Previously we have reported a near-infrared optical response in the region occupied by a peripheral nerve that is distal to the site of electrical stimulation of that peripheral nerve. This "intermediate" signal is vascular in nature but its biological origin not been elucidated. In the present study, an animal model of the signal has been created and our human studies expanded to directly investigate the contribution of non-artifactual vascular motion induced by muscle contraction to the biological origin of this signal. Under non-invasive conditions during stimulation of the exposed sciatic nerve of the Sprague-Dawley rat, optical responses are robust. These signals can be abolished both pharmacologically and surgically using methods that eliminate muscle motion while leaving the electrophysiological health of the nerve intact. In human studies, signals that are elicited on stimulation of nerves containing motor axons, both within and outside the predicted imaging volume of the spectrometer, have similar temporal characteristics of those previously observed. Moreover, stimulation of sensory nerves alone does not elicit an optical response. These results strongly suggest that the intermediate signals are derived from stimulus-induced muscle contraction (whether via an innervating nerve or by direct stimulation) causing translational vascular motion within the optically interrogated region.