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Alpha-CaMKII deficiency causes immature dentate gyrus, a novel candidate endophenotype of psychiatric disorders.

Yamasaki N, Maekawa M, Kobayashi K, Kajii Y, Maeda J, Soma M, Takao K, Tanda K, Ohira K, Toyama K, Kanzaki K, Fukunaga K, Sudo Y, Ichinose H, Ikeda M, Iwata N, Ozaki N, Suzuki H, Higuchi M, Suhara T, Yuasa S, Miyakawa T - Mol Brain (2008)

Bottom Line: Transcriptome analysis of the hippocampus of these mutants revealed that the expression levels of more than 2000 genes were significantly changed.Moreover, c-Fos expression in the DG after electric footshock was almost completely and selectively abolished in the mutants.Nearly half of the differentially-expressed probes in the schizophrenia-enriched cluster encoded genes that are involved in neurogenesis or in neuronal migration/maturation, including calbindin, a marker for mature DG neurons.

View Article: PubMed Central - HTML - PubMed

Affiliation: Frontier Technology Center, Kyoto University Graduate School of Medicine, Japan. yamasaki@kuhp.kyoto-u.ac.jp

ABSTRACT
Elucidating the neural and genetic factors underlying psychiatric illness is hampered by current methods of clinical diagnosis. The identification and investigation of clinical endophenotypes may be one solution, but represents a considerable challenge in human subjects. Here we report that mice heterozygous for a mutation of the alpha-isoform of calcium/calmodulin-dependent protein kinase II (alpha-CaMKII+/-) have profoundly dysregulated behaviours and impaired neuronal development in the dentate gyrus (DG). The behavioral abnormalities include a severe working memory deficit and an exaggerated infradian rhythm, which are similar to symptoms seen in schizophrenia, bipolar mood disorder and other psychiatric disorders. Transcriptome analysis of the hippocampus of these mutants revealed that the expression levels of more than 2000 genes were significantly changed. Strikingly, among the 20 most downregulated genes, 5 had highly selective expression in the DG. Whereas BrdU incorporated cells in the mutant mouse DG was increased by more than 50 percent, the number of mature neurons in the DG was dramatically decreased. Morphological and physiological features of the DG neurons in the mutants were strikingly similar to those of immature DG neurons in normal rodents. Moreover, c-Fos expression in the DG after electric footshock was almost completely and selectively abolished in the mutants. Statistical clustering of human post-mortem brains using 10 genes differentially-expressed in the mutant mice were used to classify individuals into two clusters, one of which contained 16 of 18 schizophrenic patients. Nearly half of the differentially-expressed probes in the schizophrenia-enriched cluster encoded genes that are involved in neurogenesis or in neuronal migration/maturation, including calbindin, a marker for mature DG neurons. Based on these results, we propose that an "immature DG" in adulthood might induce alterations in behavior and serve as a promising candidate endophenotype of schizophrenia and other human psychiatric disorders.

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Altered Granule Cell Excitability and Mossy Fiber Synaptic Transmission in Alpha-CaMKII+/- Mice. (A) Examples of action potential firing of granule cells in wild-type and mutant mice evoked by steady depolarizing currents (320 pA, 400 ms). (B) Increased input resistance in mutant mice (P = 0.0029). (C) Reduced spike latency in mutant mice (P < 0.0001). Depolarizing currents (320 pA) were injected into granule cells and the latency of the first action potential was measured. (D) The maximal number of spikes evoked by 400 ms depolarizing currents was decreased in mutant mice (P < 0.0001). (E) The efficacy of basal transmission at the mossy fiber synapse was increased in mutant mice (P < 0.0001). The ratio of the peak EPSP amplitude to fiber volley amplitude is shown. (F) Reduced paired-pulse facilitation ratios of EPSPs at intervals ranging from 50 to 5000 ms in mutant mice. (G) Mutant mice had greatly reduced frequency facilitation at 1 Hz. Sample EPSPs were recorded at the time indicated by the numbers in the graph. Error bars indicate s.e.m.
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Figure 4: Altered Granule Cell Excitability and Mossy Fiber Synaptic Transmission in Alpha-CaMKII+/- Mice. (A) Examples of action potential firing of granule cells in wild-type and mutant mice evoked by steady depolarizing currents (320 pA, 400 ms). (B) Increased input resistance in mutant mice (P = 0.0029). (C) Reduced spike latency in mutant mice (P < 0.0001). Depolarizing currents (320 pA) were injected into granule cells and the latency of the first action potential was measured. (D) The maximal number of spikes evoked by 400 ms depolarizing currents was decreased in mutant mice (P < 0.0001). (E) The efficacy of basal transmission at the mossy fiber synapse was increased in mutant mice (P < 0.0001). The ratio of the peak EPSP amplitude to fiber volley amplitude is shown. (F) Reduced paired-pulse facilitation ratios of EPSPs at intervals ranging from 50 to 5000 ms in mutant mice. (G) Mutant mice had greatly reduced frequency facilitation at 1 Hz. Sample EPSPs were recorded at the time indicated by the numbers in the graph. Error bars indicate s.e.m.

Mentions: There are distinct stages in adult neurogenesis in the DG. Each stage has a few markers and specific morphological and physiological properties [24,26]. The number of cells expressing Polysialic acid-NCAM (PSA-NCAM), a marker for late-stage progenitor cells and immature neurons, and calretinin, a marker for immature neurons, was dramatically increased in the DG of the mutant mice. In normal mice, after granule cells migrate into the granule cell layer, PSA-NCAM positive cells were hardly detected in the granule cell layer. In the mutants, however, there were many PSA-NCAM positive cells in the granule cell layer. Moreover, the amount of calbindin, a marker for mature neurons in the DG, -positive cells was dramatically reduced (Figure 3B and 3C). Thus, in the mutants, the number of immature neurons is increased and the number of mature neurons is decreased (in this report, "immature" means that the control of the precursor cell proliferation and the functional differentiation of the newly born cells were hampered.). Furthermore, rapid Golgi staining in the mutant DG showed that dendritic branching and length were decreased, which are the morphological properties of immature neurons (Figure 3D and Additional file 1, Figure S9). Consistent with these immunohistochemical and morphological findings, the neurons in mutant DG had electrophysiological features that are characteristic of immature DG neurons [27-29], including high input resistance, high excitability, small spike amplitude and a decreased number of spikes during sustained depolarization (Figure 4A–4D, and Additional file 1, Figure S10). Moreover, the mutants had extremely abnormal transmission at the synapses between granule cell axons, mossy fibers (MFs), and the CA3 pyramidal cells. That is, basal transmission was increased and large facilitation characteristic of the MF synapse was markedly decreased in mutants (Figure 4E–4G, and Additional file 1, Figure S10). The synaptic facilitation is generally known to be mediated by presynaptic mechanisms. Since the frequency facilitation at the MF synapse is very small at the early postnatal developmental stages in normal mice [30], the greatly reduced MF facilitation in the mutant mice lends further support to the hypothesis that the mutant granule cells in alpha-CaMKII+/- mice are immature. At the ultra structural level, the MF-CA3 synapses were poorly developed in the hippocampus of the mutant mice (see Additional file 1, Figure S11), which could be related to the abnormal transmission at the synapse. Collectively, results of analyses using molecular markers, morphological assessment using Golgi staining, and electrophysiological analysis using whole cell patch clamp all indicated that the vast majority of the mutant DG granule cells failed to develop into mature neurons. The qualitative difference in the DG granule cell developmental stage accompanied by the severely impaired neuronal functions might also account for the high number of genes that were differentially expressed in the mutant hippocampus.


Alpha-CaMKII deficiency causes immature dentate gyrus, a novel candidate endophenotype of psychiatric disorders.

Yamasaki N, Maekawa M, Kobayashi K, Kajii Y, Maeda J, Soma M, Takao K, Tanda K, Ohira K, Toyama K, Kanzaki K, Fukunaga K, Sudo Y, Ichinose H, Ikeda M, Iwata N, Ozaki N, Suzuki H, Higuchi M, Suhara T, Yuasa S, Miyakawa T - Mol Brain (2008)

Altered Granule Cell Excitability and Mossy Fiber Synaptic Transmission in Alpha-CaMKII+/- Mice. (A) Examples of action potential firing of granule cells in wild-type and mutant mice evoked by steady depolarizing currents (320 pA, 400 ms). (B) Increased input resistance in mutant mice (P = 0.0029). (C) Reduced spike latency in mutant mice (P < 0.0001). Depolarizing currents (320 pA) were injected into granule cells and the latency of the first action potential was measured. (D) The maximal number of spikes evoked by 400 ms depolarizing currents was decreased in mutant mice (P < 0.0001). (E) The efficacy of basal transmission at the mossy fiber synapse was increased in mutant mice (P < 0.0001). The ratio of the peak EPSP amplitude to fiber volley amplitude is shown. (F) Reduced paired-pulse facilitation ratios of EPSPs at intervals ranging from 50 to 5000 ms in mutant mice. (G) Mutant mice had greatly reduced frequency facilitation at 1 Hz. Sample EPSPs were recorded at the time indicated by the numbers in the graph. Error bars indicate s.e.m.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2562999&req=5

Figure 4: Altered Granule Cell Excitability and Mossy Fiber Synaptic Transmission in Alpha-CaMKII+/- Mice. (A) Examples of action potential firing of granule cells in wild-type and mutant mice evoked by steady depolarizing currents (320 pA, 400 ms). (B) Increased input resistance in mutant mice (P = 0.0029). (C) Reduced spike latency in mutant mice (P < 0.0001). Depolarizing currents (320 pA) were injected into granule cells and the latency of the first action potential was measured. (D) The maximal number of spikes evoked by 400 ms depolarizing currents was decreased in mutant mice (P < 0.0001). (E) The efficacy of basal transmission at the mossy fiber synapse was increased in mutant mice (P < 0.0001). The ratio of the peak EPSP amplitude to fiber volley amplitude is shown. (F) Reduced paired-pulse facilitation ratios of EPSPs at intervals ranging from 50 to 5000 ms in mutant mice. (G) Mutant mice had greatly reduced frequency facilitation at 1 Hz. Sample EPSPs were recorded at the time indicated by the numbers in the graph. Error bars indicate s.e.m.
Mentions: There are distinct stages in adult neurogenesis in the DG. Each stage has a few markers and specific morphological and physiological properties [24,26]. The number of cells expressing Polysialic acid-NCAM (PSA-NCAM), a marker for late-stage progenitor cells and immature neurons, and calretinin, a marker for immature neurons, was dramatically increased in the DG of the mutant mice. In normal mice, after granule cells migrate into the granule cell layer, PSA-NCAM positive cells were hardly detected in the granule cell layer. In the mutants, however, there were many PSA-NCAM positive cells in the granule cell layer. Moreover, the amount of calbindin, a marker for mature neurons in the DG, -positive cells was dramatically reduced (Figure 3B and 3C). Thus, in the mutants, the number of immature neurons is increased and the number of mature neurons is decreased (in this report, "immature" means that the control of the precursor cell proliferation and the functional differentiation of the newly born cells were hampered.). Furthermore, rapid Golgi staining in the mutant DG showed that dendritic branching and length were decreased, which are the morphological properties of immature neurons (Figure 3D and Additional file 1, Figure S9). Consistent with these immunohistochemical and morphological findings, the neurons in mutant DG had electrophysiological features that are characteristic of immature DG neurons [27-29], including high input resistance, high excitability, small spike amplitude and a decreased number of spikes during sustained depolarization (Figure 4A–4D, and Additional file 1, Figure S10). Moreover, the mutants had extremely abnormal transmission at the synapses between granule cell axons, mossy fibers (MFs), and the CA3 pyramidal cells. That is, basal transmission was increased and large facilitation characteristic of the MF synapse was markedly decreased in mutants (Figure 4E–4G, and Additional file 1, Figure S10). The synaptic facilitation is generally known to be mediated by presynaptic mechanisms. Since the frequency facilitation at the MF synapse is very small at the early postnatal developmental stages in normal mice [30], the greatly reduced MF facilitation in the mutant mice lends further support to the hypothesis that the mutant granule cells in alpha-CaMKII+/- mice are immature. At the ultra structural level, the MF-CA3 synapses were poorly developed in the hippocampus of the mutant mice (see Additional file 1, Figure S11), which could be related to the abnormal transmission at the synapse. Collectively, results of analyses using molecular markers, morphological assessment using Golgi staining, and electrophysiological analysis using whole cell patch clamp all indicated that the vast majority of the mutant DG granule cells failed to develop into mature neurons. The qualitative difference in the DG granule cell developmental stage accompanied by the severely impaired neuronal functions might also account for the high number of genes that were differentially expressed in the mutant hippocampus.

Bottom Line: Transcriptome analysis of the hippocampus of these mutants revealed that the expression levels of more than 2000 genes were significantly changed.Moreover, c-Fos expression in the DG after electric footshock was almost completely and selectively abolished in the mutants.Nearly half of the differentially-expressed probes in the schizophrenia-enriched cluster encoded genes that are involved in neurogenesis or in neuronal migration/maturation, including calbindin, a marker for mature DG neurons.

View Article: PubMed Central - HTML - PubMed

Affiliation: Frontier Technology Center, Kyoto University Graduate School of Medicine, Japan. yamasaki@kuhp.kyoto-u.ac.jp

ABSTRACT
Elucidating the neural and genetic factors underlying psychiatric illness is hampered by current methods of clinical diagnosis. The identification and investigation of clinical endophenotypes may be one solution, but represents a considerable challenge in human subjects. Here we report that mice heterozygous for a mutation of the alpha-isoform of calcium/calmodulin-dependent protein kinase II (alpha-CaMKII+/-) have profoundly dysregulated behaviours and impaired neuronal development in the dentate gyrus (DG). The behavioral abnormalities include a severe working memory deficit and an exaggerated infradian rhythm, which are similar to symptoms seen in schizophrenia, bipolar mood disorder and other psychiatric disorders. Transcriptome analysis of the hippocampus of these mutants revealed that the expression levels of more than 2000 genes were significantly changed. Strikingly, among the 20 most downregulated genes, 5 had highly selective expression in the DG. Whereas BrdU incorporated cells in the mutant mouse DG was increased by more than 50 percent, the number of mature neurons in the DG was dramatically decreased. Morphological and physiological features of the DG neurons in the mutants were strikingly similar to those of immature DG neurons in normal rodents. Moreover, c-Fos expression in the DG after electric footshock was almost completely and selectively abolished in the mutants. Statistical clustering of human post-mortem brains using 10 genes differentially-expressed in the mutant mice were used to classify individuals into two clusters, one of which contained 16 of 18 schizophrenic patients. Nearly half of the differentially-expressed probes in the schizophrenia-enriched cluster encoded genes that are involved in neurogenesis or in neuronal migration/maturation, including calbindin, a marker for mature DG neurons. Based on these results, we propose that an "immature DG" in adulthood might induce alterations in behavior and serve as a promising candidate endophenotype of schizophrenia and other human psychiatric disorders.

Show MeSH
Related in: MedlinePlus