Pamela Kent

Neural plasticity, neuropeptides and anxiety in animals--implications for understanding and treating affective disorder following traumatic stress in humans.

Exposure of rats to cats (predator stress) lastingly increases rodent anxiety-like behavior (ALB) in the elevated plus-maze. Previous work shows that lasting changes in ALB following predator stress depend on NMDA and CCKB receptors. In this paper we describe the effects of differing degrees of predator exposure on behavior. Effects depend on the behavioral measure. In general, exposure to predator odor is less provocative of lasting change in ALB than is unprotected exposure to a cat. In addition, we examine the development of effects of unprotected predator exposure over time. Lasting effects on ALB begin at 30 min to 1 h after predator stress and persist for at least 3 weeks. We also report a complex pattern of effects of predator stress on neuroendocrine and stress peptide (bombesin, CRF and AVP) levels in a variety of brain areas. Not surprisingly, predator exposure increases plasma levels of corticosterone and ACTH. Central changes in peptide content in the hypothalamo-pituitary axis, related hypothalamic nuclei, limbic and brain stem areas are also noted. Finally, path analysis demonstrates a replicable relationship between cat behavior, rat defensive behavior and degree of increase in ALB one week later. It is proposed that behavioral changes following predator stress may model anxiety associated with PTSD.

Corticotropin-releasing hormone, arginine vasopressin, gastrin-releasing peptide, and neuromedin B alterations in stress-relevant brain regions of suicides and control subjects.

BACKGROUND Postmortem levels of several stress- and depression-relevant neuropeptides were assessed in brain regions of depressed suicides relative to control subjects that had died of other causes. METHODS Brains of suicides and those that died from other causes were collected soon after death (typically <6 hours). Immunoreactivity levels (ir) of corticotropin-releasing hormone (CRH-ir) and arginine vasopressin (AVP-ir), and the bombesin analogs, gastrin-releasing peptide (GRP-ir), and neuromedin B (NMB-ir), were assessed. RESULTS Levels of CRH-ir among suicides were elevated in the locus coeruleus (LC), frontopolar, dorsolateral prefrontal (DMPFC) and ventromedial prefrontal cortices, but were reduced at the dorsovagal complex (DVC). The concentration of AVP-ir was elevated at the paraventricluar hypothalamic nucleus, LC, and DMPFC, and reduced at the DVC. Finally, GRP and NMB variations, which might influence anxiety states, were limited, although GRP-ir within the LC of suicides was higher than in control subjects, while NMB-ir was reduced at the DVC of suicides. CONCLUSIONS The data show several neuropeptide changes in relation to suicide, although it is premature to ascribe these outcomes specifically to the suicide act versus depression. Likewise, it is uncertain whether the neuropeptide alterations were etiologically related to suicide/depression or secondary to the depressive state.

Influence of psychogenic and neurogenic stressors on neuroendocrine and central monoamine activity in fast and slow kindling rats

The central neurochemical and neuroendocrine effects of a psychogenic (ferret exposure) and a neurogenic (restraint) stressor were assessed in rats that had been selectively bred for differences in amygdala excitability manifested by either Fast or Slow amygdala kindling epileptogenesis. While these rat lines differ in their emotionality, their behavioral styles were dependent on the nature of the stressor to which they were exposed. During restraint, the Slow rats were mostly immobile, while Fast rats persistently struggled. In contrast, Fast rats were more immobile in response to the ferret. Yet, the more emotional Slow rats exhibited a greater corticosterone response to the ferret, while comparable corticosterone responses between lines were evident following restraint. Although both stressors influenced norepinephrine (NE), dopamine (DA) and/or serotonin (5-HT) activity in brain regions typically associated with stressors (e.g., locus coeruleus, paraventricular nucleus of the hypothalamus, nucleus accumbens, prefrontal cortex), considerable amine alterations were evident in the medial and basolateral amygdala nuclei, but not in the central nucleus. Moreover, greater NE changes were apparent in the medial amygdala of the left hemisphere. Similarly, DA alterations also were greater in the left medial amygdala in response to stressors. Despite very different behavioral styles, however, the two lines often exhibited similar amine alterations in response to both stressors.

Stressor-Induced Corticotropin-Releasing Hormone, Bombesin, ACTH and Corticosterone Variations in Strains of Mice Differentially Responsive to Stressors

The effects of brief stressor exposure on hypothalamic-pituitary-adrenal (HPA) functioning was assessed in two strains of mice shown to be differentially responsive to stressors. Mild stress (1 min of cold swim, 20 C) led to marked elevations of plasma ACTH and corticosterone concentrations in the stress-reactive BALB/cByJ and the stress-resistant C57Bl/6ByJ mice. Moreover, it was observed that the strains differed in basal CRH content within the amygdala and the paraventricullar nucleus (PVN). Within 1 min of cold swim, the CRH changes were detected in these brain regions in BALB/cByJ mice, but were less apparent in C57Bl/6ByJ mice. Following a chronic stressor regimen, the marked elevations of plasma ACTH associated with acute stressors in BALB/cByJ mice were diminished. In contrast, in C57Bl/6ByJ mice in which acute stressors hardly affected ACTH concentrations, the chronic stressor regimen lead to a marked increase of plasma ACTH. Taken together, data indicate that the stress reactivity differences seen in the two strains of mice are not limited to ACTH and corticosterone, but are also detected with respect to CRH within the amygdala and PVN. Furthermore, the suggestion is offered that the reactivity differences in the two strains of mice may have lead to different profiles of ACTH secretagogues and hence the response profile to later acute and chronic stressors differed in these strains of mice.

Bombesin Receptors as a Novel Anti-Anxiety Therapeutic Target: BB1 Receptor Actions on Anxiety through Alterations of Serotonin Activity

The effects of PD 176252 [3-(1H-indol-3-yl)-N-[1-(5-methoxy-pyridin-2-yl)-cyclohexylmethyl]-2-methyl-2-[3-(nitro-phenyl)ureido]propionamide], a nonpeptide bombesin (BB) BB1/BB2 receptor antagonist, were assessed in rats using several ethologically relevant tests of anxiety. Consistent with a role for the bombesin family of peptides in subserving anxiety behaviors, the antagonist increased social interaction (3.75 and 7.5 mg/kg, i.p.), dose-dependently attenuated the number of vocalizations emitted by guinea pig pups separated from their mother (1–30 mg/kg, i.p.), reduced latency to approach a palatable snack in an anxiogenic (unfamiliar) environment, and reduced the fear-potentiated startle response (5 and 10 mg/kg, i.p., and 100–200 ng per rat, i.c.v.). When administered directly to the dorsal raphé nucleus (DRN), PD 176252 (20–500 ng) increased social interaction under aversive conditions, as did the 5-HT1A receptor agonist 8-hydroxy-2(di-n-propylamino)tetralin (50 ng). Furthermore, intra-DRN microinfusion of the peptide antagonist (PD 176252) suppressed, whereas its agonist [neuromedin B (NMB)-30] promoted, the in vivo release of 5-HT in the ventral hippocampus. In parallel, the suppressed social interaction elicited by intra-DRN administration of NMB was attenuated by a systemically administered 5-HT2C (but not 5-HT1A) receptor antagonist. Together, these findings suggest that endogenous BB-like peptides at the DRN evoke the release of 5-HT from the limbic nerve terminals originating from the raphé, specifically at the ventral hippocampus, resulting in anxiogenesis. The finding that this action was attenuated by BB receptor (BB1 and/or BB2) antagonists suggests that these compounds may represent a novel class of anxiolytic agents.

Influence of Psychogenic and Neurogenic Stressors on Endocrine and Immune Activity: Differential Effects in Fast and Slow Seizing Rat Strains

Variations of plasma ACTH and corticosterone, as well as splenic macrophage activity and mitogen-induced cell proliferation, were determined in rats following 15 min of either the neurogenic stressor of restraint or by a purely psychogenic stressor consisting of exposure to a ferret. The effects of these stressors were assessed in two strains of rats that were selectively bred for either Fast or Slow kindling epileptogenesis triggered in response to amygdala stimulation. The stressors differentially influenced behavioral responses, endocrine activity, and immune functioning, and these effects varied with the strain of rat. In response to restraint the Fast rats exhibited protracted struggling, while the Slow rats tended to be immobile. In contrast, upon ferret exposure the Fast rats showed greater immobility than the Slow rats. The stressors also induced marked elevations of plasma ACTH and corticosterone. Whereas the ACTH and corticosterone increases were more pronounced in response to the ferret in the Slow rats, restraint resulted in a markedly greater rise of plasma ACTH in the Fast strain. Proliferation of splenic lymphocytes in response to Con A and LPS were elevated in Fast seizing rats, while macrophage activity, as determined by oxygen burst following addition of PMA and luminol to splenic mononuclear cells, was greater in the Slow seizing strain. While neither stressor influenced cell proliferation in either the Fast or Slow rats, macrophage activity was greatly suppressed by ferret exposure only in the Slow rats. Taken together, it appears that while stressors influence behavior and immune and endocrine functioning, these effects may vary as a function of the interaction of the strain of rat and the specific type of stressor employed.

Role of gastrin-releasing peptide and neuromedin B in anxiety and fear-related behavior

Bombesin (BB)-like peptides have been implicated in the mediation and/or modulation of the stress response. However, the impact of manipulating this peptidergic system has only been assessed in a limited number of anxiety and fear paradigms. Given that different behavioral paradigms reflect different aspects of anxiety, the objective of the present investigation was to assess the effects of two mammalian BB-related peptides, namely gastrin-releasing peptide (GRP) and neuromedin B (NMB), in paradigms thought to reflect fear and anxiety-related behaviors. To this end, the effects of central (3rd ventricular; i.c.v.) administration of GRP (0.30 nmol), GRP receptor (BB(2)) antagonist, [Leu(13)-(CH(2)NH)Leu(14)]-BN (1.26 nmol), NMB-30 (0.29 nmol), NMB (BB(1)) receptor antagonist, BIM 23127 (1.70 nmol) and a mixed BB(1)/BB(2) receptor antagonist, PD 176252 (0.621 nmol) were assessed in the elevated plus maze (EPM) and in a fear potentiated startle paradigm (a model thought to reflect conditioned fear). The BB(1) receptor antagonist and the mixed BB(1)/BB(2) receptor antagonist elicited anxiolytic effects in the EPM, whereas, the BB(2) receptor antagonist was without effect. In the fear potentiated startle paradigm, pretreatment with either the BB(1) receptor antagonist or the BB(2) receptor agonist attenuated the fear potentiated startle response, without affecting basal startle amplitude. These data suggest that NMB and GRP do affect the stress response. However, whereas NMB manipulations affected both anxiety and fear responses, GRP alterations selectively affected fear-related responses.

ARE BOMBESIN-LIKE PEPTIDES INVOLVED IN THE MEDIATION OF STRESS RESPONSE?

The neurochemical mechanisms underlying the coincident activation of the hypothalamic-pituitary-adrenal (HPA) axis and the sympathetic nervous system in response to stress remain unclear. Central injection of the neuropeptide bombesin (BN) potently stimulates the release of epinephrine from the adrenal medulla, adrenocorticotropic hormone (ACTH) from the pituitary gland, and elicits behaviors typically associated with increased emotionality and arousal. The current studies assessed whether stress is associated with 1) fluctuations in the endogenous regional levels of BN-like peptides and/or 2) changes in BN receptor density. Male Sprague-Dawley rats received either no treatment or were subjected to acute immobilization stress for 10, 30 or 120 min. Plasma ACTH levels increased in response to stress, peaking at 30 min. BN-like immunoreactivity increased significantly at the hypothalamus and medulla, within 30 min; however with more sustained immobilization (120 min) BN-like immunoreactivity declined to control levels. Levels of BN-like peptides remained unchanged in several other regions, including the hippocampus, striatum, midbrain, pituitary, and pons. Autoradiographic analysis revealed that the density of BN receptor varied in a regionally specific manner. Significant stress related increases in binding were found at the nucleus of the solitary tract (at 30 and 120 min), and at the paraventricular (at 120 min) and arcuate nuclei (at 120 min) of the hypothalamus. These data indicate the BN-like peptides may play a role in the mediation and/or modulation of response to stress.

Differential involvement of amygdaloid CRH system(s) in the salience and valence of the stimuli

Anxiety is a heterogeneous term encompassing not only state or trait characteristics but also a wide range of pathologies such as generalized anxiety disorders, phobias, panic and obsessive-compulsive disorders, acute stress disorder, and posttraumatic stress disorder. Given that diverse forms of anxiety exist, numerous animal models have been developed, which are considered to be useful in identifying mechanisms underlying anxiety states. Examples of such animal models include paradigms that assess the behavioral response to neurogenic (or painful stimuli) or psychogenic stressors or to cues that had previously been associated with painful stimuli. The present report presents data regarding the impact of stressors on corticotropin-releasing hormone (CRH), and relates these to changes in anxiety-like states. Specifically, we demonstrate that (1) psychogenic stressors influence the in vivo release of CRH at the central nucleus of the amygdala (CeA); (2) although CRH changes within the CeA are exquisitely sensitive to stressors, they are also elicited by positive stimuli; and (3) while treatment with diazepam attenuates behavioral signs of anxiety, the CRH release associated with a stressor is unaffected by the treatment. The position is offered that although release of CRH within the CeA is increased under stressful conditions, it is not a necessary condition for the consequent behavioral expression of anxiety-like reactions, at least not in minimally threatening situations. We suggest that the CRH responses at the CeA may be involved in a preparatory capacity and, as such, may accompany a range of emotionally significant stimuli, be they appetitive or aversive.

Effects of corticosterone on corticotrophin-releasing hormone and gastrin-releasing peptide release in response to an aversive stimulus in two regions of the forebrain (central nucleus of the amygdala and prefrontal cortex)

Previous research has shown that chronic corticosterone treatment increases the expression of corticotrophin‐releasing hormone (CRH) mRNA at the central nucleus of the amygdala (CeA). Like CRH, gastrin‐releasing peptide (GRP) appears to be involved in mediation of the stress response and is released at the CeA during exposure to an acute stressor. Using in‐vivo microdialysis, this study examined the effects of corticosterone treatment on the release of CRH and GRP in response to an airpuff challenge at two forebrain regions, the CeA and medial prefrontal cortex. Adrenally intact rats were treated with corticosterone by systemic implants over a 14‐day period prior to microdialysis probe insertion. We found that, at both regions, the airpuff‐induced CRH and GRP release were enhanced in the corticosterone pellet‐implanted rats as compared with the release observed in the vehicle‐implanted control rats. These findings suggest that chronic corticosterone exposure potentiates the stressor‐elicited release of CRH and GRP. As cortisol dysregulation has frequently been reported in people with psychiatric conditions, such as anxiety disorders or depression, a better understanding of the glucocorticoid‐mediating regulation of CRH and GRP may provide insight into the underlying neurochemical mechanisms involved in both adaptive fear‐type responses and maladaptive responses leading to pathology.