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Behavior training reverses asymmetry in hippocampal transcriptome of the cav3.2 knockout mice.

Chung NC, Huang YH, Chang CH, Liao JC, Yang CH, Chen CC, Liu IY - PLoS ONE (2015)

Bottom Line: We found a significant left-right asymmetric effect on the hippocampal transcriptome caused by the Cav3.2 knockout.Remarkably, the effect of Cav3.2 knockout was partially reversed by trace fear conditioning.To our knowledge, these results demonstrate for the first time the asymmetric effects of the Cav3.2 and its partial reversal by behavior training on the hippocampal transcriptome.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology and Human Genetics, Tzu Chi University, Hualien, Taiwan.

ABSTRACT
Homozygous Cav3.2 knockout mice, which are defective in the pore-forming subunit of a low voltage activated T-type calcium channel, have been documented to show impaired maintenance of late-phase long-term potentiation (L-LTP) and defective retrieval of context-associated fear memory. To investigate the role of Cav3.2 in global gene expression, we performed a microarray transcriptome study on the hippocampi of the Cav3.2-/- mice and their wild-type littermates, either naïve (untrained) or trace fear conditioned. We found a significant left-right asymmetric effect on the hippocampal transcriptome caused by the Cav3.2 knockout. Between the naive Cav3.2-/- and the naive wild-type mice, 3522 differentially expressed genes (DEGs) were found in the left hippocampus, but only 4 DEGs were found in the right hippocampus. Remarkably, the effect of Cav3.2 knockout was partially reversed by trace fear conditioning. The number of DEGs in the left hippocampus was reduced to 6 in the Cav3.2 knockout mice after trace fear conditioning, compared with the wild-type naïve mice. To our knowledge, these results demonstrate for the first time the asymmetric effects of the Cav3.2 and its partial reversal by behavior training on the hippocampal transcriptome.

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The overall experimental design.(A) Hippocampal sample groups. We used microarrays to probe the transcriptome of naïve (N) and trained (T) groups for both wild-type (W) and the Cav3.2 homozygous knockout (K) mice. In each group, we separated left (L) and right (R) hippocampi samples. A total of eight groups of samples were analyzed: WNL, WNR, WTL, WTR, KNL, KNR, KTL, and KTR. (B) Experimental procedures: Mice were handled in conditioning chambers for three days (Day 1–3), and then received trace fear conditioning individually (Day 4), tested with contextual memory 24 hours later (Day 5) in the same conditioning chamber. Immediately after memory testing, left (L) and right (R) hippocampi from three W and three K animals were dissected, pooled together to extract total RNA for microarray hybridization.
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pone.0118832.g001: The overall experimental design.(A) Hippocampal sample groups. We used microarrays to probe the transcriptome of naïve (N) and trained (T) groups for both wild-type (W) and the Cav3.2 homozygous knockout (K) mice. In each group, we separated left (L) and right (R) hippocampi samples. A total of eight groups of samples were analyzed: WNL, WNR, WTL, WTR, KNL, KNR, KTL, and KTR. (B) Experimental procedures: Mice were handled in conditioning chambers for three days (Day 1–3), and then received trace fear conditioning individually (Day 4), tested with contextual memory 24 hours later (Day 5) in the same conditioning chamber. Immediately after memory testing, left (L) and right (R) hippocampi from three W and three K animals were dissected, pooled together to extract total RNA for microarray hybridization.

Mentions: Bioconductor (www.bioconductor.org) and R (www.r-project.org) were used as computational tools for microarray analysis. Raw data were thresholded with a floor of 100 and normalized using the quantile normalization method. To identify differentially expressed genes (DEGs), the Significance Analysis of Microarrays (SAM) algorithm [25] and fold change (FC) were used for each of eight microarray comparisons (WNL vs KNL, WNR vs KNR, WTL vs KTL, WTR vs KTR, WNL vs WTL, KNL vs KTL, WNR vs WTR, KNR vs KTR, Fig. 1A). A gene was considered significantly differentially expressed if a false discovery rate (q) is less than 0.01 and FC is greater than 2. The DEGs identified in at least one of comparisons were clustered using a hierarchical clustering method. To gain insight into the biological processes involved, each cluster was then analyzed by DAVID functional annotation tool [26] to identify enriched KEGG pathways. In order to demonstrate the repeatability of the duplicated microarray raw datasets, we perform correlational analysis. S1 Table lists the designated groups of raw datasets. S1 and S2 Figs. and show the scatter plots of the raw datasets of the control and the training groups, respectively. Correlation analysis shows that for all groups R≥ 0.99, indicating that the consistency of the duplicate raw datasets is high.


Behavior training reverses asymmetry in hippocampal transcriptome of the cav3.2 knockout mice.

Chung NC, Huang YH, Chang CH, Liao JC, Yang CH, Chen CC, Liu IY - PLoS ONE (2015)

The overall experimental design.(A) Hippocampal sample groups. We used microarrays to probe the transcriptome of naïve (N) and trained (T) groups for both wild-type (W) and the Cav3.2 homozygous knockout (K) mice. In each group, we separated left (L) and right (R) hippocampi samples. A total of eight groups of samples were analyzed: WNL, WNR, WTL, WTR, KNL, KNR, KTL, and KTR. (B) Experimental procedures: Mice were handled in conditioning chambers for three days (Day 1–3), and then received trace fear conditioning individually (Day 4), tested with contextual memory 24 hours later (Day 5) in the same conditioning chamber. Immediately after memory testing, left (L) and right (R) hippocampi from three W and three K animals were dissected, pooled together to extract total RNA for microarray hybridization.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4358833&req=5

pone.0118832.g001: The overall experimental design.(A) Hippocampal sample groups. We used microarrays to probe the transcriptome of naïve (N) and trained (T) groups for both wild-type (W) and the Cav3.2 homozygous knockout (K) mice. In each group, we separated left (L) and right (R) hippocampi samples. A total of eight groups of samples were analyzed: WNL, WNR, WTL, WTR, KNL, KNR, KTL, and KTR. (B) Experimental procedures: Mice were handled in conditioning chambers for three days (Day 1–3), and then received trace fear conditioning individually (Day 4), tested with contextual memory 24 hours later (Day 5) in the same conditioning chamber. Immediately after memory testing, left (L) and right (R) hippocampi from three W and three K animals were dissected, pooled together to extract total RNA for microarray hybridization.
Mentions: Bioconductor (www.bioconductor.org) and R (www.r-project.org) were used as computational tools for microarray analysis. Raw data were thresholded with a floor of 100 and normalized using the quantile normalization method. To identify differentially expressed genes (DEGs), the Significance Analysis of Microarrays (SAM) algorithm [25] and fold change (FC) were used for each of eight microarray comparisons (WNL vs KNL, WNR vs KNR, WTL vs KTL, WTR vs KTR, WNL vs WTL, KNL vs KTL, WNR vs WTR, KNR vs KTR, Fig. 1A). A gene was considered significantly differentially expressed if a false discovery rate (q) is less than 0.01 and FC is greater than 2. The DEGs identified in at least one of comparisons were clustered using a hierarchical clustering method. To gain insight into the biological processes involved, each cluster was then analyzed by DAVID functional annotation tool [26] to identify enriched KEGG pathways. In order to demonstrate the repeatability of the duplicated microarray raw datasets, we perform correlational analysis. S1 Table lists the designated groups of raw datasets. S1 and S2 Figs. and show the scatter plots of the raw datasets of the control and the training groups, respectively. Correlation analysis shows that for all groups R≥ 0.99, indicating that the consistency of the duplicate raw datasets is high.

Bottom Line: We found a significant left-right asymmetric effect on the hippocampal transcriptome caused by the Cav3.2 knockout.Remarkably, the effect of Cav3.2 knockout was partially reversed by trace fear conditioning.To our knowledge, these results demonstrate for the first time the asymmetric effects of the Cav3.2 and its partial reversal by behavior training on the hippocampal transcriptome.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology and Human Genetics, Tzu Chi University, Hualien, Taiwan.

ABSTRACT
Homozygous Cav3.2 knockout mice, which are defective in the pore-forming subunit of a low voltage activated T-type calcium channel, have been documented to show impaired maintenance of late-phase long-term potentiation (L-LTP) and defective retrieval of context-associated fear memory. To investigate the role of Cav3.2 in global gene expression, we performed a microarray transcriptome study on the hippocampi of the Cav3.2-/- mice and their wild-type littermates, either naïve (untrained) or trace fear conditioned. We found a significant left-right asymmetric effect on the hippocampal transcriptome caused by the Cav3.2 knockout. Between the naive Cav3.2-/- and the naive wild-type mice, 3522 differentially expressed genes (DEGs) were found in the left hippocampus, but only 4 DEGs were found in the right hippocampus. Remarkably, the effect of Cav3.2 knockout was partially reversed by trace fear conditioning. The number of DEGs in the left hippocampus was reduced to 6 in the Cav3.2 knockout mice after trace fear conditioning, compared with the wild-type naïve mice. To our knowledge, these results demonstrate for the first time the asymmetric effects of the Cav3.2 and its partial reversal by behavior training on the hippocampal transcriptome.

Show MeSH