Toshio Ikeda

Causal relationship between the loss of RUNX3 expression and gastric cancer.

Runx3/Pebp2alphaC null mouse gastric mucosa exhibits hyperplasias due to stimulated proliferation and suppressed apoptosis in epithelial cells, and the cells are resistant to growth-inhibitory and apoptosis-inducing action of TGF-beta, indicating that Runx3 is a major growth regulator of gastric epithelial cells. Between 45% and 60% of human gastric cancer cells do not significantly express RUNX3 due to hemizygous deletion and hypermethylation of the RUNX3 promoter region. Tumorigenicity of human gastric cancer cell lines in nude mice was inversely related to their level of RUNX3 expression, and a mutation (R122C) occurring within the conserved Runt domain abolished the tumor-suppressive effect of RUNX3, suggesting that a lack of RUNX3 function is causally related to the genesis and progression of human gastric cancer.

Roles of continuous neurogenesis in the structural and functional integrity of the adult forebrain

Neurogenesis occurs continuously in the forebrain of adult mammals, but the functional importance of adult neurogenesis is still unclear. Here, using a genetic labeling method in adult mice, we found that continuous neurogenesis results in the replacement of the majority of granule neurons in the olfactory bulb and a substantial addition of granule neurons to the hippocampal dentate gyrus. Genetic ablation of newly formed neurons in adult mice led to a gradual decrease in the number of granule cells in the olfactory bulb, inhibition of increases in the granule cell number in the dentate gyrus and impairment of behaviors in contextual and spatial memory, which are known to depend on hippocampus. These results suggest that continuous neurogenesis is required for the maintenance and reorganization of the whole interneuron system in the olfactory bulb, the modulation and refinement of the existing neuronal circuits in the dentate gyrus and the normal behaviors involved in hippocampal-dependent memory.

Innate versus learned odour processing in the mouse olfactory bulb

The mammalian olfactory system mediates various responses, including aversive behaviours to spoiled foods and fear responses to predator odours. In the olfactory bulb, each glomerulus represents a single species of odorant receptor. Because a single odorant can interact with several different receptor species, the odour information received in the olfactory epithelium is converted to a topographical map of multiple glomeruli activated in distinct areas in the olfactory bulb. To study how the odour map is interpreted in the brain, we generated mutant mice in which olfactory sensory neurons in a specific area of the olfactory epithelium are ablated by targeted expression of the diphtheria toxin gene. Here we show that, in dorsal-zone-depleted mice, the dorsal domain of the olfactory bulb was devoid of glomerular structures, although second-order neurons were present in the vacant areas. The mutant mice lacked innate responses to aversive odorants, even though they were capable of detecting them and could be conditioned for aversion with the remaining glomeruli. These results indicate that, in mice, aversive information is received in the olfactory bulb by separate sets of glomeruli, those dedicated for innate and those for learned responses.

Deficient cerebellar long-term depression, impaired eyeblink conditioning, and normal motor coordination in GFAP mutant mice.

Mice devoid of glial fibrillary acidic protein (GFAP), an intermediate filament protein specifically expressed in astrocytes, develop normally and do not show any detectable abnormalities in the anatomy of the brain. In the cerebellum, excitatory synaptic transmission from parallel fibers (PFs) or climbing fibers (CFs) to Purkinje cells is unaltered, and these synapses display normal short-term synaptic plasticity to paired stimuli in GFAP mutant mice. In contrast, long-term depression (LTD) at PF-Purkinje cell synapses is clearly deficient. Furthermore, GFAP mutant mice exhibited a significant impairment of eyeblink conditioning without any detectable deficits in motor coordination tasks. These results suggest that GFAP is required for communications between Bergmann glia and Purkinje cells during LTD induction and maintenance. The data support the notion that cerebellar LTD is a cellular mechanism closely associated with eyeblink conditioning, but is not essential for motor coordination tasks tested.

Mice devoid of the glial fibrillary acidic protein develop normally and are susceptible to scrapie prions

Glial fibrillary acidic protein (GFAP) is an intermediate filament protein specifically expressed in astrocytes in the CNS. To examine the function of GFAP in vivo, the Gfap gene was disrupted by gene targeting in embryonic stem cells. Mice homozygous for the mutation were completely devoid of GFAP but exhibited normal development and showed no obvious anatomical abnormalities in the CNS. When inoculated with infectious scrapie prions, the mutant mice exhibited neuropathological changes typical of prion diseases. Infectious prions accumulated in brains of the mutant mice to a degree similar to that in control littermates. These results suggest that GFAP is not essential for the morphogenesis of the CNS or for astrocytic responses against neuronal injury. The results argue against the hypothesis that GFAP plays a crucial role in the pathogenesis of prion diseases.

Extracellular Signal-Regulated Kinase 2 (ERK2) Knockdown Mice Show Deficits in Long-Term Memory; ERK2 Has a Specific Function in Learning and Memory

The extracellular signal-regulated kinase (ERK) 1 and 2 are important signaling components implicated in learning and memory. These isoforms display a high degree of sequence homology and share a similar substrate profile. However, recent findings suggest that these isoforms may have distinct roles: whereas ERK1 seems to be not so important for associative learning, ERK2 might be critically involved in learning and memory. Thus, the individual role of ERK2 has received considerable attention, although it is yet to be understood. Here, we have generated a series of mice in which ERK2 expression decreased in an allele dose-dependent manner. Null ERK2 knock-out mice were embryonic lethal, and the heterozygous mice were anatomically impaired. To gain a better understanding of the influence of ERK2 on learning and memory, we also generated knockdown mice in which ERK2 expression was partially (20–40%) reduced. These mutant mice were viable and fertile with normal appearance. The mutant mice showed a deficit in long-term memory in classical fear conditioning, whereas short-term memory was normal. The mice also showed learning deficit in the water maze and the eight-arm radial maze. The ERK1 expression level of the knockdown mice was comparable with the wild-type control. Together, our results indicate a noncompensable role of ERK2-dependent signal transduction in learning and memory.

Complementary expression and neurite outgrowth activity of netrin-G subfamily members

Classical members of the UNC6/netrin family are secreted proteins which play a role as long-range cues for directing growth cones. We here identified in mice a novel member netrin-G2 which constitute a subfamily with netrin-G1 among the UNC6/netrin family. Both of these netrin-Gs are characterized by glycosyl phosphatidyl-inositol linkage onto cells, molecular variants presumably generated by alternative splicing and lack of any appreciable affinity to receptors for classical netrins. These genes are preferentially expressed in the central nervous system with complementary distribution in most brain areas, that is netrin-G1 in the dorsal thalamus, olfactory bulb and inferior colliculus, and netrin-G2 in the cerebral cortex, habenular nucleus and superior colliculus. Consistently, immunohistochemical analysis revealed that netrin-G1 molecules are present on thalamocortical but not corticothalamic axons. Thalamic and neocortical neurons extended long neurites on immobilized recombinant netrin-G1 or netrin-G2 in vitro. Immobilized anti-netrin-G1 antibodies altered shapes of cultured thalamic neurons. We propose that netrin-Gs provide short-range cues for axonal and/or dendritic behavior through bi-directional signaling.

ERK2 Contributes to the Control of Social Behaviors in Mice

Signaling through extracellular signal-regulated kinase (ERK) is important in multiple signal transduction networks in the CNS. However, the specific role of ERK2 in in vivo brain functions is not fully understood. Here we show that ERK2 play a critical role in regulating social behaviors as well as cognitive and emotional behaviors in mice. To study the brain function of ERK2, we used a conditional, region-specific, genetic approach to target Erk2 using the Cre/loxP strategy with a nestin promoter-driven cre transgenic mouse line to induce recombination in the CNS. The resulting Erk2 conditional knock-out (CKO) mice, in which Erk2 was abrogated specifically in the CNS, were viable and fertile with a normal appearance. These mice, however, exhibited marked anomalies in multiple aspects of social behaviors related to facets of autism-spectrum disorders: elevated aggressive behaviors, deficits in maternal nurturing, poor nest-building, and lower levels of social familiarity and social interaction. Erk2 CKO mice also exhibited decreased anxiety-related behaviors and impaired long-term memory. Pharmacological inhibition of ERK1 phosphorylation in Erk2 CKO mice did not affect the impairments in social behaviors and learning disabilities, indicating that ERK2, but not ERK1 plays a critical role in these behaviors. Our findings suggest that ERK2 has complex and multiple roles in the CNS, with important implications for human psychiatric disorders characterized by deficits in social behaviors.

Inducible cAMP Early Repressor Acts as a Negative Regulator for Kindling Epileptogenesis and Long-Term Fear Memory

Long-lasting neuronal plasticity as well as long-term memory (LTM) requires de novo synthesis of proteins through dynamic regulation of gene expression. cAMP-responsive element (CRE)-mediated gene transcription occurs in an activity-dependent manner and plays a pivotal role in neuronal plasticity and LTM in a variety of species. To study the physiological role of inducible cAMP early repressor (ICER), a CRE-mediated gene transcription repressor, in neuronal plasticity and LTM, we generated two types of ICER mutant mice: ICER-overexpressing (OE) mice and ICER-specific knock-out (KO) mice. Both ICER-OE and ICER-KO mice show no apparent abnormalities in their development and reproduction. A comprehensive battery of behavioral tests revealed no robust changes in locomotor activity, sensory and motor functions, and emotional responses in the mutant mice. However, long-term conditioned fear memory was attenuated in ICER-OE mice and enhanced in ICER-KO mice without concurrent changes in short-term fear memory. Furthermore, ICER-OE mice exhibited retardation of kindling development, whereas ICER-KO mice exhibited acceleration of kindling. These results strongly suggest that ICER negatively regulates the neuronal processes required for long-term fear memory and neuronal plasticity underlying kindling epileptogenesis, possibly through suppression of CRE-mediated gene transcription.

Dual involvement of G-substrate in motor learning revealed by gene deletion.

In this study, we generated mice lacking the gene for G-substrate, a specific substrate for cGMP-dependent protein kinase uniquely located in cerebellar Purkinje cells, and explored their specific functional deficits. G-substrate–deficient Purkinje cells in slices obtained at postnatal weeks (PWs) 10–15 maintained electrophysiological properties essentially similar to those from WT littermates. Conjunction of parallel fiber stimulation and depolarizing pulses induced long-term depression (LTD) normally. At younger ages, however, LTD attenuated temporarily at PW6 and recovered thereafter. In parallel with LTD, short-term (1 h) adaptation of optokinetic eye movement response (OKR) temporarily diminished at PW6. Young adult G-substrate knockout mice tested at PW12 exhibited no significant differences from their WT littermates in terms of brain structure, general behavior, locomotor behavior on a rotor rod or treadmill, eyeblink conditioning, dynamic characteristics of OKR, or short-term OKR adaptation. One unique change detected was a modest but significant attenuation in the long-term (5 days) adaptation of OKR. The present results support the concept that LTD is causal to short-term adaptation and reveal the dual functional involvement of G-substrate in neuronal mechanisms of the cerebellum for both short-term and long-term adaptation.