This paper proposes a new testing method based on the toggle effect under transverse loads (cranial-caudal) to investigate the loosening potential of pedicular screw designs. A three-step in vitro testing procedure was developed to mimic the loosening mechanism of pedicular screws. Firstly, the pedicular screw of a certain design is inserted into a bone substitute model specifically designed for the test. Secondly, a controlled cyclic cranial-caudal loading is applied transversally to the longitudinal axis of the screw for three ascendent load levels (staircase) by a pre-determined number of load cycles. Lastly, each pedicular screw is adjusted and submitted to axial pull-out quasi-static testing. The results are used to calculate a loosening index that, together with statistical analysis, indicates the potential for loosening of the specific design evaluated. The proposed testing method effectively provides a simulated environment to evaluate the loosening potential of pedicular screw designs. The proposed loosening index calculation may be used to compare different pedicular screw designs. The proposed methodology was verified as a valuable tool to investigate the influence of the cranial-caudal loads on pedicular screw behavior. It offers a new alternative for use in pre-clinical studies on the loosening potential of pedicular screw designs.