostriatum.

Expression of regeneration-related molecules in injured and regenerating striatal and nigral neurons.

Peripheral nerve grafts in the neostriatum promote axonal regeneration from restricted classes of CNS neuron, principally cells in the substantia nigra pars compacta (SNpc) and striatal cholinergic interneurons. We have examined the molecular responses of CNS neurons induced to regenerate axons by tibial nerve grafting to the neostriatum of adult rats. Brain sections were probed for mRNAs for the transcription factor c-jun, and the cell recognition molecule CHL1, or immunoreacted for TrkA or p75, 1 day to 29 weeks after grafting (dpo; wpo). In unoperated rats, scattered neurons throughout the neostriatum showed weak signals for CHL1 mRNA and slightly stronger signals for c-jun mRNA. Cells of similar appearance strongly expressed TrkA but possessed little p75. By 1 dpo, many neostriatal neurons of various sizes and GFAP + glial cells near the host/graft interface had upregulated CHL1 mRNA, c-jun mRNA and p75. Most of the larger (20-25 microm diameter) CHL1 mRNA+ cells were also TrkA+, indicating that they were NGF-sensitive cholinergic interneurons. From two weeks postgrafting, high levels of CHL1 and c-jun mRNAs and p75 in the neostriatum were confined to a few presumptive cholinergic interneurons; p75+ cells were also TrkA+ and were larger than TrkA+ neurons on the contralateral side. Retrograde labelling showed that most p75+ and some TrkA+ neurons regenerated axons through the graft. Neurons in the SNpc showed a moderate to strong signal for CHL1 mRNA, weaker signal for c-jun mRNA, and no p75 or TrkA. Some SNpc cells upregulated c-jun mRNA after graft implantation, although they did not upregulate CHL1 mRNA, p75 or TrkA. Since neostriatal neurons which regenerate axons into grafts express receptors for NGF, and grafts mimic the effects of NGF treatment on these cells, sensitivity to graft-derived NGF may be a determinant of their high regenerative capacity. The finding that c-jun and CHL1 are consistently expressed by CNS neurons induced to regenerate their axons strongly supports the idea that these molecules are directly involved in axonal regeneration.

Tyrosine kinase B and C receptors in the neostriatum and nucleus accumbens are co-localized in enkephalin-positive and enkephalin-negative neuronal profiles and their expression is influenced by cocaine

Single- and double-label immunohistochemistry were used to determine the extent to which the tyrosine kinase B and C receptors, are expressed in enkephalin-immunopositive or enkephalin-immunonegative neuronal profiles in the rat neostriatum and nucleus accumbens. Results indicate that tyrosine kinase B and C receptors are co-localized in both enkephalin-positive and enkephalin-negative neurons in both of these nuclei, which suggests that these receptors influence both the striatal–pallidal (enkephalin) and striatal–ventral mesencephalic (substance P/dynorphin) pathways. We also examined the influence of acute or repeated injections of cocaine on the number of tyrosine kinase B and C receptors immunoreactive neuronal profiles in the rat neostriatum and nucleus accumbens. Following an acute injection of cocaine (15 mg/kg, i.p.), there were significant decreases in the number of tyrosine kinase B and C receptors immunoreactive profiles in specific regions of the neostriatum and nucleus accumbens relative to saline-pretreated rats. One or 14 days following the last of seven daily injections of 15 mg/kg cocaine or saline there were no differences in the numbers of tyrosine kinase B or C receptors immunoreactive neuronal profiles between these treatment groups. Collectively, the present results indicate that tyrosine kinase B and C receptors in the neostriatum and nucleus accumbens are co-localized in enkephalin-positive and enkephalin-negative neuronal profiles, which suggests that the striatal medium spiny neurons expressing tyrosine kinase B and C receptors include those that project to the pallidum or the ventral mesencephalon. The current results also show that an acute injection of cocaine results in a decrease in the number of tyrosine kinase B and C receptors immunoreactive neuronal profiles in specific regions of the nucleus accumbens and neostriatum, indicating that cocaine-induced increases in extracellular dopamine in the striatal complex result in compensatory decreases in the expression of tyrosine kinase B and C receptors.

LONG TERM PRODUCTION OF REACTIVE OXYGEN SPECIES DURING PERINATAL ASPHYXIA IN THE RAT CENTRAL NERVOUS SYSTEM: EFFECTS OF HYPOTHERMIA

The formation of oxygen-derived free radicals in hypoxic and ischemic/reperfused brains has been proposed as an important step that links brain injury to neuronal death. Previously, we have demonstrated that reactive oxygen species (ROS) production was significantly increased in rat neostriatum during acute perinatal asphyxia (PA) in pups. In this article, we have studied the time course of ROS production in the neostriatum and neocortex of adult rats subjected to PA using electron spin resonance spectrometry (ESR) in order to record ROS production. Further more, we analyzed the actions of hypothermia on ROS release in pups and adult rats. We used for this study 6-month-old rats that suffered sub-severe and severe PA when they were pups. The most significant production of ROS was detected either in the neostriatum or neocortex at 19 and 20 min of PA. Hypothermia during 20 and 100 min at 15°C prevented ROS formation either in pups and adult rats. These data further support the concept that free radicals may contribute to the brain injury alterations and that hypothermia can prevent long-term sequelae induced by PA.

Effects of acute toxic doses of psychostimulants on extracellular levels of excitatory amino acids and taurine in rats: comparison of d-amphetamine and sydnocarb.

We used microdialysis to study how acute toxic doses of d-amphetamine and sydnocarb [3-(beta-phenylisopropyl)-N-phenylcarbamoylsydnonimine], an original Russian psychostimulant, affect extracellular levels of glutamate, aspartate, and taurine in the neostriatum of halothane-anesthetized male Sprague-Dawley rats. The administration of d-amphetamine (5.0 mg/kg x 4 i.p.) caused gradual fivefold increases in the extracellular glutamate and taurine levels and moderate increases in the extracellular aspartate level. Sydnocarb administration (23.8 mg/kg x 4 i.p., a dose equimolar to 5.0 mg/kg d-amphetamine) elicited a marked increase in the extracellular aspartate level and a small increase in the extracellular level of glutamate. The extracellular taurine level increased only after the last (fourth) injection. We conclude that a massive increase in extracellular taurine reflects hyperactivation of glutamatergic neurotransmission elicited by acute toxic dose of d-amphetamine. Sydnocarb seems to be less neurotoxic than d-amphetamine, because it elicits lesser changes in the extracellular levels of glutamate and taurine.

Receptor-mediated internalization of [ 3H]–neurotensin in synaptosomal preparations from rat neostriatum

Following its binding to somatodendritic receptors, the neuropeptide neurotensin (NT) internalizes via a clathrin-mediated process. In the present study, we investigated whether NT also internalizes presynaptically using synaptosomes from rat neostriatum, a region in which NT1 receptors are virtually all presynaptic. Binding of [ 3H]–NT to striatal synaptosomes in the presence of levocabastine to block NT2 receptors is specific, saturable, and has NT1 binding properties. A significant fraction of the bound radioactivity is resistant to hypertonic acid wash indicating that it is internalized. Internalization of [ 3H]–NT, like that of [ 125I]–transferrin, is blocked by sucrose and low temperature, consistent with endocytosis occurring via a clathrin-dependent pathway. However, contrary to what was reported at the somatodendritic level, neither [ 3H]–NT nor [ 125I]–transferrin internalization in synaptosomes is sensitive to the endocytosis inhibitor phenylarsine oxide. Moreover, treatment of synaptosomes with monensin, which prevents internalized receptors from recycling to the plasma membrane, reduces [ 3H]–NT binding and internalization, suggesting that presynaptic NT1 receptors, in contrast to somatodendritic ones, are recycled back to the plasma membrane. Taken together, these results suggest that NT internalizes in nerve terminals via an endocytic pathway that is related to, but is mechanistically distinct from that responsible for NT internalization in nerve cell bodies.

Intrinsic GABA neurons inhibit proenkephalin gene expression in slice cultures of rat neostriatum

In the neostriatum, the proenkephalin gene is expressed in medium spiny GABA neurons, which project to the globus pallidus. The expression is activated by glutamatergic projections from the neocortex via NMDA receptors. In these experiments we have used slice cultures of rat neostriatum to study the role of GABA in proenkephalin gene expression. Our results show that GABA is released from neostriatal neurons and negatively regulates the proenkephalin gene expression induced by NMDA receptor stimulation. The GABAA receptors involved seem to be colocalized with NMDA receptors on the projection neurons, which express the proenkephalin gene. In further experiments, we have found that the proenkephalin gene expression is not only activated by neocortical projection neurons but also by intrinsic striatal neurons as well as by projections from the thalamus. All these glutamatergic afferents enhance the proenkephalin gene expression via NMDA receptors. Their efficacy is regulated by endogenous GABA.

Modulation of the gene expression of N-methyl- d-aspartate receptor NR2B subunit in the rat neostriatum by a single dose of specific antisense oligodeoxynucleotide

N-methyl- d-aspartate receptors (NRs) are a group of ionotropic glutamate receptors in the brain and they are composed of heteromeric subunits (NR1, NR2A-D and NR3). In the neostriatum, a brain region that is associated with movement in animals, NMDA channels are known to involve in the motor control. Our previous report (Lai et al., 2000, Neuroscience 98, 493-500) has shown that a single dose of antisense oligodeoxynucleotides that are specific to NR1 subunit results in blockage of the gene expression of NR1 as well as NR2A subunits in the neostriatum. In the present study, antisense oligodeoxynucleotides that are specific to NR2B (ANR2B) were then employed as molecular tools to further investigate the molecular interactions of NMDA receptor subunits in the neostriatum. A single dose of ANR2B was injected unilaterally into the rat neostriatum. After one day of injection, no modification of motor behavior was found in the ANR2B-injected rats. The mRNA level of NR2B in the ANR2B-injected neostriatum was found to be decreased (−20.4%) by reverse transcriptase polymerase chain reaction (RT-PCR). However, the mRNA levels of NR1, NR2A, NR2C and NR2D in the ANR2B-treated neostriatum were found to be unchanged. After two days of injection, NR2B immunoreactivity was found to decrease in the ANR2B-treated neostriatum by immunofluorescence (−35.1%). At higher magnification, NR2B immunoreactivity was found to decrease in presumed spiny neurons of the neostriatum (−23.4%). No change in NR1 immunoreactivity was observed. These results indicate that a single dose of ANR2B can successfully block the gene expression of NR2B in neurons of the neostriatum and there is less effect on NR1 and other NR2 subunits. The blockage of the gene expression of NR2B is therefore specific and the present results may provide important implications in applications of antisense in research and in clinical therapy of neurological diseases.

Changes in reactive oxygen species (ROS) production in rat brain during global perinatal asphyxia: an ESR study

A large body of evidence suggests that the production of reactive oxygen species (ROS) can play an important role in ischemic neuronal injury. However any studies has been performed in hypoxic conditions. In the present experiments we studied using electron spin resonance (ESR) techniques the ROS release in neostriatum of newborn rats subjected to acute perinatal asphyxia (PA) followed by various periods of reoxygenation. Pregnant rats’ uteri still containing foetuses were taken out and subjected to PA by immersion in a 37°C water bath during the following periods of time: 5, 10, 15, 19 and 20 min. After performing PA, animals were recovered and ROS measured after 0, 5, 15, 30 or 60 min of reoxygenation. Then, pups were sacrificed, their neostriatum removed and homogenised with N-tert.-butyl-α-phenylnitrone (PBN) and diethylenetriamine-pentacetic acid (DPTA) in phosphate-buffered saline (PBS) and the formed complexes were extracted with ethyl acetate an analysed using an X-band ESR spectrometer. A significant release of ROS was detected at 19 and 20 min of PA after 5 min of reoxygenation. These data provide strong evidence that ROS could be involved in neuronal damage during PA.

Enhanced excitatory synaptic transmission in spiny neurons of rat striatum after unilateral dopamine denervation

The synaptic transmission and intrinsic membrane properties of spiny neurons in rat neostriatum were studied after unilateral dopamine depletion using in vivo intracellular recording and staining techniques. Two to four weeks after dopamine denervation, the spontaneous firing rate of spiny neurons increased and the spontaneous membrane potential fluctuation stayed at a more depolarized state for longer periods of time. The amplitude of cortically evoked initial excitatory postsynaptic potentials increased and a late excitatory postsynaptic potential that was occasionally found in control neurons was elicited from 23% of spiny neurons after dopamine denervation. No significant changes in intrinsic membrane properties of spiny neurons were observed after dopamine denervation. These results suggest that dopamine inhibits excitatory synaptic transmission of spiny neurons in naı̈ve animals.

Effects of amphetamine and sydnocarb on dopamine release and free radical generation in rat striatum

Microdialysis technique was used to compare the effects of four repeated intraperitoneal administrations of two psychostimulant drugs, d-amphetamine and sydnocarb, at the equimolar doses 5 and 23.8 mg/kg, respectively, on the extracellular level of dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC) and hydroxyl radicals (·OH) in the dorsal striatum of freely moving 3-month-old male Wistar rats 250–300 g in weight. d-Amphetamine caused immediate increase of DA concentration up to 950% with quick decline towards baseline values thereafter, followed by much less increase after further injections. Sydnocarb elicited moderate elevation in DA level achieving 400% after the fourth injection. d-Amphetamine induced deep decrease in DOPAC concentration, while sydnocarb caused its increase after the first and second dosing. Both drugs enhanced generation of ·OH, the effect of d-amphetamine was more pronounced. d-Amphetamine induced more intensive stereotyped behavior in rats compare to sydnocarb. It is concluded that the psychostimulant action of sydnocarb is accompanied by facilitation of the central dopaminergic transmission in rat neostriatum and followed by less pronounced neurotoxic effect than that of d-amphetamine.

Differential changes of synaptic transmission in spiny neurons of rat neostriatum following transient forebrain ischemia

Spiny neurons in neostriatum are vulnerable to cerebral ischemia. To reveal the mechanisms underlying the postischemic neuronal damage, the spontaneous activities, evoked postsynaptic potentials and membrane properties of spiny neurons in rat neostriatum were compared before and after transient forebrain ischemia using intracellular recording and staining techniques in vivo. In control animals the membrane properties of spiny neurons were about the same between the left and right neostriatum but the inhibitory synaptic transmission was stronger in the left striatum. After severe ischemia, the spontaneous firing and membrane potential fluctuation of spiny neurons dramatically reduced. The cortically evoked initial excitatory postsynaptic potentials were suppressed after ischemia indicated by the increase of stimulus threshold and the rise time of these components. The paired-pulse facilitation test indicated that such suppression might involve presynaptic mechanisms. The inhibitory postsynaptic potentials in spiny neurons were completely abolished after ischemia and never returned to the control levels. A late depolarizing postsynaptic potential that was elicited from ∼5% of the control neurons by cortical stimulation could be evoked from ∼30% of the neurons in the left striatum and ∼50% in the right striatum after ischemia. The late depolarizing postsynaptic potential could not be induced after acute thalamic transection. The intrinsic excitability of spiny neurons was suppressed after ischemia evidenced by the significant increase of spike threshold and rheobase as well as the decrease of repetitive firing rate following ischemia. The membrane input resistance and time constant increased within 6 h following ischemia and the amplitude of fast afterhyperpolarization significantly increased after ischemia. These results indicate the depression of excitatory monosynaptic transmission, inhibitory synaptic transmission and excitability of spiny neurons after transient forebrain ischemia whereas the excitatory polysynaptic transmission in neostriatum was potentiated. The facilitation of excitatory polysynaptic transmission is stronger in the right neostriatum than in the left neostriatum after ischemia. The suppression of inhibitory component and the facilitation of excitatory polysynaptic transmission may contribute to the pathogenesis of neuronal injury in neostriatum after transient cerebral ischemia.

Differential expression of GABA(B)R1 and GABA(B)R2 receptor immunoreactivity in neurochemically identified neurons of the rat neostriatum.

Gamma-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the neostriatum. Functions of GABA are known to mediate GABA(A) and GABA(B) receptors. A functional GABA(B) receptor is known to compose of heteromeric subunits, namely the GABA(B)R1 and GABA(B)R2 subunits. Our previous report (Yung et al. [1999] Brain Res. 830:345-352) has demonstrated that all major subpopulations of striatal neurons express GABA(B)R1 immunoreactivity. The cellular localization of the second subunit of GABA(B) receptor protein, i.e., GABA(B)R2 immunoreactivity, in the rat neostriatum is not yet known. By using a new commercially available specific antibody against GABA(B)R2, immunofluorescence was performed to investigate the cellular expression of GABA(B)R2 in neurochemically identified subpopulations of neurons in the rat neostriatum. Immunoreactivity for GABA(B)R2 was primarily found in the neuropil of the rat neostriatum. Double labeling revealed that those perikarya that expressed immunoreactivity for parvalbumin, choline acetyltransferase, nitric oxide synthase, glutamate receptor two, N-methyl-D-aspartate receptor one, or GABA(A)alpha1 receptor, respectively, did not express GABA(B)R2 immunoreactivity. In addition, perikarya and most of the neuropilar elements in the neostriatum that expressed glutamic acid decarboxylase 67 immunoreactivity were found to be GABA(B)R2-negative. In contrast, immunoreactivity for GABA(B)R1 was found to be expressed by all of the above neuronal subpopulations. Moreover, a vast number of SV2-immunoreactive profiles and a number of tyrosine hydroxylase-immunoreactive profiles in the neuropil of the neostriatum were found to display GABA(B)R2 immunoreactivity. The present results indicate that there is a differential expression of GABA(B)R2 and GABA(B)R1 immunoreactivity in different subpopulations of striatal neurons that are identified by their specific neurochemical markers. Immunoreactivity for GABA(B)R2 is likely to localize in neuropilar elements of the neostriatum that may belong to non-GABAergic elements. These findings provide anatomical evidence of GABA(B)R2 receptor localization in the neostriatum that may have an important functional implication of the GABA(B)-mediated functions in neurons of the neostriatum.