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Alexander, S. L., & Irvine, C. H. (1998). The effect of social stress on adrenal axis activity in horses: the importance of monitoring corticosteroid-binding globulin capacity. Journal of Endocrinology, 157(3), 425–432.
Abstract: Plasma cortisol is largely bound to corticosteroid-binding globulin (CBG), which regulates its bioavailability by restricting exit from capillaries. Levels of CBG may be altered by several factors including stress and this can influence the amount of cortisol reaching cells. This study investigated the effect of social instability on plasma concentrations of CBG, total and free (not protein bound) cortisol in horses. Horses new to our research herd ('newcomers') were confined in a small yard with four dominant resident horses for 3-4 h daily for 3-4 (n = 5) or 9-14 (n = 3) days. Jugular blood was collected in the mornings from newcomers before the period of stress began ('pre-stress'), and then before each day's stress. Residents were bled before stress on the first and thirteenth day. Residents always behaved aggressively towards newcomers. By the end of the stress period, all newcomers were subordinate to residents. In newcomers (n = 8) after 3-4 days of social stress, CBG binding capacity had fallen (P = 0.0025), while free cortisol concentrations had risen (P = 0.0016) from pre-stress values. In contrast, total cortisol did not change. In residents, CBG had decreased slightly but significantly (P = 0.0162) after 12 days of stress. Residents and newcomers did not differ in pre-stress CBG binding capacity, total or free cortisol concentrations. However, by the second week of stress, CBG binding capacity was lower (P = 0.015) and free cortisol higher (P = 0.030) in newcomers (n = 3) than in residents. Total cortisol did not differ between the groups. In conclusion social stress clearly affected the adrenal axis of subordinate newcomer horses, lowering the binding capacity of CBG and raising free cortisol concentrations. However, no effect of stress could be detected when only total cortisol was measured. Therefore, to assess adrenal axis status accurately in horses, it is essential to monitor the binding capacity of CBG and free cortisol concentrations in addition to total cortisol levels.
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Irvine, C. H. G., & Alexander, S. L. (1994). Factors affecting the circadian rhythm in plasma cortisol concentrations in the horse. Domest. Anim. Endocrinol., 11(2), 227–238.
Abstract: In horses, a circadian rhythm in plasma cortisol concentrations has been reported in some but not all studies. When a rhythm occurred, horses were accustomed to a management routine, comprising stabling, feeding and sometimes exercise, which may entrain a circadian pattern. In this work, we monitored plasma cortisol by collecting jugular blood through indwelling cannulae from four groups: 1): 10 untrained, unperturbed mares grazing excess pasture, bled hourly for 26 hr; 2) 4 mares housed in a barn for 48 hr before sampling every 15 min for 20–24 hr; 3) 5 mares placed in an outdoor yard for sampling every 30 min from 0930–2100 hr; and 4) 4 stabled racehorses in training, bled every 30 min from 0730–2000 hr and once the following morning at 0830 hr. Plasma cortisol showed a similarly-timed circadian rhythm (P<0.0001) in all Group 1 horses, with a peak at 0600–0900 hr, and a nadir at 1800–2100 hr. By contrast, cortisol concentrations did not vary with time in either Group 2 or 3. Neither daily mean nor peak cortisol values differed in Group 1 and 2 (i.e. bled for >= 20 hr); however nadir values were higher (P<0.05) in Group 2. In Group 4, cortisol declined (P=0.004) during the sampling period but had returned to initial concentrations the next morning. Values did not differ from those for Group 1, except between 1000 and 1300 hr when cortisol in Group 4 was lower (P<0.05). We conclude that a circadian cortisol rhythm exists in horses in the absence of any known cues imposed by humans. However, this rhythm can be obliterated by the minor perturbation of removing the horse from its accustomed environment. By contrast, the rhythm occurs in trained racehorses, suggesting either that they have adapted to their environment thereby allowing an endogenous rhythm to emerge, or that the rhythm is entrained by their daily routine. These observations highlight the difficulties in determining the cortisol status of a horse, since measurements will be affected by time of day, the occurrence of short-term fluctuations, and how accustomed the horse is to its environment.
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