Hypertension is the most important risk factor for cardiovascular diseases worldwide. However, the underlying molecular mechanisms of hypertension are complex and remain largely elusive. Here, we described a novel, microRNA-dependent therapeutic strategy for hypertension. First, we found that plasma microRNA-21-3p (miR-21-3p) levels were significantly reduced both in hypertensive patients and spontaneously hypertensive rats (SHRs) when compared with normal controls. In a series of experiments to dissect the role of miR-21-3p in hypertension, we showed that intravenous delivery of recombinant adeno-associated virus (rAAV)-mediated miR-21-3p expression induced a persistent attenuation of hypertension, with marked amelioration of target organ damages, including cardiac hypertrophy and fibrosis and artery and kidney fibrosis in SHRs, whereas miR-21-3p tough decoys (TuDs) counter-acted the above effects. Computational prediction coupled with biochemical experiments revealed that the miR-21-3p-mediated hypotensive reduction effect was accomplished by regulating phenotypic switch of vascular smooth muscle cells (VSMCs) via suppression of the adrenal alpha 2B-adrenergic receptor (ADRA2B) in arteries. Furthermore, we observed that activation of transcription factor NF-kappa B and SRF significantly increased the expression of miR-21-3p in VSMCs. In summary, our study is the first to identify a novel role and mechanism of miR-21-3p in blood pressure control and provides a possible strategy for hypertension therapy using rAAV-miR-21-3p.