Fig. 4

Lentiviral overexpression of miR-504 in cultured neurons, and expression analysis of miR-504 in human and mouse brain. a Representative images of live neurons infected with lentivirus expressing TurboGFP together with miR-504. Scale bar, 100 μm. b Representative western blot images and quantification showing that lentiviral overexpression of miR-504 decreased endogenous Shank3, but not Shank2 and PSD-95, proteins in cultured hippocampal neurons. c Box plots showing the expression distribution of miR-504 in 16 human brain regions. The black line in each box indicates the median value. The lower and upper hinges of each box indicate the lower and upper quartile values, respectively. The whiskers of each box indicate the most extreme data values within 1.5 times of the interquartile range. The open circles indicate the data values beyond the whisker limits. The number in parentheses associated with each brain region is the expression rank of miR-504 among all the 1,883 miRNAs available for the brain region. OFC, orbital prefrontal cortex; DFC, dorsolateral prefrontal cortex; VFC, ventrolateral prefrontal cortex; MFC, medial prefrontal cortex; M1C, primary motor (M1) cortex; S1C, primary somatosensory (S1) cortex; IPC, posterior inferior parietal cortex; A1C, primary auditory (A1) cortex; STC, superior temporal cortex; ITC, inferior temporal cortex; V1C, primary visual (V1) cortex; HIP, hippocampus; AMY, amygdala; STR, striatum; MD, mediodorsal nucleus of the thalamus; CBC, cerebellar cortex. d In situ hybridization of miR-504 in mouse brains. miR-504 signals were detected in neurons of cortex and hippocampus, and in Purkinje cells of cerebellum from 3- and 8-week-old mice. The probe against scrambled miRNA (miR-SCR) was used as a negative control. CB, cerebellum; CTX, cortex; DG, dentate gyrus; GL, granular layer; ML, molecular layer; PL, Purkinje cell layer. Scale bar, 500 μm. All data are presented as mean ± SEM. Statistical analyses are in Additional file 1: Table S3