Investigation into circannual variation in endogenous plasma ACTH concentrations in Ontario senior horses

Kristiina Ruotsalo, Alison Moore

Animal Health Laboratory, University of Guelph, Guelph, ON (Ruotsalo); Ontario Ministry of Agriculture, Food and Rural Affairs (Moore)

AHL Newsletter 2023;27(2):21.

PPID (pars intermedia pituitary dysfunction) is an endocrine disorder that occurs in over 20% of aged horses, ponies and donkeys.  Clinical signs and measurement of plasma endogenous ACTH (eACTH) concentrations currently form the basis of diagnosis, as dynamic testing, such as the thyrotropin-releasing hormone (TRH) stimulation test, is not easily accessible due to the lack of a readily available source of thyrotropin.  A number of factors can affect eACTH concentrations, including individual variation, timing of sampling with respect to the course of disease, and increases related to illness, pain, or stress, thus possibly confounding clinical interpretation.  Additionally, the equine pituitary gland exhibits a circannual rhythm in the release of eACTH.  This circannual increase has previously been referred to as a ‘seasonal or fall’ increase, however it is now recognized that rather than exhibiting a sharp autumnal peak in eACTH concentrations, gradually increasing secretory activity begins during the summer, peaks, and then declines.  The timing of this circannual increase has been shown to vary with respect to geographic location, and is speculated to be linked to photoperiods.  The circannual increase occurs both in clinically healthy horses and those affected with PPID, although the increase in affected horses is typically of greater magnitude.

A preliminary study was undertaken with the support of the OAHN Equine Network to characterize the onset, duration, and magnitude of the circannual increase in eACTH concentrations in a group of clinically healthy senior Ontario horses of various breeds and both sexes.  Participating veterinarians recruited horses aged 15 years or older for monthly eACTH sampling with the owner’s consent.  Horses were considered healthy based on physical examination, health status questionnaire, and unremarkable study intake CBC and biochemistry profile results.  No horses receiving medication for PPID, EMS, thyroid hormone supplementation, or corticosteroids were entered into the study.  Horses remained in Ontario for the duration of the sampling period.  No ponies, donkeys, mules or miniature horses were included, due to the potential breed effect on eACTH concentrations.  Timing of sample collection was not standardized, however, a minimum of one hour post exercise or training was recommended.

EDTA plasma samples were received frozen and analyzed using an ACTH chemiluminescent immunoassay and Immulite 1000 instrumentation (Siemens, Canada) at the Animal Health Laboratory.

Twenty-three horses, ranging in age from 16 to 23 years entered the study.  Seven horses completed all 12 months of consecutive sampling.  Remaining horses were sampled 5 to 11 times.  The reasons for the loss of horses from the study were not investigated.

Due to limited eACTH results at some time points, descriptive analysis of data was deemed most appropriate.  Average eACTH concentrations approached 10 pmol/L in June and July, reached a peak value of 17.3 pmol/L in September, remained mildly increased in October, and then resumed concentrations of <10 pmol/L, thus supporting a circannual increase in eACTH from June to October (Fig. 1).

A large degree of individual variation with respect to the magnitude and onset of increase in plasma eACTH concentrations was evident.

Data from two horses was excluded as eACTH concentrations significantly exceeded 10 pmol/L at most measured time points, reaching values greater than 100 pmol/L during the months of June to October.  These two horses were reportedly clinically normal, but the possibility of occult PPID needs to be considered.  Horses with PPID do exhibit evidence of circannual increases in eACTH, although these increases are exaggerated in comparison to clinically healthy individuals.

Additional investigations involving a larger cohort of clinically well senior horses, targeting the June to October sampling interval, will be required to establish statistically significant reference intervals.

In the interim, the following guidelines for interpretation of eACTH concentrations may be clinically helpful, recognizing that the circannual change in eACTH is gradual and subject to individual variation:

  • December to mid-June: eACTH concentrations > 10 pmol/L may support a diagnosis of PPID in horses with the appropriate signalment and clinical signs;
  • July to mid-November: eACTH concentrations >20 pmol/L may support a diagnosis of PPID  in horses with appropriate signalment and clinical signs;
  • Individual horses may have higher circannual increases in eACTH concentrations, with a return to baseline in late winter / early spring;
  • As with all laboratory tests, eACTH results must be interpreted in the context of age, clinical history, and physical examination findings.

Figure 1.  Average monthly plasma eACTH concentrations in a group of Ontario senior horses.

Figure 1.  Average monthly plasma eACTH concentrations in a group of Ontario senior horses.

We would like to thank all of the veterinarians participating in this study, the horses, and their owners.   AHL  


1.Durham A, et al. Clinically and temporally specific diagnostic thresholds for plasma ACTH in the horse. Equine Vet J 2021:53:250-260.

2. Durham AE, Potier JF, Huber L. The effect of month and breed on plasma adrenocorticotropic hormone concentration in equids. The Veterinary Journal 2022;286:105857.

3. Equine Endocrinology Group: Recommendations for the Diagnosis and Treatment of Pituitary Pars Intermedia Dysfunction (PPID).


This study is available on the and websites:

OAHN Equine Network Project: Investigation into the seasonal variations in ACTH levels in Ontario’s senior horses - Ontario Animal Health Network

OAHN Projects - ResearchONequine