Olfactory discrimination training up-regulates and reorganizes expression of microRNAs in adult mouse hippocampus

Abstract

Adult male mice (strain C57Bl/6J) were trained to execute nose-poke responses for water reinforcement; then they were randomly assigned to either of two groups: olfactory discrimination training (exposed to two odours with reward contingent upon correctly responding to one odour) or pseudo-training (exposed to two odours with reward not contingent upon response). These were run in yoked fashion and killed when the discrimination-trained mouse reached a learning criterion of 70% correct responses in 20 trials, occurring after three sessions (a total of approximately 40 min of training). The hippocampus was dissected bilaterally from each mouse (N = 7 in each group) and profiling of 585 miRNAs (microRNAs) was carried out using multiplex RT-PCR (reverse transcription-PCR) plates. A significant global up-regulation of miRNA expression was observed in the discrimination training versus pseudo-training comparison; when tested individually, 29 miRNAs achieved significance at P = 0.05. miR-10a showed a 2.7-fold increase with training, and is predicted to target several learning-related mRNAs including BDNF (brain-derived neurotrophic factor), CAMK2b (calcium/calmodulin-dependent protein kinase IIβ), CREB1 (cAMP-response-element-binding protein 1) and ELAVL2 [ELAV (embryonic lethal, abnormal vision, Drosophila)-like; Hu B]. Analysis of miRNA pairwise correlations revealed the existence of several miRNA co-expression modules that were specific to the training group. These in vivo results indicate that significant, dynamic and co-ordinated changes in miRNA expression accompany early stages of learning.

Keywords: BDNF, brain-derived neurotrophic factor; Ct, threshold cycle value; DHPG, (S)-3,5-dihydroxyphenylglycine; LTP, long-term potentiation; MEF2, myocyte enhancer factor-2; NMDA, N-methyl-d-aspartate; RISC, RNA-induced silencing complex; RT-PCR, reverse transcription-PCR; TLDA, TaqMan® Low Density Arrays; TOP, terminal oligopyrimidine; dicer; learning; miRNA, microRNA; microRNA; olfactory discrimination; pre-miR, miRNA small hairpin precursor; pri-miR, primary miRNA gene transcript; snoRNA, small nucleolar RNA; synaptic plasticity.

Figure 1. miRNAs that are positively and significantly correlated with at least two other miRNAs within the training group, but are not correlated with each other in the control groups

See the Results section for details. Three distinct modules are formed; the central module has a particularly densely interconnected core, having two hubs (mir-188-5p and 489) surrounded by 15 miRNAs that are correlated with both hubs.