Mechanosensitive ensembles of neurons in insects, known as chordotonal organs (COs), function in proprioception, the detection of sound and substrate vibrations. Here, we characterized the mechanical sensitivity of the lateral pentascolopidial CO (lch5) of Drosophila melanogaster larvae to establish its postulated role in proprioception. We developed a physiologically realistic method to replicate proprioceptive input to lch5 by pulling the apodeme (tendon) to which the tips of the neurons attach. We found that lch5 sensory neurons respond transiently with a short latency to the velocity component of stretch displacements and the release of stretch (relaxation). In the mechanosensory mutant inactive, lch5 has a decreased response to mechanical stimuli and a lower overall spontaneous spike rate. Finally, we simulated the input that lch5 receives during crawling and observed spike rate changes of peristaltic body contraction. We provide a characterization of proprioceptive feedback in D. melanogaster larvae and firmly establish the proprioceptive function of lch5 in larval locomotion.