Delayed gramicidin delivery through an intra-pipette capillary facilitates perforated patch recordings.

The gramicidin-perforated patch-clamp technique is indispensable for recording neuronal activities without changing the intracellular Cl- concentration. Conventionally, gramicidin contained in the pipette fluid is delivered to the cell membrane by passive diffusion. Gramicidin deposited on the pipette orifice sometimes hampers giga-seal formation, and perforation progresses only slowly. These problems may be circumvented by delivering a high concentration of gramicidin from an intra-pipette capillary after a giga-seal is formed. We herein describe the detailed protocol of this improved method. This protocol would greatly facilitate the investigation of Cl- gradient-dependent neuronal activities.

Developmental synapse pathology triggered by maternal exposure to the herbicide glufosinate ammonium.

Environmental and genetic factors influence synapse formation. Numerous animal experiments have revealed that pesticides, including herbicides, can disturb normal intracellular signals, gene expression, and individual animal behaviors. However, the mechanism underlying the adverse outcomes of pesticide exposure remains elusive. Herein, we investigated the effect of maternal exposure to the herbicide glufosinate ammonium (GLA) on offspring neuronal synapse formation in vitro. Cultured cerebral cortical neurons prepared from mouse embryos with maternal GLA exposure demonstrated impaired synapse formation induced by synaptic organizer neuroligin 1 (NLGN1)-coated beads. Conversely, the direct administration of GLA to the neuronal cultures exhibited negligible effect on the NLGN1-induced synapse formation. The comparison of the transcriptomes of cultured neurons from embryos treated with maternal GLA or vehicle and a subsequent bioinformatics analysis of differentially expressed genes (DEGs) identified "nervous system development," including "synapse," as the top-ranking process for downregulated DEGs in the GLA group. In addition, we detected lower densities of parvalbumin (Pvalb)-positive neurons at the postnatal developmental stage in the medial prefrontal cortex (mPFC) of offspring born to GLA-exposed dams. These results suggest that maternal GLA exposure induces synapse pathology, with alterations in the expression of genes that regulate synaptic development via an indirect pathway distinct from the effect of direct GLA action on neurons.

Assessment of memory recognition using a smartphone-based test system: A pilot study.

BACKGROUND: Elucidation of the detailed nature of age-related memory decline requires analysis of memory performance in large populations of various ages. To promote large-scale studies, we developed a smartphone-based self-test for memory recognition. We examined whether this test could detect age-related memory decline and the effects of aerobic exercise on memory. METHODS: Seventy-eight younger and 42 older participants were randomly divided into active and passive groups. Both groups took a memory recognition test (consisting of 2 sessions separated by a 48-hour interval) conducted on smartphones. The participants answered the positive and negative affect schedule questionnaire at the beginning and end of each session. In the first session, the participants performed cognitive tasks on 90 photographs displayed on a smartphone screen. Immediately after the cognitive tasks, the active group performed a bout of light aerobic exercise for 10 minutes, while the passive group remained calm for 10 minutes. In the second session, the participants were tested on the recognition of 90 previously observed photographs and 90 distractor photographs. RESULTS: Passive older participants had ~40% to ~50% lower recognition scores (RSs) than passive younger participants did. Moreover, the aerobic exercise used in this study improved the RSs of active younger participants by up to ~40% compared with those of passive younger participants, while such an improvement was not observed in older participants. The RS did not depend on the affect levels evaluated using positive and negative affect schedule questionnaire. CONCLUSIONS: These results demonstrated that the smartphone-based test could detect age-related decline and could promote behavior modification that may lead to memory enhancement, as reported in previous studies using conventional laboratory tests. The results of the smartphone-based test were not influenced by the subjects affect. This indicates the possibility of large-scale memory studies and healthcare for memory performance by using personal mobile devices.

Carbon powder-filled microelectrode: An easy-to-fabricate probe for cellular electrochemistry.

Carbon fiber and carbon fiber disc microelectrodes are widely used for electrochemical detection of biochemicals released from cells. However, fabricating these types of microelectrodes is difficult and time-consuming. Here, we report an easy-to-fabricate, carbon powder-filled microelectrode consisting of a pulled glass capillary backfilled with carbon powder. Carbon tip size and responsiveness can be controlled by adjusting the settings of the puller. Carbon powder-filled microelectrodes with tip opening diameters of 7-24 µm detected sub-micromolar to sub-millimolar levels of dopamine and catecholamines released from PC-12 cells. This simple microelectrode should promote further work on cellular and tissue electrochemistry.

A test potential booster for fast-scan cyclic voltammetry with an electrophysiological amplifier.

Fast-scan cyclic voltammetry (FSCV) is a powerful technique for studying the local dynamics of neurotransmitters and neuromodulators. FSCV is attractive to researchers employing electrophysiological techniques because it can be performed using an electrophysiological voltage-clamp amplifier. However, the narrow test potential range of electrophysiological amplifiers (typically, ±1 V) limits testable species of analytes. Here we devised a booster that extends the test potential range. Using the booster, we could detect the oxidation current of adenosine peaking near a test potential of +1.5 V. The booster should promote combined electrophysiological and electrochemical studies of synaptic release and neural secretion.

G Protein-Coupled Glutamate and GABA Receptors Form Complexes and Mutually Modulate Their Signals.

Molecular networks containing various proteins mediate many types of cellular processes. Elucidation of how the proteins interact will improve our understanding of the molecular integration and physiological and pharmacological propensities of the network. One of the most complicated and unexplained interactions between proteins is the inter-G protein-coupled receptor (GPCR) interaction. Recently, many studies have suggested that an interaction between neurotransmitter GPCRs may mediate diverse modalities of neural responses. The B-type gamma-aminobutyric acid (GABA) receptor (GBR) and type-1 metabotropic glutamate receptor (mGluR1) are GPCRs for GABA and glutamate, respectively, and each plays distinct roles in controlling neurotransmission. We have previously reported the possibility of their functional interaction in central neurons. Here, we examined the interaction of these GPCRs using stable cell lines and rat cerebella. Cell-surface imaging and coimmunoprecipitation analysis revealed that these GPCRs interact on the cell surface. Furthermore, fluorometry revealed that these GPCRs mutually modulate signal transduction. These findings provide solid evidence that mGluR1 and GBR have intrinsic abilities to form complexes and to mutually modulate signaling. These findings indicate that synaptic plasticity relies on a network of proteins far more complex than previously assumed.

Monitoring of glutamate-induced excitotoxicity by mitochondrial oxygen consumption.

Dysfunction of mitochondrial activity is often associated with the onset and progress of neurodegenerative diseases. Membrane depolarization induced by Na+ influx increases intracellular Ca2+ levels in neurons, which upregulates mitochondrial activity. However, overlimit of Na+ influx and its prolonged retention ultimately cause excitotoxicity leading to neuronal cell death. To return the membrane potential to the normal level, Na+ /K+ -ATPase exchanges intracellular Na+ with extracellular K+ by consuming a large amount of ATP. This is a reason why mitochondria are important for maintaining neurons. In addition, astrocytes are thought to be important for supporting neighboring neurons by acting as energy providers and eliminators of excessive neurotransmitters. In this study, we examined the meaning of changes in the mitochondrial oxygen consumption rate (OCR) in primary mouse neuronal populations. By varying the medium constituents and using channel modulators, we found that pyruvate rather than lactate supported OCR levels and conferred on neurons resistance to glutamate-mediated excitotoxicity. Under a pyruvate-restricted condition, our OCR monitoring could detect excitotoxicity induced by glutamate at only 10 µM. The OCR monitoring also revealed the contribution of the N-methyl-D-aspartate receptor and Na+ /K+ -ATPase to the toxicity, which allowed evaluating spontaneous excitation. In addition, the OCR monitoring showed that astrocytes preferentially used glutamate, not glutamine, for a substrate of the tricarboxylic acid cycle. This mechanism may be coupled with astrocyte-dependent protection of neurons from glutamate-mediated excitotoxicity. These results suggest that OCR monitoring would provide a new powerful tool to analyze the mechanisms underlying neurotoxicity and protection against it.

Lack of modulatory effect of the SCN5A R1193Q polymorphism on cardiac fast Na+ current at body temperature.

SCN5A encodes the main subunit of the NaV1.5 channel, which mediates the fast Na+ current responsible for generating cardiac action potentials. The single nucleotide polymorphism SCN5A(R1193Q), which results in an amino acid replacement in the subunit, is common in East Asia. SCN5A(R1193Q) is often identified in patients with type 3 long QT syndrome and Brugada syndrome. However, its linkage to arrhythmic disorders is under debate. Previous electrophysiological studies performed at room temperature inconsistently reported the gain- or loss-of-function effect of SCN5A(R1193Q) on the NaV1.5 channel. More recently, it was theoretically predicted that SCN5A(R1193Q) would exert a loss-of-function effect at body temperature. Here, we experimentally assessed whether SCN5A(R1193Q) modulates the NaV1.5 channel at various temperatures including normal and febrile body temperatures. We compared voltage-gated Na+ currents in SCN5A(R1193Q)-transfected and wild-type SCN5A-transfected HEK293T cells using a whole-cell voltage-clamp technique. First, we made comparisons at constant temperatures of 25°C, 36.5°C, and 38°C, and found no difference in the conductance density, voltage dependence of gating, or time dependence of gating. This suggested that SCN5A(R1193Q) does not modulate the NaV1.5 channel regardless of temperature. Second, we made comparisons while varying the temperature from 38°C to 26°C in 3 min, and again observed no difference in the time course of the amplitude or time dependence of gating during the temperature change. This also indicated that SCN5A(R1193Q) does not modulate the NaV1.5 channel in response to an acute body temperature change. Therefore, SCN5A(R1193Q) may not be a monogenic factor that triggers arrhythmic disorders.

A mutant HCN4 channel in a family with bradycardia, left bundle branch block, and left ventricular noncompaction.

We found that a female infant presenting with left bundle branch block and left ventricular noncompaction carries uninvestigated gene mutations HCN4(G811E), SCN5A(L1988R), DMD(S2384Y), and EMD(R203H). Here, we explored the possible pathogenicity of HCN4(G811E), which results in a G811E substitution in hyperpolarization-activated cyclic nucleotide-gated channel 4, the main subunit of the cardiac pacemaker channel. Voltage-clamp measurements in a heterologous expression system of HEK293T cells showed that HCN4(G811E) slightly reduced whole-cell HCN4 channel conductance, whereas it did not affect the gating kinetics, unitary conductance, or cAMP-dependent modulation of voltage-dependence. Immunocytochemistry and immunoblot analysis showed that the G811E mutation did not impair the membrane trafficking of the channel subunit in the heterologous expression system. These findings indicate that HCN4(G811E) may not be a monogenic factor to cause the cardiac disorders.

Application of surface plasmon resonance imaging to monitoring G protein-coupled receptor signaling and its modulation in a heterologous expression system.

BACKGROUND: G protein-coupled receptors (GPCRs) are ubiquitous surface proteins mediating various biological responses and thus, important targets for therapeutic drugs. GPCRs individually produce their own signaling as well as modulate the signaling of other GPCRs. Real-time observation of GPCR signaling and modulation in living cells is key to molecular study of biological responses and pharmaceutical development. However, fluorescence imaging, the technique widely used for this purpose, requires a fluorescent dye which may inhibit biological responses or a fluorescent-tagged target protein created through time-consuming genetic manipulation. In this study, we applied two-dimensional surface plasmon resonance (SPR) imaging to monitoring the translocation of protein kinase C (PKC), a major GPCR-coupled signaling molecule in the widely used HEK293 cell lines and examined whether the signaling of, and, modulation between heterologously expressed GPCRs can be measured without fluorescent labeling. RESULTS: We cultured HEK293 cells on the gold-plated slide glass and evoked SPR at the interface between the cell's plasma membrane and the gold surface with incident light. The translocation of activated native PKC to the plasma membrane is expected to alter the incident angle-SPR extent relation, and this could be detected as a change in the intensity of light reflection from the specimen illuminated at a fixed incident angle. Direct activation of PKC with 12-O-tetradecanoylphorbol-13-acetate increased the reflection intensity. This increase indeed reported PKC translocation because it was reduced by a pre-treatment with bisindolylmaleimide-1, a PKC inhibitor. We further applied this technique to a stable HEK293 cell line heterologously expressing the GPCRs type-1 metabotropic glutamate receptor (mGluR1) and adenosine A1 receptor (A1R). (RS)-3,5-dihydroxyphenylglycine, a mGluR1 agonist, increased the reflection intensity, and the PKC inhibitor reduced this increase. A pre-treatment with (R)-N(6)-phenylisopropyladenosine, an A1R-selective agonist suppressed mGluR1-mediated reflection increase. These results suggest that our technique can detect PKC translocation initiated by ligand binding to mGluR1 and its modulation by A1R. CONCLUSIONS: SPR imaging turned out to be utilizable for monitoring GPCR-mediated PKC translocation and its modulation by a different GPCR in a heterologous expression system. This technique provides a powerful yet easy-to-use tool for molecular study of biological responses and pharmaceutical development.

SCN5A(K817E), a novel Brugada syndrome-associated mutation that alters the activation gating of NaV1.5 channel.

BACKGROUND: Brugada syndrome (BrS) is an inherited lethal arrhythmic disorder characterized by syncope and sudden cardiac death from ventricular tachyarrhythmias. Here we identified a novel K817E mutation of SCN5A gene in a man with type 1 BrS electrocardiogram pattern using next-generation sequencing targeted for 73 cardiac disorder-related genes. SCN5A encodes the α-subunit of NaV1.5 voltage-gated Na(+) channel, and some of its mutations are linked to BrS. The proband had no mutation in any of the other arrhythmia-related genes sequenced. OBJECTIVE: We investigated whether the K817E mutation causes a functional change of NaV1.5 channel responsible for the BrS phenotype. METHODS: We compared the electrophysiological properties of the whole-cell currents mediated by wild-type and mutant channels heterologously expressed in human embryonic kidney 293 cells by using a voltage-clamp technique. RESULTS: The K817E mutation reduced the Na(+) current density by 39.0%-91.4% at membrane potentials from -55 to -5 mV. This reduction resulted from a ~24-mV positive shift in the voltage dependence of activation. The mutation also decelerated recovery from both fast and intermediate inactivation, whereas it had little effect on the cell surface expression, single-channel conductance, voltage-dependence of fast inactivation, entry into intermediate inactivation, use-dependent loss of channel availability, or closed-state inactivation. CONCLUSION: The K817E mutation of SCN5A gene leads to loss of function of NaV1.5 channel and may underlie the BrS phenotype of the proband.

Complex formation and functional interaction between adenosine A1 receptor and type-1 metabotropic glutamate receptor.

The adenosine A1 receptor (A1R) is a G protein-coupled receptor (GPCR) for adenosine, a ubiquitous neuromodulator, and thus regulates neuronal excitability, as well as arousal and sensitivity to pain. In addition, we have previously described a new mode of action for A1R: in cerebellar Purkinje cells, its activation attenuates neuronal responses to glutamate, as mediated by the type-1 metabotropic glutamate receptor (mGluR1). mGluR1 is also a GPCR, and elicits such responses as long-term depression of the postsynaptic response to glutamate, a cellular basis for cerebellar motor learning. Here, we explore in greater detail the interaction between A1R and mGluR1 using non-neuronal cells. Co-immunoprecipitation and Förster resonance energy transfer (FRET) analysis reveal that A1R and mGluR1 form a complex. Furthermore, we found that mGluR1 activation inhibits A1R signaling, as measured by changes in intracellular cAMP. These findings demonstrate that A1R and mGluR1 have the intrinsic ability to form a heteromeric complex and mutually modulate signaling. This interaction may represent a new form of intriguing GPCR-mediated cellular responses.

The Susceptibilities of Human Ether-à-Go-Go-Related Gene Channel with the G487R Mutation to Arrhythmogenic Factors.

The human ether-à-go-go-related gene (hERG) channel mediates the rapid delayed rectifier potassium current (IKr) responsible for shaping the repolarization phase of cardiac action potentials. hERG mutation may cause hERG channel malfunction, leading to long QT syndrome and other arrhythmic disorders. Elucidation of the genotype-phenotype relationships of individual hERG mutations is key to the development of treatment for such arrhythmic disorders. We previously identified hERG(G487R), a missense mutant with a glycine-to-arginine substitution at position 487. In the absence of arrhythmogenic factors, hERG(G487R) subunit-containing channels show normal surface expression and gating kinetics. However, it remains unknown whether the mutation exacerbates hERG channel malfunction induced by arrhythmogenic factors. Here we used a voltage-clamp technique to compare the effects of the major arrythmogenic factors on wild-type hERG [hERG(WT)] and hERG(G487R) channel currents (IhERG) in HEK-293T cells. The extent of IhERG blockade by the antiarrhythmic drug dofetilide or E4031 was not different between these channels. On the other hand, the extracellular K(+) concentration ([K(+)]ex)-dependent changes in the rates of recovery from inactivation and deactivation of IhERG were rather less obvious for hERG(G487R) channel than for hERG(WT) channel. These findings suggest that the inheritance of hERG(G487R) does not increase the risk of arrhythmic disorders induced by antiarrhythmic drugs or hypokalemia.

Altered Actions of Memantine and NMDA-Induced Currents in a New Grid2-Deleted Mouse Line.

Memantine is a non-competitive antagonist of the N-methyl-D-aspartate (NMDA) receptor, and is an approved drug for the treatment of moderate-to-severe Alzheimer's disease. We identified a mouse strain with a naturally occurring mutation and an ataxic phenotype that presents with severe leg cramps. To investigate the phenotypes of these mutant mice, we screened several phenotype-modulating drugs and found that memantine (10 mg/kg) disrupted the sense of balance in the mutants. Moreover, the mutant mice showed an attenuated optokinetic response (OKR) and impaired OKR learning, which was also observed in wild-type mice treated with memantine. Microsatellite analyses indicated that the Grid2 gene-deletion is responsible for these phenotypes. Patch-clamp analysis showed a relatively small change in NMDA-dependent current in cultured granule cells from Grid2 gene-deleted mice, suggesting that GRID2 is important for correct NMDA receptor function. In general, NMDA receptors are activated after the activation of non-NMDA receptors, such as AMPA receptors, and AMPA receptor dysregulation also occurs in Grid2 mutant mice. Indeed, the AMPA treatment enhanced memantine susceptibility in wild-type mice, which was indicated by balance sense and OKR impairments. The present study explores a new role for GRID2 and highlights the adverse effects of memantine in different genetic backgrounds.

A590T mutation in KCNQ1 C-terminal helix D decreases IKs channel trafficking and function but not Yotiao interaction.

KCNQ1 encodes the α subunit of the voltage-gated channel that mediates the cardiac slow delayed rectifier K(+) current (IKs). Here, we report a KCNQ1 allele encoding an A590T mutation [KCNQ1(A590T)] found in a 39-year-old female with a mild QT prolongation. A590 is located in the C-terminal α helical region of KCNQ1 that mediates subunit tetramerization, membrane trafficking, and interaction with Yotiao. This interaction is known to be required for the proper modulation of IKs by cAMP. Since previous studies reported that mutations in the vicinity of A590 impair IKs channel surface expression and function, we examined whether and how the A590T mutation affects the IKs channel. Electrophysiological measurements in HEK-293T cells showed that the A590T mutation caused a reduction in IKs density and a right-shift of the current-voltage relation of channel activation. Immunocytochemical and immunoblot analyses showed the reduced cell surface expression of KCNQ1(A590T) subunit and its rescue by coexpression of the wild-type KCNQ1 [KCNQ1(WT)] subunit. Moreover, KCNQ1(A590T) subunit interacted with Yotiao and had a cAMP-responsiveness comparable to that of KCNQ1(WT) subunit. These findings indicate that the A590 of KCNQ1 subunit plays important roles in the maintenance of channel surface expression and function via a novel mechanism independent of interaction with Yotiao.

The synaptic targeting of mGluR1 by its carboxyl-terminal domain is crucial for cerebellar function.

The metabotropic glutamate receptor subtype 1 (mGluR1, Grm1) in cerebellar Purkinje cells (PCs) is essential for motor coordination and motor learning. At the synaptic level, mGluR1 has a critical role in long-term synaptic depression (LTD) at parallel fiber (PF)-PC synapses, and in developmental elimination of climbing fiber (CF)-PC synapses. mGluR1a, a predominant splice variant in PCs, has a long carboxyl (C)-terminal domain that interacts with Homer scaffolding proteins. Cerebellar roles of the C-terminal domain at both synaptic and behavior levels remain poorly understood. To address this question, we introduced a short variant, mGluR1b, which lacks this domain into PCs of mGluR1-knock-out (KO) mice (mGluR1b-rescue mice). In mGluR1b-rescue mice, mGluR1b showed dispersed perisynaptic distribution in PC spines. Importantly, mGluR1b-rescue mice exhibited impairments in inositol 1,4,5-trisphosphate receptor (IP3R)-mediated Ca(2+) release, CF synapse elimination, LTD induction, and delay eyeblink conditioning: they showed normal transient receptor potential canonical (TRPC) currents and normal motor coordination. In contrast, PC-specific rescue of mGluR1a restored all cerebellar defects of mGluR1-KO mice. We conclude that the long C-terminal domain of mGluR1a is required for the proper perisynaptic targeting of mGluR1, IP3R-mediated Ca(2+) release, CF synapse elimination, LTD, and motor learning, but not for TRPC currents and motor coordination.

Glycine/Serine polymorphism at position 38 influences KCNE1 subunit's modulatory actions on rapid and slow delayed rectifier K+ currents.

BACKGROUND: KCNE1 encodes a modulator of KCNH2 and KCNQ1 delayed rectifier K(+) current channels. KCNE1 mutations might cause long QT syndrome (LQTS) by impairing KCNE1 subunit's modulatory actions on these channels. There are major and minor polymorphismic KCNE1 variants whose 38(th) amino acids are glycine and serine [KCNE1(38G) and KCNE1(38S) subunits], respectively. Despite its frequent occurrence, the influence of this polymorphism on the K(+) channels' function is unclear. METHODS AND RESULTS: Patch-clamp recordings were obtained from human embryonic kidney -293T cells. KCNH2 channel current density in KCNE1(38S)-transfected cells was smaller than that in KCNE1(38G)-transfected cells by 34%. The voltage-sensitivity of the KCNQ1 channel current in KCNE1(38S)-transfected cells was lowered compared to that in KCNE1(38G)-transfected cells, with a +13mV shift in the half-maximal activation voltage. KCNH2 channel current density or KCNQ1 channel voltage-sensitivity was not different between KCNE1(38G)-transfected cells and cells transfected with both KCNE1(38G) and KCNE1(38S). Moreover, the KCNH2 channel current in KCNE1(38S)-transfected cells was more susceptible to E4031, a QT prolonging drug and a condition with hypokalemia, than that in KCNE1(38G)-transfected cells. CONCLUSIONS: Homozygous inheritance of KCNE1(38S) might cause a mild reduction of the delayed rectifier K(+) currents and might thereby increase an arrhythmogenic potential particularly in the presence of QT prolonging factors. By contrast, heterozygous inheritance of KCNE1(38G) and KCNE1(38S) might not affect the K(+) currents significantly. (Circ J 2014; 78: 610-618).

Identification and characterization of a novel genetic mutation with prolonged QT syndrome in an unexplained postoperative death.

INTRODUCTION: The human ether-à-go-go-related gene (hERG) encodes the α-subunit of a cardiac potassium channel. Various mutations of hERG, including missense mutations, have been reported to cause long QT syndrome (LQTS) and severe arrhythmic disorders such as sudden cardiac death. We identified a novel hERG frameshift mutation (hERG(ΔAT)) in the S5-pore region from a LQTS patient who died suddenly and analyzed its genetic profile and the molecular and electrophysiological behaviors of the protein product to assess the pathogenicity of hERG(ΔAT). METHODS AND RESULTS: We performed direct sequencing of hERG and evaluated its transcript level by using a whole blood sample from the patient. We performed immunoblotting, immunocytochemistry, and patch-clamp recordings of HEK-293 T cells transfected with hERG(ΔAT), wild-type hERG (hERG(WT)), or both. The patient demonstrated an AT deletion (c.1735_1736del) in hERG and a decrease in hERG mRNA transcripts. HEK-293 T cells showed lower production and cell surface expression of hERG(ΔAT) compared with hERG(WT) protein. In addition, the hERG(∆AT) protein failed to form functional channels, while the activation kinetics of functional channels, presumably consisting of hERG(WT) subunits, were unaffected. CONCLUSION: The ΔAT mutation may decrease the number of functional hERG channels by impairing the posttranscriptional and posttranslational processing of the mutant product. This decrease may partly explain the cardiac symptoms of the patient who was heterozygous for hERG(ΔAT).

[Activation of cerebellar B-type γ-aminobutyric acid receptor modulates optokinetic reflex adaptation].

The cerebellar cortex, the brain region responsible for motor coordination and learning expresses a high density of B-type γ-aminobutyric acid receptor (GABAbR). Previous in vitro and in situ studies indicated that cerebellar GABAbR may mediate multiple forms of inhibitory and excitatory modulation of cerebellar circuits. Nevertheless, the in vivo influence of cerebellar GABAbR activation is unclear. As the first step in addressing this issue, we examined how pharmacological activation of cerebellar GABAbR modulates optokinetic reflex (OKR), an involuntary cerebellum-dependent eye movement for stabilizing the retinal image against the drift of the visual scene. We injected baclofen, a GABAbR-selective agonist, or control saline into the cerebellar flocculi of adult mice and then performed 1-h OKR measurement sessions on two consecutive days. In the day 1 session, the baclofen (5 nM)-injected mice and control mice showed similar initial OKR gains and similar training-induced increases in the OKR gain (OKR adaptation). This result suggests that GABAbR activation does not affect cerebellar computation for executing OKR and formation of short-term memory for OKR adaptation. At the beginning of the day 2 session, the baclofen (5 nM or 50 µM)-injected mice showed an OKR gain higher than that achieved in the day 1 session while the control mice did not. This result suggests that GABAbR activation may facilitate the formation of OKR adaptation-related long-term memory. These findings provide a new insight into the functional architecture of the cerebellar circuits and indicate GABAbR to be a new target of pharmacological therapy against diseases with cerebellar dysfunction.