The MrD of the neostriatum is a flat, pan-shaped structure consisting of dorsoventral spindle-shaped neurons arranged in a parallel formation, located at the caudomedial margin of the neostriatum and surrounding the dostrolateral border of the globus pallidus (GP), and distinct from other parts of the striatum in the mammalian brains. Lesion and Y-maze tests and assessment of c-Fos expression and Patch clamp analysis showed that the MrD contributes to the learning and memory function[23–30]. FMRI investigations and clinical case reports verified the contribution of the MrD to digital working and mathematical calculating memory in the human brain[34–36]. All these early experiments indicate that MrD is likely to be an important subcortical center of learning and memory based on its position, its advanced development in higher mammalian brains, its abundant blood supply and the diverse connections with other memory-related structures. Moreover, miRNAs are particularly abundant in neurons, and together with the fact that a given miRNA usually regulates the expression of hundreds of target mRNAs, neuronal miRNA pathways create an extremely powerful mechanism for dynamically adjusting the protein content of neuronal compartments without the need for new gene transcription. MiRNAs likely have a big impact on higher cognitive function including learning and memory [10, 37, 38]. It has been speculated that miRNA expression differing from brain region to region, may reflect brain region-specific miRNA expression patterns with a corresponding role in each brain area . Recently, Juhila et al. compared miRNA expression between hippocampus and frontal cortex, and confirmed this hypothesis finding that miRNA expression in the hippocampus was extremely different from that of frontal cortex . Based on the above discoveries, this research was aimed at finding those miRNAs that were differentially expressed between MrD and Hip, the well established memory related structures in the brain.
Although miRNAs have been implicated in several important biological functions in the CNS including neurogenesis , dendrite formation , brain morphogenesis , neural plasticity , and silencing of non-neuronal transcripts [45, 46], far less information is available on miRNA expression patterns of neostriatum in anatomically regional differences. In this study, a miRNA array with 357 known rodent miRNA probes (15 replicates per array) was applied to sample the differential expression of miRNAs in Hip and the MrD of the rat brain. In the results, the most dysregulated 30 probes were identified in each rat. A total of 78 miRNAs were listed in Additional file 1: Table S3 and 11 miRNAs were differentially expressed in more than one rat (gray line in Additional file 1: Table S3). Although the most upregulated miRNAs in three rats were miR-383, miR-451 and miR-219-5p (with logs [FC]: -2.15, -2.21, -2.60) respectively, we wanted to focus on miRNAs that were altered in different rats simultaneously. Notably, miR-383 was not only most upregulated in rat1, but was also high in rat2 and rat3. The other ten miRNAs, up-regulated or down-regulated, were found to be high differentially affected in two different sample rats (Additional file 2: Table S4).
The importance of miRNAs in the nervous system was first described in Danio rerio (zebrafish) in which a mutation in dicer1 led to failure to produce mature miRNAs and resulted in gross morphological defects in the nervous system . Effects involving regulation of specific miRNAs in neurons have been found in a number of organisms including mammals. For example, during neurogenesis, the levels of both miR-124 and miR-9 are greatly increased, and both of them were indicated involving in neuronal differentiation in vitro experiments [44, 48]. Deep sequencing of miRNAs derived from tissues and cell lines have revealed these and other miRNAs are restricted to the CNS . Definitive proof of the role of miR-124 in neurogenesis has now been achieved in vivo, revealing its critical role in the differentiation of neurons from neural precursors . In addition to differentiation of neurons, miRNAs have been shown to affect crucial aspects of neurons. For example, neurite outgrowth is regulated by miR-132 . Furthermore, one crucial functional aspect of neurons, the synapse, is under miRNAs control. In the hippocampus, miR-134 regulates the size of dendritic spines, sites of synaptic transmission . Further linking of miRNAs to synaptic changes and the implications of such in brain development and plasticity was the recent demonstration that miR-138 controls dendritic spine morphogenesis . The expression of miRNAs mentioned above was not significantly deregulated in our study, though they were verified to be more important in development, differentiation and function of CNS.
Rno-miR-383, the most interesting one of these miRNAs, is significantly up-regulated in Hip in all three rats. Its precursor possesses 73nt stem loop construction, and the mature sequence is 5′-cagaucagaaggugacugugg-3′. After bioinformatics analysis, there are 770 predicted target sites in rattus norvegicu genome (http://mirnamap.mbc.nctu.edu.tw/index.php). There are 17 members from different species that have been sequenced in the mir-383 family, including: bta, cfa, eca, gga, hsa, mdo, mml, mmu, oan, ppy, ptr, rno, ssc, tgu, xtr, aca, sha (http://www.mirbase.org/cgi-bin/mirna_summary.pl?fam=MIPF0000137). Experimentally, Lian et al  have reported that the expression of hsa-miR-383 is altered in testicular itssues of human patients, and they consider that a potential target of miR-383 may be GADD45G. GADD45G can induce apoptosis and inhibit cell growth in response to stress shock. Abnormal expressions of these proteins may have a significant impact on male infertility [53, 54]. However, no evidence demonstrates how rno-miR-383 is involved into complicated regulatory net-works in CNS, especially in MrD.
Beside rno-miR-383, there are also 10 miRNAs differentially expressed in only two rats in this study. In these 10 deregulated miRNAs, seven (let-7d*, miR-181b, miR-187, miR-214, miR-466b, miR-592, miR-1224) are up-regulated in two rats, and one (miR-195) is down-regulated. The last two investigated, miR-382 and miR-411, were up-regulated in one rat and down-regulated in another. It is possible to speculate that non tissue-specific expression may be a reason for this unexpected expression pattern of some miRNAs, and meanwhile they were also expressed in many different tissues in random manner. The other reasons may be the intrinsic variability of these miRNAs in this brain region or the effect correlate with some behavioral variability. Except for let-7d*, the RT-PCR results showed the same trend with that of microarray, which means that it validated the expression patterns we found in our microarray experiments.
Our previous study has shown that the MrD-NBM-hippocampus circuit may play an important role in modulating learning and memory [23–30]. Tissue-specific miRNAs expression patterns as determined in this study can make important contributions to understanding regulatory expression networks. Although difficult for small RNAs, there is no substitute for in situ hybridization studies to follow up on various experimental findings in order to assess the regions and cell types responsible for distinct miRNA expression patterns. Meanwhile, differential expression of identified miRNAs can be then assessed in models of disease or in diseased tissue itself. In summary, the study on miRNAs, their mRNA targets, and resulting changes in protein products will continue to be an exciting field of research, leading to a greater understanding of the regulatory effects of the miRNA.