Dean A. Myers

Corticosterone delivery to the amygdala increases corticotropin-releasing factor mRNA in the central amygdaloid nucleus and anxiety-like behavior

The present study examined the effects of stereotaxic delivery of corticosterone to the amygdala on anxiety-like behavior and corticotropin-releasing factor (CRF) mRNA level in the central nucleus of the amygdala (CeA). Micropellets (30 microg) of crystalline corticosterone or cholesterol (control) were implanted bilaterally at the dorsal margin of the CeA in Wistar rats. Seven days post-implantation, anxiety-like behavior was accessed using an elevated plus-maze. CRF mRNA level in the CeA was determined by in situ hybridization 4 h after being tested on the elevated plus-maze. Corticosterone implants increased indices of anxiety on the elevated plus-maze and produced a concomitant increase in both basal level of CRF mRNA per neuron and the number of neurons with CRF hybridization signal in the CeA. The plus-maze increased CRF mRNA levels in the CeA of cholesterol implanted rats to the elevated basal levels observed in corticosterone treated animals. Exposure to the plus-maze did not increase CRF mRNA level in the CeA of corticosterone implanted rats beyond elevated basal levels. Taken together, these findings support the involvement of the amygdala in anxiety-like behaviors in response to chronically elevated corticosterone and suggests that elevated glucocorticoids may increase anxiety by inducing CRF expression in the CeA.

Stereotaxic delivery of corticosterone to the amygdala modulates colonic sensitivity in rats.

Episodes of anxiety are often associated with the onset or exacerbation of visceral pain in patients with irritable bowel syndrome (IBS). The central amygdaloid nucleus (CeA) is a key limbic structure involved in the expression of anxiety as well as a major site for regulating autonomic and visceral responses to stress. Previous experiments have shown that glucocorticoids can act directly at the CeA to increase the level of anxiety in rats. Therefore, the goal of this study was to examine the effect of stereotaxic delivery of corticosterone into the CeA on the development of visceral hypersensitivity by measuring visceromotor response to colorectal distention in rats. Stereotaxic delivery of corticosterone to the CeA increases indices of anxiety and produces a hypersensitive colon as demonstrated by an exaggerated visceromotor response to colorectal distention in the F344 rat strain. Our findings suggest that modulation of anxiety by manipulating amygdala function with corticosterone induced colonic hypersensitivity via descending neuronal pathways from the CeA.

Evidence for visceral hypersensitivity in high-anxiety rats.

In patients with irritable bowel syndrome, anxiety is often associated with visceral pain. Based on this information we hypothesized that rats genetically predisposed to anxiety have an increased visceral sensitivity. To test this hypothesis, visceromotor reflex recordings in response to colorectal distention were used to estimate the level of visceral stimulation in high; moderate-, and low-anxiety rats. We compared the effect of innocuous colorectal distension in rats with and without sensitized colons. In nonsensitized rats visceromotor responses were increased by colorectal distention with the greatest response in the high-anxiety Wistar-Kyoto strain. Sensitization of the colon significantly increased visceromotor responses to colorectal distention in all rat strains. In summary, our data suggested that a manifestation of a genetically determined anxiety level appeared to be abnormal neural responsiveness of the gastrointestinal tract leading to visceral hypersensitivity in high-anxiety animals.

Corticotropin‐releasing factor 1 receptor‐mediated mechanisms inhibit colonic hypersensitivity in rats

Abstract  The potential relationship between stress and irritable bowel syndrome (IBS) symptomatology suggests a possible role for stress‐mediating hormones, such as corticotropin‐releasing factor (CRF), in the altered perception of stimuli in IBS patients. In previous studies, Wistar–Kyoto (WKY) rats with genetic indices of high anxiety demonstrated colonic hypersensitivity coupled with a high basal level of CRF within the central nervous system. In the current study we tested the hypothesis that a selective, non‐peptide CRF1 receptor antagonist, antalarmin, would inhibit hypersensitivity in the WKY rat colon. Colonic sensitivity was determined by monitoring a visceromotor behavioural response during innocuous levels of colorectal distention (30 mmHg). In high anxiety WKY rats we found that antalarmin (20 mg kg−1, i.p.) significantly decreased the visceromotor response induced by colorectal distention. In a second study central administration (i.c.v.) of CRF was used to induce colonic hypersensitivity in lower anxiety Fischer 344 (F‐344) rats, and in this model, antalarmin significantly inhibited the CRF‐induced colonic hypersensitivity. In summary, a selective CRF1 receptor antagonist, antalarmin, inhibits colonic hypersensitivity apparent in WKY rats or in F‐344 rats given a central administration of CRF. Our findings suggest that CRF1 receptor antagonism may represent a novel therapeutic approach for the treatment of IBS.

Stereotaxic localization of corticosterone to the amygdala enhances hypothalamo-pituitary–adrenal responses to behavioral stress

The amygdala is involved in behavioral, autonomic, and neuroendocrine responses to stressful stimuli. The goal of the current study was to determine the effect of directly elevating glucocorticoids in the amygdala on hypothalamo-pituitary-adrenocortical (HPA) responses to the elevated plus maze, a behavioral stressor known to activate the amygdala. Micropellets (30 microg) of crystalline corticosterone or cholesterol (control) were implanted bilaterally at the dorsal margin of the CeA in male Wistar rats; vascular catheters were also placed at this time. Five days post-surgery, blood samples were drawn at 07:00 and 19:00 h to assess diurnal rhythm of plasma corticosterone. At 7 days post-implantation, rats were subjected to behavioral stress using an elevated plus maze and blood was collected 15 min prior to stress, and at 15, 45, and 90 min after the initiation of the stressor. Corticotropin releasing factor (CRF) and arginine vasopressin (AVP) mRNA levels were analyzed by in situ hybridization in the medial parvocellular division of the hypothalamic paraventricular nucleus (mpPVN) in corticosterone- and cholesterol-implanted rats either not exposed to the elevated plus maze (control) or 4 h post-behavioral stress. Localization of corticosterone to the amygdala had no effect on diurnal rhythm of corticosterone secretion. Behavioral stress significantly increased peak plasma corticosterone levels in both groups to a similar level. However, in the corticosterone implanted rats, plasma corticosterone concentrations at 45 and 90 min post-stress were significantly greater compared to control rats indicating a prolonged corticosterone response to behavioral stress. In non-stressed rats, corticosterone delivery to the amygdala elevated basal CRF mRNA in the mpPVN to levels similar to those observed post-stress in control animals; no further increase was observed in CRF mRNA following stress. Behavioral stress resulted in a significant elevation in CRF mRNA in cholesterol controls. Basal AVP mRNA levels were unaffected by corticosterone implants. AVP mRNA did not increase in cholesterol implanted rats in response to behavioral stress. However, AVP mRNA levels were higher in corticosterone implanted rats post stress compared to cholesterol treated controls. In conclusion, direct administration of corticosterone to the amygdala increases plasma corticosterone in response to a behavioral stressor without altering the diurnal rhythm in plasma corticosterone. Elevated basal levels of mpPVN CRF mRNA, and the induction of a mpPVN AVP mRNA response to the behavioral stressor implicate enhanced ACTH secretagogue expression in the increased HPA response to corticosterone modulation of amygdala function.

Corticosterone implants to the amygdala and type 1 CRH receptor regulation: effects on behavior and colonic sensitivity.

Corticosterone (CORT) micropellets were stereotaxically placed bilaterally at the dorsal margin of the central nucleus of the amygdala (CeA). Both behavioral and physiological responses were recorded (plus maze and colonic discomfort) at 7 days post-implantation. Corticosterone reduced the exploration of the plus maze and increased colonic distress. The ability of a CRH type 1 receptor antagonist, antalarmin, to block behavioral and colonic effects of central placement of CORT was also examined. The diminished exploration in the plus maze and colon distress observed in response to CORT placement at the CeA were averted by the administration of antalarmin. These results provide further evidence for the role of the CRH type 1 receptor to ameliorate both behavioral and physiological functions.

Strain differences in anxiety-like behavior: Association with corticotropin-releasing factor

The purpose of the current study was to relate basal corticotropin-releasing factor (CRF) mRNA level in the central nucleus of the amygdala (CeA) with anxiety-like behavior using three strains of rat reported to exhibit a range of behavioral and neuroendocrine responses to stress. Anxiety-like behavior was determined for Fischer (F344), Wistar, and Wistar-Kyoto (WKY) rats with an elevated plus-maze and CRF mRNA level was measured using in situ hybridization. WKY rats exhibited more anxiety-like behavior on the elevated plus-maze than both F344 (ps < 0.01) and Wistar rats (ps < 0.05). WKY rats had higher basal levels of CRF mRNA in the CeA than F344 rats (p < 0.05) with a trend toward higher levels than Wistar rats (p = 0.06). Wistar rats had similar indices of anxiety with F344 rats and comparable levels of CRF mRNA in the CeA. Basal plasma corticosterone was similar for all three strains and repeated experience with the plus-maze had no effect on basal corticosterone levels or CRF mRNA levels in the paraventricular nucleus of the hypothalamus (PVN) for Wistar or WKY rats. Consistent with reported hyperactivity of the hypothalamopituitary adrenal axis of F344 rats with repeated stress, we observed elevated corticosterone following repeated exposure to the elevated plus-maze in F344 rats (p < 0.01) with a trend toward increased CRF mRNA levels in the PVN (p = 0.09). Heightened expression of CRF in the CeA of WKY rats may be involved in anxiety-like behaviors observed in this strain.

Chronically elevated corticosterone in the amygdala increases corticotropin releasing factor mRNA in the dorsolateral bed nucleus of stria terminalis following duress

Elevated corticosterone in the amygdala leads to cautious avoidance behaviors on the elevated plus maze. The current study examined the effect of elevated corticosterone in the amygdala on corticotropin releasing factor (CRF) mRNA levels in the bed nuclei of stria terminalis (BNST). Exposure to the elevated plus maze increased CRF mRNA in both the dorsolateral and ventrolateral BNST. Corticosterone implants in the amygdala potentiated this effect in the dorsolateral, but not ventrolateral BNST. Corticosterone also resulted in an increased number of neurons expressing CRF in the dorsolateral BNST in response to the behavioral stimuli.

Long-term hypoxia modulates expression of key genes regulating adipose function in the late-gestation ovine fetus

A major function of abdominal adipose in the newborn is nonshivering thermogenesis. Uncoupling protein (UCP) UCP1 and UCP2 play major roles in thermogenesis. The present study tested the hypothesis that long-term hypoxia (LTH) modulates expression of UCP1 and UCP2, and key genes regulating expression of these genes in the late-gestation ovine fetus. Ewes were maintained at high altitude (3,820 m) from 30 to 138 days gestation (dG); perirenal adipose tissue was collected from LTH and age-matched, normoxic control fetuses at 139-141 dG. Quantitative real-time PCR was used to analyze mRNA for UCP1, UCP2, 11beta hydroxysteroid dehydrogenase type 1 (HSD11B1) and 2 (HSD11B2), glucocorticoid receptor (GR), beta3 adrenergic receptor (beta3AR), deiodinase type 1 (DIO1) and DIO2, peroxisome proliferator activated receptor (PPAR) alpha and gamma and PPARgamma coactivator 1 (PGC1alpha). Concentrations of mRNA for UCP1, HSD11B1, PPARgamma, PGC1, DIO1, and DIO2 were significantly higher in perirenal adipose of LTH compared with control fetuses, while mRNA for HSD11B2, GR, or PPARalpha in perirenal adipose did not differ between control and LTH fetuses. The increased expression of UCP1 is likely an adaptive response to LTH, assuring adequate thermogenesis in the event of birth under oxygen-limiting conditions. Because both glucocorticoids and thyroid hormone regulate UCP1 expression, the increase in HSD11B1, DIO1, and DIO2 implicate increased adipose capacity for local synthesis of these hormones. PPARgamma and its coactivator may provide an underlying mechanism via which LTH alters development of the fetal adipocyte. These findings have important implications regarding fetal/neonatal adipose tissue function in response to LTH.

eNOS activation and NO function: Differential control of steroidogenesis by nitric oxide and its adaptation with hypoxia

Nitric oxide (NO) plays a role in a wide range of physiological processes. Aside from its widely studied function in the regulation of vascular function, NO has been shown to impact steroidogenesis in a number of different tissues. The goal of this review is to explore the effects of NO on steroid production and further, to discern its source(s) and mechanism of action. Attention will be given to the regulation of NO synthases in specific endocrine tissues including ovaries, testes, and adrenal glands. The effects of hypoxia on generation of NO and subsequent effects on steroid biosynthesis will also be examined. Finally, a potential model for the interaction of hypoxia on NO synthesis and steroid production is proposed.