The ability to communicate through vocalization plays a key role in the survival of animals across all vertebrate groups. Although avian reptiles have received much attention relating to their stunning sound repertoire, non-avian reptiles have been wrongfully assumed to have less elaborate vocalization types, and little is known about the biomechanics of sound production and their underlying neural pathways in this group. We investigated alarm calls of Gekko gecko using audio and cineradiographic recordings. Acoustic analysis revealed three distinct call types: a sinusoidal call type (type 1); a train-like call type, characterized by distinct pulse trains (type 3); and an intermediate type, which showed both sinusoidal and pulse train components (type 2). Kinematic analysis of cineradiographic recordings showed that laryngeal movements differ significantly between respiratory and vocal behavior. During respiration, animals repeatedly moved their jaws to partially open their mouths, which was accompanied by small glottal movements. During vocalization, the glottis was pulled back, contrasting with what has previously been reported. In vitro retrograde tracing of the nerve innervating the laryngeal constrictor and dilator muscles revealed round to fusiform motoneurons in the hindbrain-spinal cord transition ipsilateral to the labeled nerve. Taken together, our observations provide insight into the alarm calls generated by G. gecko, the biomechanics of this sound generation and the underlying organization of motoneurons involved in the generation of vocalizations. Our observations suggest that G. gecko may be an excellent non-avian reptile model organism for enhancing our understanding of the evolution of vertebrate vocalization.