In ants, chemical communication, through pheromones for example, was thought to be the main communication tool in ant societies; however, a major question is whether chemical substances alone can control highly differentiated social behaviors. In this study, we focused on the surface-borne vibrational signals of fungus-growing ants, which are key organisms in the Neotropical ecosystem, and conducted detailed analyses of the signals, auditory organs, and playback experiments in Panama. Our results indicate that vibrational communication is a much more frequent and critical factor in fungus-growing ants than previously thought. The frequency of vibrational signal recordings from the ants was significantly correlated with social complexity. Structural analysis of the stridulatory organs indicated that both the area and number of slits were significantly correlated with body size. The ability of leaf-cutting ants to maintain fungus gardens was significantly lower in the vibration-inhibited group than in the pheromone secretion-inhibited group. We used fluorescent dyes to label sensory nerves and search for candidate sensory organs in a leaf-cutting ant. The candidate sensory organs were the Johnston’s organ in the pedicel of the antenna, the subgenual organ in the tibia, and the chordotonal organ in the femur. As a result, we could observe several anatomical features that could detect airborne sound in the subgenual organ and Johnston’s organ. This is the first time in the world that a detailed structure of the auditory organs of ants has been analyzed. The playback experiment involved transmitting artificially synthesized signals at 100, 400, and 1000 Hz through a bone-conduction earphone to a platform placed on the trail of leaf-cutting ants, and recording the responses on a video camera. As a result, the most active reaction was observed at 100 Hz and a significant difference was found among signals. These results suggest that ants that have become more communicative may induce altruistic behaviors and create more complex societies. The findings of this study may apply not only to social evolution studies but also to the effective control of ant behaviors.