Roles of centromedian parafascicular nuclei of thalamus and cholinergic interneurons in the dorsal striatum in associative learning of environmental events.

The thalamus provides a massive input to the striatum, but despite accumulating evidence, the functions of this system remain unclear. It is known, however, that the centromedian (CM) and parafascicular (Pf) nuclei of the thalamus can strongly influence particular striatal neuron subtypes, notably including the cholinergic interneurons of the striatum (CINs), key regulators of striatal function. Here, we highlight the thalamostriatal system through the CM-Pf to striatal CINs. We consider how, by virtue of the direct synaptic connections of the CM and PF, their neural activity contributes to the activity of CINs and striatal projection neurons (SPNs). CM-Pf neurons are strongly activated at sudden changes in behavioral context, such as switches in action-outcome contingency or sequence of behavioral requirements, suggesting that their activity may represent change of context operationalized as associability. Striatal CINs, on the other hand, acquire and loose responses to external events associated with particular contexts. In light of this physiological evidence, we propose a hypothesis of the CM-Pf-CINs system, suggesting that it augments associative learning by generating an associability signal and promotes reinforcement learning guided by reward prediction error signals from dopamine-containing neurons. We discuss neuronal circuit and synaptic organizations based on in vivo/in vitro studies that we suppose to underlie our hypothesis. Possible implications of CM-Pf-CINs dysfunction (or degeneration) in brain diseases are also discussed by focusing on Parkinson's disease.

Effects of the volatile anesthetic sevoflurane on tonic GABA currents in the mouse striatum during postnatal development.

The volatile anesthetic sevoflurane, which is widely used in pediatric surgery, has proposed effects on GABAA receptor-mediated extrasynaptic tonic inhibition. In the developing striatum, medium-sized spiny projection neurons have tonic GABA currents, which function in the excitatory/inhibitory balance and maturation of striatal neural circuits. In this study, we examined the effects of sevoflurane on the tonic GABA currents of medium spiny neurons in developing striatal slices. Sevoflurane strongly increased GABAA receptor-mediated tonic conductance at postnatal days 3-35. The antagonist of the GABA transporter-1, 1-[2-[[(diphenylmethylene)imino]oxy]ethyl]-1,2,5,6-tetrahydro-3-pyridinecarboxylic acid hydrochloride further increased tonic GABA conductance during the application of sevoflurane, thereby increasing the total magnitude of tonic currents. Both GABA (5 µM) and 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridine-3-ol hydrochloride, the δ-subunit-containing GABAA receptor agonist, induced tonic GABA currents in medium spiny neurons but not in cholinergic neurons. However, sevoflurane additively potentiated the tonic GABA currents in both cells. Interestingly, 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridine-3-ol hydrochloride-sensitive neurons made a large current response to sevoflurane, indicating the contribution of the δ-subunit on sevoflurane-enhanced tonic GABA currents. Our findings suggest that sevoflurane can affect the tone of tonic GABA inhibition in a developing striatal neural network.

Imbalanced suppression of excitatory and inhibitory synaptic transmission onto mouse striatal projection neurons during induction of anesthesia with sevoflurane in vitro.

Suppression of movement during induction of anesthesia is mediated through subcortical structures. We studied the effects of a brief, 5-min application of a clinically relevant concentration of sevoflurane (two minimum alveolar concentration) on the electrophysiological activities of the medium spiny neurons (MSNs) of the striatum in brain slice preparations, using a whole-cell patch-clamp technique. We found that sevoflurane slightly depolarized principal neurons in the cortex and the striatum without a significant alteration in spike threshold. Furthermore, it depressed the peak, as well as the net, charge transfer of intrastriatally evoked inhibitory postsynaptic currents (eIPSCs) much more strongly than those of excitatory postsynaptic currents (EPSCs), and this inhibition was accompanied by an elevated paired-pulse ratio. The strong suppression of eIPSCs paralleled a significant suppression of the frequency, but not the amplitude, of miniature IPSCs (mIPSCs), and was associated with a transient increase in the frequency of spontaneous EPSCs. Treatment with the Ca(2+) channel blocker Cd(2+) restored the frequency of mIPSCs to the control level, indicating sevoflurane's strong presynaptic suppression of γ-aminobutyric acid release in the striatum. In contrast, in hippocampal CA1 pyramidal neurons sevoflurane produced an enhancement of the net charge transfer of IPSCs, while it suppressed EPSCs to an equivalent degree to that in striatal MSNs. These results suggest that, in contrast to its effects on other brain structures, sevoflurane shifts the balance between synaptic excitation and inhibition in the direction of excitation in the striatum, thereby causing involuntary movements during induction of anesthesia by sevoflurane.

Severe neurological phenotypes of Q129 DRPLA transgenic mice serendipitously created by en masse expansion of CAG repeats in Q76 DRPLA mice.

We herein provide a thorough description of new transgenic mouse models for dentatorubral-pallidoluysian atrophy (DRPLA) harboring a single copy of the full-length human mutant DRPLA gene with 76 and 129 CAG repeats. The Q129 mouse line was unexpectedly obtained by en masse expansion based on the somatic instability of 76 CAG repeats in vivo. The mRNA expression levels of both Q76 and Q129 transgenes were each 80% of that of the endogenous mouse gene, whereas only the Q129 mice exhibited devastating progressive neurological phenotypes similar to those of juvenile-onset DRPLA patients. Electrophysiological studies of the Q129 mice demonstrated age-dependent and region-specific presynaptic dysfunction in the globus pallidus and cerebellum. Progressive shrinkage of distal dendrites of Purkinje cells and decreased currents through alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid and gamma-aminobutyrate type A receptors in CA1 neurons were also observed. Neuropathological studies of the Q129 mice revealed progressive brain atrophy, but no obvious neuronal loss, associated with massive neuronal intranuclear accumulation (NIA) of mutant proteins with expanded polyglutamine stretches starting on postnatal day 4, whereas NIA in the Q76 mice appeared later with regional specificity to the vulnerable regions of DRPLA. Expression profile analyses demonstrated age-dependent down-regulation of genes, including those relevant to synaptic functions and CREB-dependent genes. These results suggest that neuronal dysfunction without neuronal death is the essential pathophysiologic process and that the age-dependent NIA is associated with nuclear dysfunction including transcriptional dysregulations. Thus, our Q129 mice should be highly valuable for investigating the mechanisms of disease pathogenesis and therapeutic interventions.

Postnatal development of tyrosine hydroxylase mRNA-expressing neurons in mouse neostriatum.

The striatum harbors a small number of tyrosine hydroxylase (TH) mRNA-containing GABAergic neurons that express TH immunoreactivity after dopamine depletion, some of which reportedly resembled striatal medium spiny projection neurons (MSNs). To clarify whether the TH mRNA-expressing neurons were a subset of MSNs, we characterized their postnatal development of electrophysiological and morphological properties using a transgenic mouse strain expressing enhanced green fluorescent protein (EGFP) under the control of the rat TH gene promoter. At postnatal day (P)1, EGFP-TH+ neurons were present as clusters in the striatum and, thereafter, gradually scattered ventromedially by P18 without regard to the striatal compartments. They were immunonegative for calbindin, but immunopositive for enkephalin (54.5%) and dynorphin (80.0%). Whole-cell patch-clamp recordings revealed at least two distinct neuronal types, termed EGFP-TH+ Type A and B. Whereas Type B neurons were aspiny and negative for the MSN marker dopamine- and cyclic AMP-regulated phosphoprotein of 32 kDa (DARPP-32), Type A neurons constituted 75% of the EGFP+ cells, had dendritic spines (24.6%), contained DARPP-32 (73.6%) and a proportion acquired TH immunoreactivity after injections of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine and 3-nitropropionic acid. The membrane properties and N-methyl-d-aspartate : non-N-methyl-d-aspartate excitatory postsynaptic current ratio of Type A neurons were very similar to MSNs at P18. However, their resting membrane potentials and spike widths were statistically different from those of MSNs. In addition, the calbindin-like, DARPP-32-like and dynorphin B-like immunoreactivity of Type A neurons developed differently from that of MSNs in the matrix. Thus, Type A neurons closely resemble MSNs, but constitute a cell type distinct from classical MSNs.

Activin plays a key role in the maintenance of long-term memory and late-LTP.

A recent study has revealed that fear memory may be vulnerable following retrieval, and is then reconsolidated in a protein synthesis-dependent manner. However, little is known about the molecular mechanisms of these processes. Activin betaA, a member of the TGF-beta superfamily, is increased in activated neuronal circuits and regulates dendritic spine morphology. To clarify the role of activin in the synaptic plasticity of the adult brain, we examined the effect of inhibiting or enhancing activin function on hippocampal long-term potentiation (LTP). We found that follistatin, a specific inhibitor of activin, blocked the maintenance of late LTP (L-LTP) in the hippocampus. In contrast, administration of activin facilitated the maintenance of early LTP (E-LTP). We generated forebrain-specific activin- or follistatin-transgenic mice in which transgene expression is under the control of the Tet-OFF system. Maintenance of hippocampal L-LTP was blocked in the follistatin-transgenic mice. In the contextual fear-conditioning test, we found that follistatin blocked the formation of long-term memory (LTM) without affecting short-term memory (STM). Furthermore, consolidated memory was selectively weakened by the expression of follistatin during retrieval, but not during the maintenance phase. On the other hand, the maintenance of memory was also influenced by activin overexpression during the retrieval phase. Thus, the level of activin in the brain during the retrieval phase plays a key role in the maintenance of long-term memory.

Compartment-specific modulation of GABAergic synaptic transmission by mu-opioid receptor in the mouse striatum with green fluorescent protein-expressing dopamine islands.

The striatum is a heterogeneous mosaic of two neurochemically, developmentally, and functionally distinct compartments: the mu-opioid receptor (MOR)-enriched striosomes and the matrix. Preferential activation of the striosomes and persistent suppression of the matrix have recently been suggested to represent neural correlates of motor stereotypy. However, little is known concerning the physiological properties of the striosomes. We made patch-clamp recordings from medium spiny neurons in identified MOR-immunoreactive "dopamine islands" as striosomes in a slice preparation taken from transgenic mice expressing green fluorescent protein in tyrosine hydroxylase mRNA-containing neurons. Striosomal neurons differed electrophysiologically from cells in the matrix in having significantly less hyperpolarized resting membrane potentials and larger input resistances, suggesting developmental differences between the two types of cells. Moreover, corticostriatal EPSCs were inhibited by MOR activation to similar extents in the two compartments, although inhibition of IPSCs was observed only in the striosomes. This MOR-induced inhibition of IPSCs was presynaptically mediated, because MOR agonist invariably decreased IPSC amplitudes when postsynaptic G-protein was inactivated, significantly increased the paired-pulse ratio of the IPSCs, and decreased the frequency but not the amplitude of miniature IPSCs. These effects of MOR were mediated principally by 4-aminopyridine-sensitive K+ conductance via a cAMP-dependent pathway, which was further augmented by previous blockade of the protein kinase C cascade. These findings suggest that MOR activation by endogenous and/or exogenous MOR-selective opioid substances differentially regulates the activities of the striosome and matrix compartments and thus plays an important role in motivated behavior and learning.

Roles of micro-opioid receptors in GABAergic synaptic transmission in the striosome and matrix compartments of the striatum.

The striatum is divided into two compartments, the striosomes and extrastriosomal matrix, which differ in several cytochemical markers, input-output connections, and time of neurogenesis. Since it is thought that limbic, reward-related information and executive aspects of behavioral information may be differentially processed in the striosomes and matrix, respectively, intercompartmental communication should be of critical importance to proper functioning of the basal ganglia-thalamocortical circuits. Cholinergic interneurons are in a suitable position for this communication since they are preferentially located in the striosome-matrix boundaries and are known to elicit a conditioned pause response during sensorimotor learning. Recently, micro-opioid receptor (MOR) activation was found to presynaptically suppress the amplitude of GABAergic inhibitory postsynaptic currents in striosomal cells but not in matrix cells. Disinhibition of cells in the striosomes is further enhanced by inactivation of the protein kinase C cascade. We discuss in this review the possibility that MOR activation in the striosomes affects the activity of cholinergic interneurons and thus leads to changes in synaptic efficacy in the striatum.

Pael receptor is involved in dopamine metabolism in the nigrostriatal system.

Pael receptor (Pael-R) has been identified as one of the substrates of Parkin, a ubiquitin ligase responsible for autosomal recessive juvenile Parkinsonism (AR-JP). When Parkin is inactivated, unfolded Pael-R accumulates in the endoplasmic reticulum and results in neuronal death by unfolded protein stress, suggesting that Pael-R has an important role in the pathogenesis of AR-JP. Here we report the analyses on Pael-R-deficient (KO) and Pael-R-transgenic (Tg) mice. The striatal dopamine (DA) level of Pael-R KO mice was only 60% of that in normal mice, while in Pael-R Tg mice, striatal 3,4-dihydroxyphenylacetic acid (DOPAC) as well as vesicular DA content increased. Moreover, the nigrostriatal dopaminergic neurons of Pael-R Tg mice are more vulnerable to Parkinson's disease-related neurotoxins while those of Pael-R KO mice are less. These results strongly suggest that the Pael-R signal regulates the amount of DA in the dopaminergic neurons and that excessive Pael-R expression renders dopaminergic neurons susceptible to chronic DA toxicity.

Chronic nicotine treatment increases GABAergic input to striatal neurons.

Recent studies suggest the involvement of the dorsal striatum in the advanced stages of drug addiction as well as motor functions. We investigated the effect of chronic nicotine treatment on GABAergic synaptic transmission in the striatum of mice. Intrastriatal stimulation evoked GABAA receptor-mediated polysynaptic inhibitory postsynaptic currents more frequently in medium-sized spiny projection neurons of mice treated chronically with nicotine (1 mg/kg, twice-daily subcutaneous injections for 10-15 days) than in those of PBS-treated mice. The multiphasic inhibitory postsynaptic currents consisted of monosynaptic early and polysynaptic, nicotinic acetylcholine receptor-mediated late components. Dihydro-beta-erythroidine, an antagonist of the non-alpha7nicotinic acetylcholine receptor, suppressed only the late inhibitory postsynaptic current. These results suggest that chronic nicotine treatment increases GABAergic input to projection neurons in the dorsal striatum.

Dopamine D4 receptor-induced postsynaptic inhibition of GABAergic currents in mouse globus pallidus neurons.

Dopamine D4 receptors (D4R) are localized in the globus pallidus (GP), but their function remains unknown. In contrast, dopamine D2 receptor activation hyperpolarizes medium spiny neurons projecting from the striatum to the GP and inhibits GABA release. However, using slice preparations from D2R-deficient [D2 knock-out (D2KO)] mice, we found that dopamine inhibited GABA(A)-receptor-mediated currents in GP neurons. The paired-pulse ratio was statistically unchanged after dopamine application but was significantly elevated in D2KO wild-type littermates (WT). Furthermore, in D2KO mice, outward currents elicited by iontophoretically applied GABA were suppressed by dopamine. Dopamine (30 microm) decreased the amplitude of miniature IPSCs in both WT and D2KO mice, but the decrease in the frequency was observed only in the former but not significantly in the latter. Dopamine-induced suppression of IPSCs was blocked by selective D4R antagonists (clozapine or 3-[4-(4-iodophenyl)piperazin-1-yl]methyl-1H-pyrrolo[2,3-b]pyridine trihydrochloride), and a D4R-selective agonist N-[[4-(2-cyanophenyl)-1-piperazinyl]methyl]-3-methyl-benzamide reversibly and dose-dependently suppressed IPSCs, whereas agonists [SKF38,393 ((+/-)-1-phenyl-2,3,4,5-tetrahydro-(1H)-3-benzazepine-7,8-diol hydrochloride) or (+)-(4aR,10bR)-3,4,4a,10b-tetrahydro-4-propyl-2H,5H-[1]benzopyrano[4,3-b]-1,4-oxazin-9-ol] or antagonists [SCH23,390 (R(+)-7-chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine hydrochloride) or sulpiride] of other receptor subtypes had little effect. In GP neurons from D4R-deficient mice, dopamine-induced inhibition of GABAergic outward currents was undetectable. D4R activation suppressed the activity of protein kinase A in GP neurons, resulting in a decrease in the amplitude of GABAergic IPSCs. These findings showed that postsynaptic activation of D4R on the GP neurons reduces GABAergic currents through the suppression of PKA activity.

Singular localization of sodium channel ß4 subunit in unmyelinated fibres and its role in the striatum.

Voltage-gated Na(+) channel ß-subunits are multifunctional molecules that modulate Na(+) channel activity and regulate cell adhesion, migration and neurite outgrowth. ß-subunits including ß4 are known to be highly concentrated in the nodes of Ranvier and axon initial segments in myelinated axons. Here we show diffuse ß4 localization in striatal projection fibres using transgenic mice that express fluorescent protein in those fibres. These axons are unmyelinated, forming large, inhibitory fibre bundles. Furthermore, we report ß4 dimer expression in the mouse brain, with high levels of ß4 dimers in the striatal projection fascicles, suggesting a specific role of ß4 in those fibres. Scn4b-deficient mice show a resurgent Na(+) current reduction, decreased repetitive firing frequency in medium spiny neurons and increased failure rates of inhibitory postsynaptic currents evoked with repetitive stimulation, indicating an in vivo channel regulatory role of ß4 in the striatum.

Protein kinase C activity alters the effect of µ-opioid receptors on inhibitory postsynaptic current in the striosomes.

We have previously reported that earlier blockade of protein kinase C (PKC) augments the suppressive effect of µ-opioid receptors (MORs) on the GABAergic inhibitory postsynaptic current (IPSC) in the MOR-rich striosomes of the striatum. Interestingly, striatal medium-spiny neurons have muscarinic acetylcholine receptor subtypes M1 and M4, among which M1 activates the phosphoinositide signaling pathway yielding PKC. In this study, we examined whether acetylcholine regulates the effects of MOR on presynaptic IPSC by binding to the M1 receptor, and found that IPSC suppression by the MOR agonist, [D-Ala-N-Me-Phe, Gly-ol]-enkephalin, was significantly augmented and prolonged by the PKC inhibitor chelerythrine and attenuated by the PKC activator, phorbol 12, 13-dibutyrate. This modulatory action by chelerythrine was mimicked by the muscarinic antagonist atropine and the M1-specific antagonist pirenzepine, whereas M2-M4 antagonists had no discernible effect. These results suggest that PKC activity modulates the effect of MOR by muscarinic receptors in the striosomes.

Acetylcholine-dopamine balance hypothesis in the striatum: an update.

The imbalance between cholinergic activity and dopaminergic activity in the striatum causes a variety of neurological disorders, such as Parkinson's disease. During sensorimotor learning, the arrival of a conditioned stimulus reporting a reward evokes a pause response in the firing of the tonically active cholinergic interneurons in targeted areas of the striatum, whereas the same stimulus triggers an increase in the firing frequency of the dopaminergic neurons in the substantia nigra pars compacta. The pause response of the cholinergic interneurons begins with an initial depolarizing phase followed by a pause in spike firing and ensuing rebound excitation. The timing of the pause phase coincides well with the surge in dopaminergic firing, indicating that a dramatic rise in dopamine (DA) release occurs while nicotinic receptors remain unbound by acetylcholine. The pause response begins with dopamine D5 receptor-dependent synaptic plasticity in the cholinergic neurons and an increased GABAergic IPSP, which is followed by a long pause in firing through D2 and D5 receptor-dependent modulation of ion channels. Inactivation of muscarinic receptors on the projection neurons eventually yields endocannabinoid-mediated, dopamine-dependent long-term depression in the medium spiny projection neurons. Breakdown of acetylcholine-dopamine balance hampers proper functioning of the cortico-basal ganglia-thalamocortical loop circuits. In Parkinson's disease, dopamine depletion blocks autoinhibition of acetylcholine release through muscarinic autoreceptors, leading to excessive acetylcholine release which eventually prunes spines of the indirect-pathway projection neurons of the striatum and thus interrupts information transfer from motor command centers in the cerebral cortex.

[Physiological interaction between acetylcholine and dopamine in the striatum].

The breakdown of the balance between acetylcholine and dopamine in the striatum is known to cause a variety of neurological diseases. Physiologically, the association between sensory cues and reward during behavioral learning gradually forms a conditional pause response in the firing of the tonically active cholinergic interneurons in the striatum. Simultaneous recordings of striatal cholinergic interneurons and midbrain dopaminergic neurons during the task revealed that the pause response was well synchronized with the increase in the firing frequency of the dopaminergic neurons. Recent studies have indicated that the content of released dopamine is proportional to the firing frequency of the dopaminergic neurons unless the nicotinic receptors are activated, but it remains unaltered if the receptors are bound by acetylcholine. Therefore, dopamine release would be dramatically increased during the pause response of the cholinergic neurons. The pause response is composed of an initial depolarization phase, a pause phase, and a rebound facilitation phase. Dopamine D5 receptor-dependent long-term potentiation underlies the initial depolarization phase, which causes the ensuing pause phase. The termination of the pause is further delayed by suppression of Ih and sodium channels through dopamine D2 receptor activation. This facilitates the synaptic efficacy of the striatal medium spiny projection neurons, which enables the execution of action commands with an improved signal-to-noise ratio.

Usp46 is a quantitative trait gene regulating mouse immobile behavior in the tail suspension and forced swimming tests.

The tail suspension test (TST) and forced swimming test (FST) are widely used for assessing antidepressant activity and depression-like behavior. We found that CS mice show negligible immobility in inescapable situations. Quantitative trait locus (QTL) mapping using CS and C57BL/6J mice revealed significant QTLs on chromosomes 4 (FST) and 5 (TST and FST). To identify the quantitative trait gene on chromosome 5, we narrowed the QTL interval to 0.5 Mb using several congenic and subcongenic strains. Ubiquitin-specific peptidase 46 (Usp46) with a lysine codon deletion was located in this region. This deletion affected nest building, muscimol-induced righting reflex and anti-immobility effects of imipramine. The muscimol-induced current in the hippocampal CA1 pyramidal neurons and hippocampal expression of the 67-kDa isoform of glutamic acid decarboxylase were significantly decreased in the Usp46 mutant mice compared to control mice. These phenotypes were rescued in transgenic mice with bacterial artificial chromosomes containing wild-type Usp46. Thus, Usp46 affects the immobility in the TST and FST, and it is implicated in the regulation of GABA action.

Calcium-permeable AMPA receptors promote misfolding of mutant SOD1 protein and development of amyotrophic lateral sclerosis in a transgenic mouse model.

Mutant Cu/Zn-superoxide dismutase (SOD1) protein aggregation has been suggested as responsible for amyotrophic lateral sclerosis (ALS), although the operative mediating factors are as yet unestablished. To evaluate the contribution of motoneuronal Ca2+-permeable (GluR2 subunit-lacking) alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA)-type glutamate receptors to SOD1-related motoneuronal death, we generated chat-GluR2 transgenic mice with significantly reduced Ca2+-permeability of these receptors in spinal motoneurons. Crossbreeding of the hSOD1G93A transgenic mouse model of ALS with chat-GluR2 mice led to marked delay of disease onset (19.5%), mortality (14.3%) and the pathological hallmarks such as release of cytochrome c from mitochondria, induction of cox2 and astrogliosis. Subcellular fractionation analysis revealed that unusual SOD1 species first accumulated in two fractions dense with neurofilaments/glial fibrillary acidic protein/nuclei and mitochondria long time before disease onset, and then concentrated into the former fraction by disease onset. All these processes for unusual SOD1 accumulation were considerably delayed by GluR2 overexpression. Ca2+-influx through atypical motoneuronal AMPA receptors thus promotes a misfolding of mutant SOD1 protein and eventual death of these neurons.