Quantitative assessment of the nociceptive withdrawal reflex in healthy, non-medicated experimental sheep

This study aimed to characterize the nociceptive withdrawal reflex (NWR) and to define the nociceptive threshold in 25 healthy, non-medicated experimental sheep in standing posture. Electrical stimulation of the dorsal lateral digital nerves of the right thoracic and the pelvic limb was performed and surface-electromyography (EMG) from the deltoid (all animals) and the femoral biceps (18 animals) or the peroneus tertius muscles (7 animals) was recorded. The behavioural reaction following each stimulation was scored on a scale from 0 (no reaction) to 5 (strong whole body reaction). A train-of-five 1 ms constant-current pulse was used and current intensity was stepwise increased until NWR threshold intensity was reached. The NWR threshold intensity (It) was defined as the minimal stimulus intensity able to evoke a reflex with a minimal Root-Mean-Square amplitude (RMSA) of 20 μV, a minimal duration of 10 ms and a minimal reaction score of 1 (slight muscle contraction of the stimulated limb) within the time window of 20 to 130 ms post-stimulation. Based on this value, further stimulations were performed below (0.9It) and above threshold (1.5It and 2It). The stimulus-response curve was described. Data are reported as medians and interquartile ranges. At the deltoid muscle It was 4.4 mA (2.9–5.7) with an RMSA of 62 μV (30–102). At the biceps femoris muscle It was 7.0 mA (4.0–10.0) with an RMSA of 43 μV (34–50) and at the peroneus tertius muscle It was 3.4 mA (3.1–4.4) with an RMSA of 38 μV (32–46). Above threshold, RMSA was significantly increased at all muscles. Below threshold, RMSA was only significantly smaller than at It for the peroneus tertius muscle but not for the other muscles.

Repeated electrical stimulations as a tool to evoke temporal summation of nociceptive inputs in healthy, non-medicated experimental sheep.

The nociceptive withdrawal reflex (NWR) model is used in animal pain research to quantify nociception. The aim of this study was to evaluate the NWR evoked by repeated stimulations in healthy, non-medicated standing sheep. Repeated electrical stimulations were applied at 5Hz for 2s to the digital nerves of the right thoracic and the pelvic limbs of 25 standing sheep. The stimulation intensities applied were fractions (0.5, 0.6, 0.7, 0.8, 0.9 and 1) of the individual previously determined nociceptive threshold (It) after single stimulation. Surface-electromyographic activity (EMG) was recorded from the deltoid, the femoral biceps or the peroneus tertius muscles. The repeated stimulation threshold (RS It) was reached if at least one stimulus in the train was followed by a reflex with a minimal root-mean-square-amplitude (RMSA) of 20μV. The behavioural reaction following each series of stimulations was scored on a scale from 0 (no reaction) to 5 (vigorous whole-body reaction). For the deltoid muscle, RS It was 2.3mA (1.6-3mA) with a reaction score of 2 (1-2) and at a fraction of 0.6 (0.5-0.8)×It. For the biceps femoris muscle, RS It was 2.9mA (2.6-4mA) with a reaction score of 1 (1-2) at a fraction of and 0.55 (0.4-0.7)×It while for the peroneus tertius muscle RS It was 3mA (2.8-3.5mA) with a reaction score of 1 (1-2) and at a fraction of 0.8 (0.8-0.95)×It. Both, RMSA and reaction scores increased significantly with increasing stimulation intensities in all muscles (p<0.001). The repeated application of electrical stimuli led to temporal summation of nociceptive inputs and therefore a reduction of the stimulus intensity evoking a withdrawal reaction in healthy, standing sheep. Data achieved in this study can now serve as reference for further clinical or experimental applications of the model in this species.