Comparison of Stress Response Pathways in Male and Female Athletes During Ultra‐Marathon Events

During extreme exercise competitions, changes in hormone concentrations can affect an athlete's overall performance. For example, high intensity exercise, such as running marathon distances and beyond, can cause physiological stress, and subsequent elevations in stress hormones, not apparent in low intensity cardio‐exercise programs. The goal of this study was to compare the activity of the hypothalamic‐pituitary‐adrenocortical (HPA) axis to autonomic nervous system production of salivary alpha‐amylase (AA) during ultra‐running events (distances beyond 26.2 miles). Saliva samples have the advantage of giving an accurate, minimally‐invasive measurement of CORT and AA levels in individual athletes and the levels of salivary CORT have been shown to accurately represent the levels taken from samples of plasma. Salivary CORT levels are directly correlated to the values seen when measuring plasma concentrations of CORT. Despite the well‐known effects of exercise on CORT concentrations and AA activity, it is unclear what effects running beyond the marathon distance has on these analytes. In this study, we investigated 1) the effect of high intensity cardio‐exercise, such as running up to 100 kilometers (K), on separate stress response systems and 2) sex differences in neuroendocrine responses to high intensity, long‐duration exercise. Saliva samples from recruited race participants were collected by the passive drool method from 15 male runners and 25 non‐pregnant female runners competing in the Pumpkin Holler Hunnerd race near Tahlequah, OK, on October 17, 2015. Samples were collected in 2 mL cryovial tubes (Salimetrics, Inc) with each runner submitting a 1 mL sample of saliva throughout the race at the following markers: See Figure 1 Samples were kept frozen on dry ice from collection until transport to a −20°C freezer. For processing, samples were removed from the freezer, thawed and analyzed for CORT concentration and AA activity. Concentration levels for CORT were determined using an expanded range high sensitivity salivary CORT enzyme immunoassay kit (Salimetrics, Inc.) performed according to the manufacturer's instructions. Upon completion, each plate was read in a plate reader (iMark, Bio‐Rad) at 450nm within 10 minutes of adding the stop solution. To ensure accurate results, the plate was then exposed to a secondary filter correction at 490 nm. AA activity was determined using a salivary AA kinetic enzyme assay kit (Salimetrics Inc.) performed according to the manufacturer's instructions; the plate was read at 415nm. All samples were assayed in duplicate. Current research suggests that CORT release is modified by exercise experience. Our preliminary results suggest that at 10K, CORT remains unaffected when compared to the start. However, CORT levels increase after 25K, and progressively increase up to 50K, after which point there is a distinct decline in CORT levels. These preliminary findings suggest that 25K could be a sufficient stimulus to increase the CORT levels while athletes who reach the 50K distance have exhausted their CORT production capability and the CORT levels begin to show a distinguishable decline. Preliminary AA levels do not parallel the CORT levels, which is consistent with previous studies showing AA activity as a pathway independent of the HPA axis. 1