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Physiological changes provide indices of stress responses, however, behavioural measures may be easier to determine. Spontaneous eye blink rate has potential as a non-invasive indicator of stress. Eyelid movements, along with heart rate (HR) and behaviour, from 33 horses were evaluated over four treatments: 1) control—horse in its normal paddock environment; 2) feed restriction—feed was withheld at regular feeding time; 3) separation—horse was removed from visual contact with their paddock mates; and 4) startle test—a ball was suddenly thrown on the ground in front of the horse. HR data was collected every five s throughout each three min test. Eyelid movements and behaviours were retrospectively determined from video recordings. A generalized linear mixed model (GLIMMIX) procedure with Sidak’s multiple comparisons of least squares means demonstrated that both full blinks (16 ± 12b vs. 15 ± 15b vs. 13 ± 11b vs. 26 ± 20a full blinks/3 min ± SEM; a,b differ p < 0.006) and half blinks (34 ± 15ab vs. 27 ± 14bc vs. 25 ± 13c vs. 42 ± 22a half blinks/3 min ± SEM; a,b,c differ p < 0.0001) decreased during feed restriction, separation and the startle test compared to the control, respectively. Eyelid twitches occurred more frequently in feed restriction (p < 0.0001) along with an increased HR (p < 0.0001). This study demonstrates that spontaneous blink rate decreases while eyelid twitches increase when the horse experiences a stressful situation.

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Examining the characteristics of an animal's lying behaviour, such as frequency and duration of lying bouts, has become increasingly relevant for animal welfare research. Triaxial accelerometers have the advantage of being able to continuously monitor an animal's standing and lying behaviour without relying on live observations or video recordings. Multiple models of accelerometers have been validated for use in monitoring dairy cattle; however, no units have been validated for use in equines. This study tested Onset Pendant G data loggers attached to the hind limb of each of two mature Standardbred horses for a period of 5 days. Data loggers were set to record their position every 20 s. Horses were monitored via live observations during the day and by video recordings during the night to compare activity against accelerometer data. All lying events occurred overnight (three to five lying bouts per horse per night). Data collected from the loggers was converted and edited using a macro program to calculate the number of bouts and the length of time each animal spent lying down by hour and by day. A paired t-test showed no significant difference between the video observations and the output from the data loggers (P = 0.301). The data loggers did not distinguish standing hipshot from standing square. Predictability, sensitivity, and specificity were all >99%. This study has validated the use of Onset Pendant G data loggers to determine the frequency and duration of standing and lying bouts in adult horses when set to sample and register readings at 20 s intervals.