Yes-associated protein 1 (YAP), a major transcriptional cofactor in the Hippo signaling pathway, is known to regulate cell growth and homeostasis. We have shown previously that YAP is activated and mediates adaptive cardiac hypertrophy in response to acute pressure overload (PO). However, how YAP induces adaptive hypertrophy is unknown. Glycolysis is intimately involved in cell growth, including cardiac hypertrophy. Here we examined whether YAP regulates glycolysis during acute PO to promote adaptive cardiac hypertrophy. Evaluation of extracellular acidification rate using the Seahorse XF analyzer in freshly isolated adult ventricular myocytes (AVMs) from wild-type mice revealed that glycolytic flux was increased by acute PO. The PO-induced glycolysis was attenuated in isolated AVMs from cardiac-specific heterozygous YAP knockout mice. We found that YAP promoted glycolysis by upregulating glucose transporter 1 (GLUT1), which in turn caused accumulation of glucose metabolites, including L-serine, L-aspartate, and malate, during acute PO. YAP overexpression increased GLUT1 protein levels, glycolytic flux, and cardiomyocytes size in isolated AVMs. These results suggest that YAP induces adaptive cardiac hypertrophy through activation of aerobic glycolysis in response to acute PO.