Figure 3. Validation of top miR-132-upregulating candidate compounds in rodent and human neurons.

a, Top compounds that showed significant upregulation of miR-132 in primary rat cortical neurons and human NGN2-iNs after 24h treatment (RT-qPCR analysis, N=4). b, Dose curve experiments were performed in DIV14 rat neurons after 24h treatment. Solid lines were used for cardiac glycosides, and dotted lines were used for other compounds. EC₅₀ and max fold change were calculated using sigmoidal fit, 4 parameters. (N=4-6, error bars represent SD). c, Cardiac glycosides, forskolin, and BIX02188 upregulated the precursors of miR-132/212 24h after treatment (unpaired two-tailed Student’s t-test compared to DMSO control, N=4). d, Upregulation of miR-132 by forskolin or oleandrin was completely blocked by the transcription inhibitor actinomycin D and partially blocked by CREB inhibitor (unpaired two-tailed Student’s t-test compared to DMSO control, N=8-19). e, Knocking down ATP1A1 or ATP1A3, the predominant isoforms in neurons, also upregulated pre- and mature miR-132 (unpaired two-tailed Student’s t-test compared to DMSO control, N=4-6).