A users guide to HPA axis research

Abstract

Glucocorticoid hormones (cortisol and corticosterone - CORT) are the effector hormones of the hypothalamic-pituitary-adrenal (HPA) axis neuroendocrine system. CORT is a systemic intercellular signal whose level predictably varies with time of day and dynamically increases with environmental and psychological stressors. This hormonal signal is utilized by virtually every cell and physiological system of the body to optimize performance according to circadian, environmental and physiological demands. Disturbances in normal HPA axis activity profiles are associated with a wide variety of physiological and mental health disorders. Despite numerous studies to date that have identified molecular, cellular and systems-level glucocorticoid actions, new glucocorticoid actions and clinical status associations continue to be revealed at a brisk pace in the scientific literature. However, the breadth of investigators working in this area poses distinct challenges in ensuring common practices across investigators, and a full appreciation for the complexity of a system that is often reduced to a single dependent measure. This Users Guide is intended to provide a fundamental overview of conceptual, technical and practical knowledge that will assist individuals who engage in and evaluate HPA axis research. We begin with examination of the anatomical and hormonal components of the HPA axis and their physiological range of operation. We then examine strategies and best practices for systematic manipulation and accurate measurement of HPA axis activity. We feature use of experimental methods that will assist with better understanding of CORT’s physiological actions, especially as those actions impact subsequent brain function. This research approach is instrumental for determining the mechanisms by which alterations of HPA axis function may contribute to pathophysiology.

Dynamic glucocorticoid-dependent regulation of Sgk1 expression in oligodendrocytes of adult male rat brain by acute stress and time of day

“…The CORT-induced increase in Sgk1 mRNA was similar at both times of day. As expected, CORT treatment produced relatively high levels of plasma CORT 30 min after injection that were in the high physiological stress range [17], and these circulating levels were similar at both times of day.…”

“…Rats were acclimated to lighting conditions at the University of Colorado Boulder animal facility for two weeks before undergoing ADX surgery during the second half of the light phase. For three days after surgery, CORT (25 μg/ml; Steraloids Inc., Newport, RI) was replaced in the post-operative drinking water (tap water containing 0.1% ethanol, 0.9% saline, and 0.04% ibuprofen (Pfizer, Madison, NJ)) during the first two hours of the dark phase in order to preserve diurnal variations in baseline CORT levels [17]. On the third day after surgery, rats received no injection, vehicle injection (10% ethanol, 30% propylene glycol, 60% sterile saline, 1 ml/kg i.p.…”

“…Instead of use of the steroid displacement reagent provided by the kit, samples were instead diluted 1:50 in assay buffer and heated to 65°C for one hour in order to completely inactivate corticosteroid binding globulin [17]. Intra-assay coefficients of variation were less than 10% for each experiment.…”

Assessment of social behavior directed toward sick partners and its relation to central cytokine expression in rats

“…To do this, samples were immersed in a 75 °C water bath for 60 min. Prior assays conducted within our laboratory have demonstrated that this procedure produces superior denaturation of CBG compared to the steroid displacemernt step provided in the kit (unpublished observations, as discussed in [39, 40]). This assay has been shown to have a sensitivity of 27.0 pg/ml, with an inter-assay coefficient of 5.40%.…”

Screening for Cognitive Impairment by Model-Assisted Cerebral Blood Flow Estimation

“…There is evidence linking AD progression with decreased variability in CBF [8] and increased variability in BP [9]. Glucocorticoid hormone levels vary in 24-h cycles and dynamically increase with stress [10,11]. Circadian rhythms of cortisol (a key glucocorticoid hormone) are associated with changes in heart rate [12,13] and BP.…”