Objective The purpose of this study was to determine how the preload that precedes a high-velocity, low-amplitude spinal manipulation (HVLA-SM) affects muscle spindle input from lumbar paraspinal muscles both during and after the HVLA-SM. Methods Primary afferent activity from muscle spindles in lumbar paraspinal muscles were recorded from the L6 dorsal root in anesthetized cats. High-velocity, low-amplitude spinal manipulation of the L6 vertebra was preceded either by no preload or systematic changes in the preload magnitude, duration, and the presence or absence of a downward incisural point. Immediate effects of preload on muscle spindle responses to the HVLA-SM were determined by comparing mean instantaneous discharge frequencies (MIF) during the HVLA-SM's thrust phase with baseline. Longer lasting effects of preload on spindle responses to the HVLA-SM were determined by comparing MIF during slow ramp and hold movement of the L6 vertebra before and after the HVLA-SM. Results The smaller compared with the larger preload magnitude and the longer compared with the shorter preload duration significantly increased (P =.02 and P =.04, respectively) muscle spindle responses during the HVLA-SM thrust. The absence of preload had the greatest effect on the change in MIF. Interactions between preload magnitude, duration, and downward incisural point often produced statistically significant but arguably physiologically modest changes in the passive signaling properties of the muscle spindle after the manipulation. Conclusion Because preload parameters in this animal model were shown to affect neural responses to an HVLA-SM, preload characteristics should be taken into consideration when judging this intervention's therapeutic benefit in both clinical efficacy studies and in clinical practice. © 2014 National University of Health Sciences.