The AMP-activated protein kinase (AMPK) is a key cellular energy sensor and regulator of metabolic homeostasis. Activation of AMPK provides beneficial outcomes fighting against metabolic disorders such as insulin resistance and type 2 diabetes. Currently, there is no allosteric AMPK activator available for the treatment of metabolic diseases and limited compounds available to robustly stimulate cellular/tissue AMPK in a specific manner. Here, we investigated if simultaneous administration of two different pharmacological AMPK activators, which bind and act on different sites, would result in an additive or synergistic effect on AMPK and its downstream signaling and physiological events in intact cells. We observed that co-treating primary hepatocytes with an AMP mimetic AICAR and a low dose (1 μM) of an allosteric activator A769662 produces a synergistic effect on AMPK Thr172 phosphorylation and catalytic activity, which was associated with a more profound increase/decrease in phosphorylation of downstream AMPK targets and inhibition of hepatic lipogenesis compared to single compound treatment. Mechanistically, we found that co-treatment does not stimulate LKB1, upstream kinase for AMPK, but it protects against dephosphorylation of Thr172 phosphorylation by protein phosphatase, PP2Cα, in an additive manner in cell-free assay. Collectively, we demonstrate that AICAR sensitizes the effect of A769662 and promote AMPK activity and its downstream events. The study demonstrates the feasibility of promoting AMPK activity by using two activators with distinct modes of action in order to achieve a greater activation of AMPK and downstream signaling.