Angiotensin II plays a key role in the development of cardiac hypertrophy. The contribution of the angiotensin II type 1 receptor (AT1) in angiotensin II-induced cardiac hypertrophy is well established, but the role of AT2 signaling remains controversial. Previously, we have shown that natriuretic peptide receptor/guanylyl cyclase-A (GCA) signaling protects the heart from hypertrophy at least in part by inhibiting AT1-mediated pro-hypertrophic signaling. Here, we investigated the role of AT2 in cardiac hypertrophy observed in mice lacking GCA. Real-time RT-PCR and immunoblotting approaches indicated that the cardiac AT2 gene was overexpressed in GCA-deficient mice. Mice lacking AT2 alone did not exhibit an abnormal cardiac phenotype. In contrast, GCA-deficiency-induced increases in heart to body weight ratio, cardiomyocyte cross-sectional area, and collagen accumulation as evidenced by van Gieson staining were attenuated when AT2 was absent. Furthermore, the up-regulated cardiac expression of hypertrophy-related genes in GCA-null animals was also suppressed. Pharmacological blockade of AT2 with PD123319 similarly attenuated cardiac hypertrophy in GCA-deficient mice. In addition, whereas the AT1 antagonist olmesartan attenuated cardiac hypertrophy in GCA-deficient mice, this treatment was without effect on cardiac hypertrophy in GCA/AT2-double null mice, notwithstanding its potent antihypertensive effect in these animals. These results suggest that the interplay of AT2 and AT1 may be important in the development of cardiac hypertrophy. Collectively, our findings support the assertion that GCA inhibits AT2-mediated pro-hypertrophic signaling in heart and offer new insights into endogenous cardioprotective mechanisms during disease pathogenesis.