Yukio Takamatsu

A rotarod test for evaluation of motor skill learning.

The rotarod test is widely used to evaluate the motor coordination of rodents, and is especially sensitive in detecting cerebellar dysfunction. However, mice with striatal dopamine depletion show only mild or no motor deficit on the typical accelerating rotarod. This suggests that dopamine-depleted mice are useful as animal models for non-motor symptoms, because the influence of motor deficit is minimum and easy to discriminate from cognitive aspects of the behavioral change. The typical accelerating rotarod test is designed to evaluate maximal motor performance and is not optimized to detect motor skill learning. In an attempt to make the test more selective to motor skill learning rather than maximal gait performance, we modified the rotarod test by using a slowly rotating large drum to obtain a steep learning curve. Furthermore, administration of nomifensine, a dopamine uptake inhibitor, improved the learning. On the other hand, apomorphine, an agonist of dopamine autoreceptor, a dopaminergic toxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) impaired the learning. These pharmacological profiles fit the involvement of the so-called phasic dopamine neurotransmission. Using our modified procedure, we found impaired learning of Parkin-deficit mice, which has not been detected in typical accelerating rotarod. The modified rotarod test would be useful for evaluation of dopamine involvement in the acquisition of motor skill learning.

Rapamycin reverses impaired social interaction in mouse models of tuberous sclerosis complex

Impairment of reciprocal social interaction is a core symptom of autism spectrum disorder. Genetic disorders frequently accompany autism spectrum disorder, such as tuberous sclerosis complex caused by haploinsufficiency of the TSC1 and TSC2 genes. Accumulating evidence implicates a relationship between autism spectrum disorder and signal transduction that involves tuberous sclerosis complex 1, tuberous sclerosis complex 2 and mammalian target of rapamycin. Here we show behavioural abnormalities relevant to autism spectrum disorder and their recovery by the mammalian target of rapamycin inhibitor rapamycin in mouse models of tuberous sclerosis complex. In Tsc2+/− mice, we find enhanced transcription of multiple genes involved in mammalian target of rapamycin signalling, which is dependent on activated mammalian target of rapamycin signalling with a minimal influence of Akt. The findings indicate a crucial role of mammalian target of rapamycin signalling in deficient social behaviour in mouse models of tuberous sclerosis complex, supporting the notion that mammalian target of rapamycin inhibitors may be useful for the pharmacological treatment of autism spectrum disorder associated with tuberous sclerosis complex and other conditions that result from dysregulated mammalian target of rapamycin signalling.

Fluoxetine as a potential pharmacotherapy for methamphetamine dependence: studies in mice.

Abstract:  The monoamine transporters are the main targets of psychostimulant drugs, including methamphetamine (METH) and cocaine. Interestingly, the rewarding effects of cocaine are retained in dopamine transporter (DAT) knockout (KO) mice, while serotonin transporter (SERT) and DAT double KO mice do not exhibit conditioned place preference (CPP) to cocaine. These data suggest that SERT inhibition decreases the rewarding effects of psychostimulants. To further test this hypothesis, in the present study, we investigated the effects of intraperitoneal (i.p.) injections of 20 mg/kg fluoxetine, a selective serotonin reuptake inhibitor (SSRI), on 2 mg/kg METH (i.p.) CPP and locomotor sensitization to 1 mg/kg METH (i.p.) in C57BL/6J mice. Fluoxetine treatment before both the conditioning and preference tests abolished METH CPP. A two‐way analysis of variance (ANOVA) revealed that METH CPP tended to be lower in mice pretreated with fluoxetine before the preference test than in control mice pretreated with saline before the preference test. Furthermore, pretreatment with fluoxetine had inhibitory effects on METH‐induced locomotor sensitization. These results suggest that fluoxetine, a widely used medication for depression, may be also a useful tool for treating METH dependence.

Effects of MDMA on Extracellular Dopamine and Serotonin Levels in Mice Lacking Dopamine and/or Serotonin Transporters

3,4-Methylendioxymethamphetamine (MDMA) has both stimulatory and hallucinogenic properties which make its psychoactive effects unique and different from those of typical psychostimulant and hallucinogenic agents. The present study investigated the effects of MDMA on extracellular dopamine (DAex) and serotonin (5-HTex) levels in the striatum and prefrontal cortex (PFC) using in vivo microdialysis techniques in mice lacking DA transporters (DAT) and/or 5-HT transporters (SERT). subcutaneous injection of MDMA (3, 10 mg/kg) significantly increased striatal DAex in wild-type mice, SERT knockout mice, and DAT knockout mice, but not in DAT/SERT double-knockout mice. The MDMA-induced increase in striatal DAex in SERT knockout mice was significantly less than in wildtype mice. In the PFC, MDMA dose-dependently increased DAex levels in wildtype, DAT knockout, SERT knockout and DAT/SERT double-knockout mice to a similar extent. In contrast, MDMA markedly increased 5-HTex in wildtype and DAT knockout mice and slightly increased 5-HTex in SERT-KO and DAT/SERT double-knockout mice. The results confirm that MDMA acts at both DAT and SERT and increases DAex and 5-HTex.

Association of morphine-induced antinociception with variations in the 5'flanking and 3' untranslated regions of the μ opioid receptor gene in 10 inbred mouse strains

Objective Genetic factors are hypothesized to be involved in interindividual differences in opioid sensitivity. Inbred mouse strains that are genetically different and isogenic within each strain are useful for elucidating the genetic mechanisms underlying the interindividual differences in opioid-induced analgesia. Methods We examined the effects of morphine in 10 inbred mouse strains, including wild-derived strains that have a wide range of genetic diversity, including BLG2, CHD, KJR, MSM, NJL, PGN2, and SWN. We also performed full sequencing of the 5′ flanking region and exons of the mouse μ opioid receptor gene Oprm1 and analyzed the association between genotypes and phenotypes in these mice. Results The effects of morphine on locomotor activation and antinociception varied among the inbred strains. The nucleotide differences that cause amino acid substitutions were not found in the Oprm1 gene in the inbred strains analyzed in this study. In the 5′ flanking region and 3′ untranslated region of the Oprm1 gene, four highly variable regions containing novel short tandem repeat polymorphisms (GA, T, TA, and CA/CT) were identified. The GA, T, and TA repeat numbers were significantly associated with morphine-induced antinociception. Conclusion These results suggest that the short tandem repeats in the 5′ flanking and 3′ untranslated regions of the μ opioid receptor gene are involved in interstrain differences in opioid sensitivity in mice. Wild-derived inbred mouse strains with different numbers of these repeats may be useful models for examining interindividual differences in opioid sensitivity.

G protein-activated inwardly rectifying K+ channel inhibition and rescue of weaver mouse motor functions by antidepressants.

Antidepressants, including tricyclic antidepressants (TCAs) and selective serotonin reuptake inhibitors (SSRIs), have been widely used for the treatment of not only depression but also other psychiatric disorders, although the molecular mechanisms of the drug effects have not yet been sufficiently revealed. Here, we investigated the in vivo effects of these antidepressants on G protein-activated inwardly rectifying K+ (GIRK) channels, which are important for regulating the excitability of various cells, by using weaver (wv) mice, which have mutant GIRK channels and show abnormal neuronal cell death and motor disturbances. First, we found that a widely used SSRI fluoxetine (also known as Prozac) effectively inhibited wv GIRK2 channels like wild-type GIRK channels, expressed in Xenopus oocytes. Next, we found that weaver motor disturbances were remarkably alleviated by chronic treatment with fluoxetine or desipramine. Furthermore, the chronic fluoxetine treatment substantially suppressed the abnormal neuronal cell death in the weaver mouse cerebellum and pontine nuclei. These results suggest that continuous inhibition of wv GIRK2 channels by a group of antidepressants caused substantial suppression of the neuronal cell death and resulted in improvement of motor abilities in weaver mice. These results provide evidence for in vivo GIRK channel inhibition by a group of antidepressants.

Differential Effects of Donepezil on Methamphetamine and Cocaine Dependencies

Abstract:  Donepezil, a choline esterase inhibitor, has been widely used as a medicine for Alzheimers disease. Recently, a study showed that donepezil inhibited addictive behaviors induced by cocaine, including cocaine‐conditioned place preference (CPP) and locomotor sensitization to cocaine. In the present study, we investigated the effects of donepezil on methamphetamine (METH)‐induced behavioral changes in mice. In counterbalanced CPP tests, the intraperitoneal (i.p.) administration of 3 mg/kg donepezil prior to 2 mg/kg METH i.p. failed to inhibit METH CPP, whereas pretreatment with 3 mg/kg donepezil abolished the CPP for cocaine (10 mg/kg, i.p.). Similarly, in locomotor sensitization experiments, i.p. administration of 1 mg/kg donepezil prior to 2 mg/kg METH i.p. failed to inhibit locomotor sensitivity to METH, whereas pretreatment with 1 mg/kg donepezil significantly inhibited locomotor sensitivity to cocaine (10 mg/kg, i.p.). These results suggest that donepezil may be a useful tool for treating cocaine dependence but not for treating METH dependence. The differences in the donepezil effects on addictive behaviors induced by METH and cocaine might be due to differences in the involvement of acetylcholine in the mechanisms of METH and cocaine dependencies.

Involvement of the N-methyl-d-aspartate receptor GluN2D subunit in phencyclidine-induced motor impairment, gene expression, and increased Fos immunoreactivity

BackgroundNoncompetitive N-methyl-d-aspartate (NMDA) receptor antagonists evoke a behavioral and neurobiological syndrome in experimental animals. We previously reported that phencyclidine (PCP), an NMDA receptor antagonist, increased locomotor activity in wildtype (WT) mice but not GluN2D subunit knockout mice. Thus, the aim of the present study was to determine whether the GluN2D subunit is involved in PCP-induced motor impairment.ResultsPCP or UBP141 (a GluN2D antagonist) induced potent motor impairment in WT mice but not GluN2D KO mice. By contrast, CIQ, a GluN2C/2D potentiator, induced severe motor impairment in GluN2D KO mice but not WT mice, suggesting that the GluN2D subunit plays an essential role in the effects of PCP and UBP141, and an appropriate balance between GluN2C and GluN2D subunits might be needed for appropriate motor performance. The level of the GluN2D subunit in the mature mouse brain is very low and restricted. GluN2D subunits exist in brainstem structures, the globus pallidus, thalamus, and subthalamic nucleus. We found that the expression of the c-fos gene increased the most among PCP-dependent differentially expressed genes between WT and GluN2D KO mice, and the number of Fos-positive cells increased after PCP administration in the basal ganglia motor circuit in WT mice but not GluN2D KO mice.ConclusionThese results suggest that the GluN2D subunit within the motor circuitry is a key subunit for PCP-induced motor impairment, which requires an intricate balance between GluN2C- and GluN2D-mediated excitatory outputs.

Involvement of the 3′ non‐coding region of the mu opioid receptor gene in morphine‐induced analgesia

Abstract  The mu opioid receptor (MOR) is known to play an essential role in morphine‐induced analgesia. MOR‐1 mRNA, the major MOR transcript, possesses a long 3′ untranslated region (3′UTR) in both mouse and human species. The sequence of the MOR‐1 3′UTR, especially that of its 3′ end region, is conserved between mice and humans. In the MOR‐1 3′UTR, AU‐rich elements (AREs) are densely localized at the 3′ end region, suggesting low stability of this mRNA. Numerous putative transcription factor‐binding motifs are located in the 3′ non‐coding regions of the mouse and human MOR genes. The CXBK mouse strain, known as a MOR‐deficient strain, possesses a decreased amount of MOR‐1 mRNA containing an abnormally long MOR‐1 3′UTR with a long nucleotide insertion. This insert might disrupt the stability of the MOR‐1 mRNA or might reduce the transcription of the MOR‐1 mRNA by separating the transcription factor‐binding motifs in the 3′ non‐coding region of the MOR gene, thereby decreasing MOR‐1 mRNA expression and attenuating morphine‐induced analgesia in CXBK mice. These recent findings suggest that the MOR‐1 3′UTR is involved in mRNA expression and in the difference in response to morphine. This possible genetic mechanism may provide a good starting point for designing effective pain treatments with opiates.

Inhibitory Role of Inducible cAMP Early Repressor (ICER) in Methamphetamine-Induced Locomotor Sensitization

Background The inducible cyclic adenosine monophosphate (cAMP) early repressor (ICER) is highly expressed in the central nervous system and functions as a repressor of cAMP response element-binding protein (CREB) transcription. The present study sought to clarify the role of ICER in the effects of methamphetamine (METH). Methods and Findings We tested METH-induced locomotor sensitization in wildtype mice, ICER knockout mice, and ICER I-overexpressing mice. Both ICER wildtype mice and knockout mice displayed increased locomotor activity after continuous injections of METH. However, ICER knockout mice displayed a tendency toward higher locomotor activity compared with wildtype mice, although no significant difference was observed between the two genotypes. Moreover, compared with wildtype mice, ICER I-overexpressing mice displayed a significant decrease in METH-induced locomotor sensitization. Furthermore, Western blot analysis and quantitative real-time reverse transcription polymerase chain reaction demonstrated that ICER overexpression abolished the METH-induced increase in CREB expression and repressed cocaine- and amphetamine-regulated transcript (CART) and prodynorphin (Pdyn) expression in mice. The decreased CART and Pdyn mRNA expression levels in vivo may underlie the inhibitory role of ICER in METH-induced locomotor sensitization. Conclusions Our data suggest that ICER plays an inhibitory role in METH-induced locomotor sensitization.