Locus Coeruleus Noradrenergic Modulation of Diurnal Corticosterone, Stress Reactivity and Cardiovascular Homeostasis in Male Rats

Introduction: Activation of the locus coeruleus-noradrenergic (LC-NA) system during awakening is associated with an increase in plasma corticosterone and cardiovascular tone. These studies evaluate the role of the LC in this corticosterone and cardiovascular response. Methods: Male rats, on day 0, were treated IP with either DSP4 (50 mg/ kg body weight) (DSP), a LC-NA specific neurotoxin, or normal saline (SAL). On day 10, animals were surgically prepared with jugular vein [Hypothalamic–pituitary–adrenal (HPA) axis] or carotid artery (hemodynamics) catheters and experiments performed on day 14. HPA axis activity, diurnally (circadian) and after stress [transient hemorrhage (14 mL/kg body weight) or airpuff-startle], and basal and post-hemorrhage hemodynamics were evaluated. On day 16, brain regions from a subset of rats were dissected for norepinephrine and corticotropin-releasing factor (CRF) assay. Results: In DSP rats compared to SAL rats: 1) regional brain norepinephrine was decreased but there was no change in median eminence or olfactory bulb CRF content; 2) during HPA axis acrophase, the plasma corticosterone response was blunted; 3) after hemorrhage and airpuff-startle, the plasma adrenocorticotropic hormone response was attenuated, whereas the corticosterone response was dependent on stressor category; 4) under basal conditions, hemodynamic measures exhibited altered blood flow dynamics and systemic vasodilation; and 5) after hemorrhage, hemodynamics exhibited asynchronous responses. Conclusion: LC-NA modulation of diurnal and stress-induced HPA axis reactivity occurs via distinct neurocircuits. The integrity of the LC-NA system is important to maintain blood flow dynamics. The importance of increases in plasma corticosterone at acrophase to maintain short- and long-term cardiovascular homeostasis is discussed.

Floor vibrations for motivation and feedback in the rat vibration actuating search task

Motivating rodents to perform cognitive tasks often relies on the application of aversive stimuli. The Vibration Actuating Search Task (VAST) is a novel open-field task in which gradient floor vibration provides motivation for the rodent to navigate in the direction of diminishing vibration to an unmarked target destination. Using floor vibration as a motivational stimulus may overcome several of the potential confounds associated with stimuli used in other tasks. In a series of three experiments, we determined whether (1) rats exhibit place preference for floor vibration over other aversive stimuli (i.e., water, foot shock, and bright light), (2) exposure to floor vibration is associated with a lower corticosterone response than exposure to these other stimuli, (3) rats successfully acquire the VAST, and (4) VAST performance is sensitive to 6 h of sleep deprivation (SD). Our results showed that rats exhibited place preference for vibration over water, foot shock, and bright light environments, and that corticosterone levels were lower in rats exposed to vibration than those exposed to water. VAST performance also significantly improved over two days of testing for some metrics, and SD impaired VAST performance. Overall, we conclude that (1) rats exhibit place preference for vibration over other stimuli commonly used to motivate task performance, (2) the vibrations employed by the VAST produce lower concentrations of circulating corticosterone than forced swimming, (3) rats can learn to use gradient floor vibration as a mode of performance feedback within two days of testing, and (4) VAST performance is substantially impaired by SD. Thus, the VAST is an effective and practical testbed for studying the mechanisms by which SD causes deficits in feedback-dependent decision making.

Mitochondrial stress-induced GDF15-GFRAL axis promotes anxiety-like behavior and CRH-dependent anorexia

Growth differentiation factor 15 (GDF15) is a stress-induced cytokine that modulates food intake and energy metabolism. Until now, most mechanistic studies on GDF15 rely on pharmacological interventions using exogenous GDF15, but little is known about its mode of action when induced both chronically and endogenously. Mitochondrial stress is one of the most described physiological conditions that induces GDF15, and therefore an important model to study the underlying mechanisms of endogenous GDF15 action. Here, using a mouse model of mitochondrial dysfunction via elevated respiratory uncoupling in skeletal muscle, we show a circadian oscillation of muscle-derived GDF15 to promote a daytime-restricted anorexia without signs of nausea or reduced physical activity, contrary to findings using recombinant GDF15. We find that mitochondrial stress-induced GDF15 associates with increased anxiety and hypothalamic corticotropin releasing hormone (CRH) induction, without further activation of the hypothalamic-pituitary-adrenal (HPA) axis and corticosterone response. Strikingly, the daytime-restricted anorexia, lean phenotype, systemic shift in substrate metabolism and anxiety-like behavior are completey abolished in conditions of mitochondrial stress coupled with genetic ablation of the GDF15 receptor GDNF receptor alpha-like (GFRAL), which is predominantly expressed in the hindbrain. Finally, we demonstrate that stress-induced GDF15-GFRAL signaling is required for hypothalamic CRH induction to control diurnal food intake in a CRH-receptor 1 (CRHR1)-dependent manner. With this, we uncover for the first time a molecular target of the GDF15-GFRAL axis that links anxiolytic and anorectic behavior as downstream effects of the chronic activation of this pathway by mitochondrial stress.

Activation of the hypothalamic–pituitary–adrenal axis by exogenous and endogenous GDF15

An acute increase in the circulating concentration of glucocorticoid hormones is essential for the survival of severe somatic stresses. Circulating concentrations of GDF15, a hormone that acts in the brain to reduce food intake, are frequently elevated in stressful states. We now report that GDF15 potently activates the hypothalamic–pituitary–adrenal (HPA) axis in mice and rats. A blocking antibody to the GDNF-family receptor α-like receptor completely prevented the corticosterone response to GDF15 administration. In wild-type mice exposed to a range of stressful stimuli, circulating levels of both corticosterone and GDF15 rose acutely. In the case of Escherichia coli or lipopolysaccharide injections, the vigorous proinflammatory cytokine response elicited was sufficient to produce a near-maximal HPA response, regardless of the presence or absence of GDF15. In contrast, the activation of the HPA axis seen in wild-type mice in response to the administration of genotoxic or endoplasmic reticulum toxins, which do not provoke a marked rise in cytokines, was absent in Gdf15−/− mice. In conclusion, consistent with its proposed role as a sentinel hormone, endogenous GDF15 is required for the activation of the protective HPA response to toxins that do not induce a substantial cytokine response. In the context of efforts to develop GDF15 as an antiobesity therapeutic, these findings identify a biomarker of target engagement and a previously unrecognized pharmacodynamic effect, which will require monitoring in human studies.

Ketamine decreases HPA axis reactivity to a novel stressor in male but not female mice

Ketamine is an antidepressant drug that interacts with the hypothalamic-pituitary-adrenal (HPA) axis, but whether this interaction is important for its behavioral effect is unknown. The goal of this experiment was to determine whether the behavioral response to ketamine depends on intact HPA axis function. Male and female C57BL/6J mice underwent chronic unpredictable stress prior to ketamine (30 mg/kg, i.p.) or vehicle treatment, with or without the glucocorticoid synthesis inhibitor metyrapone (20 mg/kg, i.p.) to block adrenal corticosterone production. Mice were tested in the forced swim test (FST) and open field test one and two days after injection, respectively. Fecal corticosterone was measured at select time points. No significant drug effects on behavior were observed. Males consistently had higher fecal corticosterone levels and stress-induced increases than females. Ketamine lowered the fecal corticosterone response to the FST only in males. These data show that ketamine after chronic stress decreases the corticosterone response to a novel stressor (the FST) in males, but not females. Corticosterone levels in all mice correlated with immobility in the FST, suggesting that shared neural circuitry could mediate both endocrine and behavioral responses. This circuitry may be ketamine-responsive only in males.

Is PTSD-Phenotype Associated with HPA-Axis Sensitivity? Feedback Inhibition and Other Modulating Factors of Glucocorticoid Signaling Dynamics

Previously, we found that basal corticosterone pulsatility significantly impacts the vulnerability for developing post-traumatic stress disorder (PTSD). Rats that exhibited PTSD-phenotype were characterized by blunted basal corticosterone pulsatility amplitude and a blunted corticosterone response to a stressor. This study sought to identify the mechanisms underlining both the loss of pulsatility and differences in downstream responses. Serial blood samples were collected manually via jugular vein cannula at 10-min intervals to evaluate suppression of corticosterone following methylprednisolone administration. The rats were exposed to predator scent stress (PSS) after 24 h, and behavioral responses were assessed 7 days post-exposure for retrospective classification into behavioral response groups. Brains were harvested for measurements of the glucocorticoid receptor, mineralocorticoid receptor, FK506-binding protein-51 and arginine vasopressin in specific brain regions to assess changes in hypothalamus–pituitary–adrenal axis (HPA) regulating factors. Methylprednisolone produced greater suppression of corticosterone in the PTSD-phenotype group. During the suppression, the PTSD-phenotype rats showed a significantly more pronounced pulsatile activity. In addition, the PTSD-phenotype group showed distinct changes in the ventral and dorsal CA1, dentate gyrus as well as in the paraventricular nucleus and supra-optic nucleus. These results demonstrate a pre-trauma vulnerability state that is characterized by an over-reactivity of the HPA and changes in its regulating factors.

Effects of test experience, closed-arm wall color, and illumination level on behavior and plasma corticosterone response in an elevated plus maze in male C57BL/6J mice: a challenge against conventional interpretation of the test

The elevated plus maze test is a widely used test for assessing anxiety-like behavior and screening novel therapeutic agents in rodents. Previous studies have shown that a variety of internal factors and procedural variables can influence elevated plus maze behavior. Although some studies have suggested a link between behavior and plasma corticosterone levels, the relationships between them remain unclear. In this study, we investigated the effects of experience with a battery of behavioral tests, the wall color of the closed arms, and illumination level on the behavior and plasma corticosterone responses in the elevated plus maze in male C57BL/6J mice. Mice were either subjected to a series of behavioral tests, including assessments of general health and neurological function, a light/dark transition test, and an open field test, or left undisturbed until the start of the elevated plus maze test. The mice with and without test battery experience were allowed to freely explore the elevated plus maze. The other two independent groups of naïve mice were tested in mazes with closed arms with different wall colors (clear, transparent blue, white, and black) or different illumination levels (5, 100, and 800 lx). Immediately after the test, blood was collected to measure plasma corticosterone concentrations. Mice with test battery experience showed a lower percentage of open arm time and entries and, somewhat paradoxically, had lower plasma corticosterone levels than the mice with no test battery experience. Mice tested in the maze with closed arms with clear walls exhibited higher open arm exploration than mice tested in the maze with closed arms with black walls, while there were no significant differences in plasma corticosterone levels between the different wall color conditions. Illumination levels had no significant effects on any measure. Our results indicate that experience with other behavioral tests and different physical features of the maze affect elevated plus maze behaviors. Increased open arm time and entries are conventionally interpreted as decreased anxiety-like behavior, while other possible interpretations are considered: open arm exploration may reflect heightened anxiety and panic-like reaction to a novel situation under certain conditions. With the possibility of different interpretations, the present findings highlight the need to carefully consider the test conditions in designing experiments and drawing conclusions from the behavioral outcomes in the elevated plus maze test in C57BL/6J mice.

Japanese quail (Coturnix japonica) as a novel model to study the relationship between the avian microbiome and microbial endocrinology-based host-microbe interactions

Background Microbial endocrinology, which is the study of neuroendocrine-based interkingdom signaling, provides a causal mechanistic framework for understanding the bi-directional crosstalk between the host and microbiome, especially as regards the effect of stress on health and disease. The importance of the cecal microbiome in avian health is well-recognized, yet little is understood regarding the mechanisms underpinning the avian host-microbiome relationship. Neuroendocrine plasticity of avian tissues that are focal points of host-microbiome interaction, such as the gut and lung, has likewise received limited attention. Avian in vivo models that enable the study of the neuroendocrine dynamic between host and microbiome are needed. As such, we utilized Japanese quail (Coturnix japonica) that diverge in corticosterone response to stress to examine the relationship between stress-related neurochemical concentrations at sites of host-microbe interaction, such as the gut, and the cecal microbiome. Results Our results demonstrate that birds which contrast in corticosterone response to stress show profound separation in cecal microbial community structure as well as exhibit differences in tissue neurochemical concentrations and structural morphologies of the gut. Changes in neurochemicals known to be affected by the microbiome were also identified in tissues outside of the gut, suggesting a potential relationship in birds between the cecal microbiome and overall avian physiology. Conclusions The present study provides the first evidence that the structure of the avian cecal microbial community is shaped by selection pressure on the bird for neuroendocrine response to stress. Identification of unique region-dependent neurochemical changes in the intestinal tract following stress highlights environmental stressors as potential drivers of microbial endocrinology-based mechanisms of avian host-microbiome dialogue. Together, these results demonstrate that tissue neurochemical concentrations in the avian gut may be related to the cecal microbiome and reveal the Japanese quail as a novel avian model in which to further examine the mechanisms underpinning these relationships.