Two immunohistochemical methods that allow the concurrent localization of neuroactive substances within individual neurons have been used to identify, count, and chart the distribution of corticotropin-releasing factor (CRF)-immunoreactive cells in the paraventricular nucleus of the hypothalamus (PVH) that may also contain an additional peptide. In colchicine-treated male rats a moderate number of oxytocin-stained cells, localized primarily in a discrete, anterior part of the magnocellular division of the nucleus, was found also to stain positively for CRF. Similarly, oxytocin and CRF immunoreactivity were jointly expressed in magnocellular neurons distributed diffusely in the supraoptic nucleus. Smaller numbers of vasopressin- and neurotensin-stained neurons centered in specific parts of the parvocellular division of the PVH were stained with antisera against CRF. Possible mechanisms whereby the function of subsets of magnocellular and parvocellular neurosecretory neurons can be modulated differentially are discussed.

Although corticotropin-releasing hormone (CRH), a regulator of stress responses, acts through two receptors (CRH1 and CRH2), the role of CRH2 in stress responses remains unclear. Knock-out mice without the CRH2 gene exhibit increased stress-like behaviors. This profile could result either directly from the absence of CRH2 receptors or indirectly from developmental adaptations. In the present study, CRH2 receptors were acutely blocked by alpha-helical CRH (alpha(h)CRH, CRH1/CRH2 antagonist; 0, 30, 100, and 300 ng) infusion into the lateral septum (LS), which abundantly expresses CRH2 but not CRH1 receptors. Freezing, locomotor activity, and analgesia were tested after infusion. Intra-LS alpha(h)CRH blocked shock-induced freezing without affecting activity or pain responses; infusions into lateral ventricle or nucleus of the diagonal band had no effects. The same behavioral profile was obtained with d-Phe-CRH((12-41)) (100 ng), another CRH1/CRH2 antagonist. A selective CRH1 antagonist (NBI27914), in doses that reduced freezing on intra-amygdala (central nucleus) infusion (0, 0.2, and 1.0 microg), did not affect freezing when infused into the LS. Ex vivo autoradiography revealed that binding of [125I]sauvagine, a mixed CRH1/CRH2 agonist, was prevented in the LS by previous intra-LS infusion of alpha(h)CRH but not NBI27914. In vitro studies demonstrated that [125I]sauvagine binding in the LS could be inhibited by a CRH1/CRH2 antagonist but not by the selective CRH1 receptor antagonist, confirming that in the LS, alpha(h)CRH antagonized exclusively CRH2 receptors. Acute antagonism of CRH2 receptors in the LS thus produces a behaviorally, anatomically, and pharmacologically specific reduction in stress-induced behavior, in contrast to results of recent knock-out studies, which induced congenital and permanent CRH2 removal. CRH2 receptors may thus represent a potential target for the development of novel CRH system anxiolytics.

Stress is defined as a state of threatened homeostasis. The principal effectors of the stress system include corticotropin-releasing hormone, arginine vasopressin, the glucocorticoids, and the catecholamines norepinephrine and epinephrine. Activation of the stress system leads to adaptive behavioral and physical changes. The principal stress hormones glucocorticoids and catecholamines affect major immune functions such as antigen presentation, leukocyte proliferation and traffic, secretion of cytokines and antibodies, and selection of the T helper (Th) 1 versus Th2 responses. A fully fledged systemic inflammatory reaction results in stimulation of the stress response, which in turn, through induction of a Th2 shift protects the organism from systemic overshooting with Th1/pro-inflammatory cytokines. Stress is often regarded as immunosuppressive, but recent evidence indicates that stress hormones influence the immune response in a less monochromatic way--systemically they inhibit Th1/pro-inflammatory responses and induce a Th2 shift, whereas in certain local responses they promote pro-inflammatory cytokine production and activation of the corticotropin-releasing hormone-mast cell-histamine axis. Through this mechanism a hyper- or hypoactive stress system associated with abnormalities of the systemic anti-inflammatory feedback and/or hyperactivity of the local pro-inflammatory factors may play a role in the pathogenesis of chronic inflammation and immune-related diseases.

Localized inflammation of a rat's hindpaw elicits an accumulation of beta-endorphin-(END) containing immune cells. We investigated the production, release, and antinociceptive effects of lymphocyte-derived END in relation to cell trafficking. In normal animals, END and proopiomelanocortin mRNA were less abundant in circulating lymphocytes than in those residing in lymph nodes (LN), suggesting that a finite cell population produces END and homes to LN. Inflammation increased proopiomelanocortin mRNA in cells from noninflamed and inflamed LN. However, END content was increased only in inflamed paw tissue and noninflamed LN-immune cells. Accordingly, corticotropin-releasing factor and IL-1beta released significantly more END from noninflamed than from inflamed LN-immune cells. This secretion was receptor specific, calcium dependent, and mimicked by potassium, consistent with vesicular release. Finally, both agents, injected into the inflamed paw, induced analgesia which was blocked by the co-administration of antiserum against END. Together, these findings suggest that END-producing lymphocytes home to inflamed tissue where they secrete END to reduce pain. Afterwards they migrate to the regional LN, depleted of the peptide. Consistent with this notion, immunofluorescence studies of cell suspensions revealed that END is contained predominantly within memory-type T cells. Thus, the immune system is important for the control of inflammatory pain. This has implications for the understanding of pain in immunosuppressed conditions like cancer or AIDS.

The major neuroendocrine response mediating stress adaptation is activation of the hypothalamic pituitary adrenal axis, with stimulation of corticotropin releasing hormone (CRH) and vasopressin (VP) from parvocellular neurons of the hypothalamic paraventricular nucleus, leading to stimulation of pituitary ACTH secretion and increases in glucocorticoid secretion from the adrenal cortex. Basal production and transient increases during stress of glucocorticoids and its hypothalamic regulators are essential for neuronal plasticity and normal brain function. While activation of the HPA axis is essential for survival during stress, chronic exposure to stress hormones can predispose to psychological, metabolic and immune alterations. Thus, prompt termination of the stress response is essential to prevent negative effects of inappropriate levels of CRH and glucocorticoids. This review addresses the regulation of HPA axis activity with emphasis on the mechanisms of termination of CRH transcription, which is a critical step in this process. In addition, the actions by which glucocorticoids, CRH and VP can affect the aging process will be discussed.

Relapse is a highly prevalent phenomenon in addiction. This paper examines the new research on identifying biological factors that contribute to addiction relapse risk. Prospective studies examining relapse risk are reviewed, and clinical, biological, and neural factors that predict relapse risk are identified. Clinical factors, patient-related factors, and subjective and behavioral measures such as depressive symptoms, stress, and drug craving all predict future relapse risk. Among biological measures, endocrine measures such as cortisol and cortisol/corticotropin (ACTH) ratio as a measure of adrenal sensitivity and serum brain-derived neurotrophic factor were also predictive of future relapse risk. Among neural measures, brain atrophy in the medial frontal regions and hyperreactivity of the anterior cingulate during withdrawal were identified as important in drug withdrawal and relapse risk. Caveats pertaining to specific drug abuse type and phase of addiction are discussed. Finally, significant implications of these findings for clinical practice are presented, with a specific focus on determining biological markers of relapse risk that may be used to identify those individuals who are most at risk of relapse in the clinic. Such markers may then be used to assess treatment response and develop specific treatments that will normalize these neural and biological sequelae so as to significantly improve relapse outcomes.

BACKGROUND

Suppression of pituitary-adrenal function is a well-known consequence of glucocorticoid therapy, manifested principally by decreased corticotropin secretion. To determine the degree of suppression of pituitary-adrenal function in patients treated with different doses of synthetic glucocorticoid medication for different periods, we measured the pituitary-adrenal response to the administration of exogenous human corticotropin-releasing hormone (CRH).

METHODS

We studied 279 patients who were receiving daily therapy with 5 to 30 mg of prednisone or its equivalent to treat various chronic diseases, principally collagen vascular disorders, and 50 normal subjects. Therapy ranged in duration from 1 week to 15 years. Stimulation tests using 100 micrograms of CRH as a bolus injection were performed in the morning, 24 hours after the most recent dose of glucocorticoids. In 61 patients an insulin hypoglycemia test, thought by many to be the reference standard, was also performed to assess the reliability of the CRH results.

RESULTS

After the administration of CRH, 43 patients had no increase in plasma concentrations of corticotropin and cortisol. The response was blunted in 133 patients and normal in 103. There was poor correlation between the plasma cortisol response after the administration of CRH and the dose or duration of therapy or the basal plasma cortisol concentration. Although plasma cortisol concentrations after stimulation with CRH were generally lower than those after insulin administration, there was a significant correlation between the plasma cortisol responses to the two stimuli (r = 0.82).

CONCLUSIONS

Pituitary-adrenal function in patients treated with synthetic glucocorticoids cannot be reliably estimated from the dose of glucocorticoid, the duration of therapy, or the basal plasma cortisol concentration. In such patients, testing with CRH is nearly as useful as insulin hypoglycemia testing in the assessment of pituitary-adrenal function.

Cushing's syndrome is caused by excess cortisol production from the adrenocortical gland. In corticotropin-independent Cushing's syndrome, the excess cortisol production is primarily attributed to an adrenocortical adenoma, in which the underlying molecular pathogenesis has been poorly understood. We report a hotspot mutation (L206R) in PRKACA, which encodes the catalytic subunit of cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA), in more than 50% of cases with adrenocortical adenomas associated with corticotropin-independent Cushing's syndrome. The L206R PRKACA mutant abolished its binding to the regulatory subunit of PKA (PRKAR1A) that inhibits catalytic activity of PRKACA, leading to constitutive, cAMP-independent PKA activation. These results highlight the major role of cAMP-independent activation of cAMP/PKA signaling by somatic mutations in corticotropin-independent Cushing's syndrome, providing insights into the diagnosis and therapeutics of this syndrome.

Anxiety disorders are frequently long-lasting and debilitating for more than 40 million American adults. Although stressor exposure plays an important role in the etiology of some anxiety disorders, the mechanisms by which exposure to stressful stimuli alters central circuits that mediate anxiety-like emotional behavior are still unknown. Substantial evidence has implicated regions of the central extended amygdala, including the bed nucleus of the stria terminalis (BNST) and the central nucleus of the amygdala as critical structures mediating fear- and anxiety-like behavior in both humans and animals. These areas organize coordinated fear- and anxiety-like behavioral responses as well as peripheral stress responding to threats via direct and indirect projections to the paraventricular nucleus of the hypothalamus and brainstem regions (Walker et al. Eur J Pharmacol 463:199-216, 2003, Prog Neuropsychopharmacol Biol Psychiatry 33(8):1291-1308, 2009; Ulrich-Lai and Herman Nat Rev Neurosci 10:397-409, 2009). In particular, the BNST has been argued to mediate these central and peripheral responses when the perceived threat is of long duration (Waddell et al. Behav Neurosci 120:324-336, 2006) and/or when the anxiety-like response is sustained (Walker and Davis Brain Struct Funct 213:29-42, 2008); hence, the BNST may mediate pathological anxiety-like states that result from exposure to chronic stress. Indeed, chronic stress paradigms result in enhanced BNST neuroplasticity that has been associated with pathological anxiety-like states (Vyas et al. Brain Res 965:290-294, 2003; Pego et al. Eur J Neurosci 27:1503-1516, 2008). Here we review evidence that suggests that pituitary adenylate cyclase-activating polypeptide (PACAP) and corticotropin-releasing hormone (CRH) work together to modulate BNST function and increase anxiety-like behavior. Moreover, we have shown that BNST PACAP as well as its cognate PAC1 receptor is substantially upregulated following chronic stress, particularly in the BNST oval nucleus where PACAP-containing neurons closely interact with CRH-containing neurons (Kozicz et al. Brain Res 767:109-119, 1997; Hammack et al. Psychoneuroendocrinology 34:833-843, 2009). We describe how interactions between PACAP and CRH in the BNST may mediate stress-associated behaviors, including anorexia and anxiety-like behavior. These studies have the potential to define specific mechanisms underlying anxiety disorders, and may provide important therapeutic strategies for stress and anxiety management.

In the adult castrated male rat, exposure to inescapable, intermittent electroshocks inhibited the pulsatile pattern of luteinizing hormone release and markedly lowered its plasma concentrations. The central administration of the corticotropin-releasing factor (CRF) antagonist alpha-helical ovine CRF residues 9 to 41 reversed the inhibitory action of stress. Neither its peripheral injection, nor the intraventricular injection of the inactive CRF analog des-Glu to Arg ovine CRF was effective. These results suggest that endogenous CRF may mediate some deleterious effects of noxious stimuli on reproduction.

Serotonin neurons play a major role in many normal and pathological brain functions. In the rat these neurons have a varying number of cotransmitters, including neuropeptides. Here we studied, with histochemical techniques, the relation between serotonin, some other small-molecule transmitters, and a number of neuropeptides in the dorsal raphe nucleus (DRN) and the adjacent ventral periaqueductal gray (vPAG) of mouse, an important question being to establish possible differences from rat. Even if similarly distributed, the serotonin neurons in mouse lacked the extensive coexpression of nitric oxide synthase and galanin seen in rat. Although partly overlapping in the vPAG, no evidence was obtained for the coexistence of serotonin with dopamine, substance P, cholecystokinin, enkephalin, somatostatin, neurotensin, dynorphin, thyrotropin-releasing hormone, or corticotropin-releasing hormone. However, some serotonin neurons expressed the gamma-aminobutyric acid (GABA)-synthesizing enzyme glutamic acid decarboxylase (GAD). Work in other laboratories suggests that, as in rat, serotonin neurons in the mouse midline DRN express the vesicular glutamate transporter 3, presumably releasing glutamate. Our study also shows that many of the neuropeptides studied (substance P, galanin, neurotensin, dynorphin, and corticotropin-releasing factor) are present in nerve terminal networks of varying densities close to the serotonin neurons, and therefore may directly or indirectly influence these cells. The apparently low numbers of coexisting messengers in mouse serotonin neurons, compared to rat, indicate considerable species differences with regard to the chemical neuronatomy of the DRN. Thus, extrapolation of DRN physiology, and possibly pathology, from rat to mouse, and even human, should be made with caution.

The hormonal response to stress is enhanced by oestrogen but inhibited by androgens. To determine underlying changes in activity of neuropeptide neurones in the paraventricular nucleus of the hypothalamus (PVN), we examined the effect of oestrogen and androgen treatment on restraint-induced c-fos mRNA, corticotropin-releasing hormone (CRH) heteronuclear RNA, and arginine vasopressin hnRNA expression in the PVN. Male rats were gonadectomized and injected with oestradiol benzoate (EB) or dihydrotestosterone propionate (DHTP; s.c., daily for 4 days). Rats were stressed by restraint for 10 min or 30 min before killing. Other rats were stressed for 30 min and then returned to their home cage for 20 min before killing. Corticosterone and adrenocorticotropic hormone responses to restraint stress were significantly greater in EB-treated rats and lower in DHTP-treated rats at the 30-min timepoint compared to controls. c-fos mRNA increases following stress were augmented by EB but inhibited by DHTP. CRH hnRNA expression increased significantly in the PVN in response to restraint stress, and this increase was augmented by EB treatment, but decreased by DHTP treatment. Vasopressin hnRNA expression was also increased in response to stress, and this increase was attenuated by DHTP. These findings indicate that gonadal hormones influence the reactivity of the hypothalamic-pituitary adrenal axis to stress.

In Alzheimer's disease (AD) there are dramatic reductions in the content of corticotropin releasing factor (CRF), reciprocal increases in CRF receptors, and morphological abnormalities in CRF neurons in affected brain areas. Cognitive impairment in AD patients is associated with a lower cerebrospinal fluid concentration of CRF, which is known to induce increases in learning and memory in rodents. This suggests that CRF deficits contribute to cognitive impairment. The identification in post-mortem brain of CRF-binding protein (CRF-BP), a high-affinity binding protein that inactivates CRF, and the differential distribution of CRF-BP and CRF receptors, provides the potential for improving learning and memory without stress effects of CRF receptor agonists. Here we show that ligands that dissociate CRF from CRF-BP increase brain levels of 'free CRF' in AD to control levels and show cognition-enhancing properties in models of learning and memory in animals without the characteristic stress effects of CRF receptor agonists.

BACKGROUND

Recent studies have reported a high comorbidity between posttraumatic stress disorder (PTSD) and psychotic symptoms, and it has been hypothesized that PTSD with comorbid psychosis is a severe form of PTSD. Few studies have examined the neurobiology of PTSD with comorbid psychosis. If PTSD with secondary psychotic symptoms (PTSD-SP) is a severe form of PTSD, then it might be expected to show more extreme perturbations in the neuroendocrine patterns that characterize PTSD.

METHODS

Patients with PTSD with secondary psychotic symptoms (PTSD-SP), PTSD without psychosis, and healthy comparison subjects were compared for differences in cerebrospinal fluid concentrations of corticotropin-releasing factor (CRF) and somatotropin-release-inhibiting hormone (SRIF).

RESULTS

The PTSD-SP subjects had significantly higher mean levels of CRF than either the PTSD or control subjects (p <.01). The three groups showed similar SRIF levels.

CONCLUSIONS

These data implicate abnormalities in the secretion of CRF with the production of secondary psychotic symptoms in PTSD. This finding supports the validity of PTSD-SP as a PTSD subtype and as a severe form of PTSD.

Intermittent electrical footshock induces c-fos expression in parvocellular neurosecretory neurons expressing corticotropin-releasing factor and in other visceromotor cell types of the paraventricular hypothalamic nucleus (PVH). Since catecholaminergic neurons of the nucleus of the solitary tract and ventrolateral medulla make up the dominant loci of footshock-responsive cells that project to the PVH, these were evaluated as candidate afferent mediators of hypothalamic neuroendocrine responses. Rats bearing discrete unilateral transections of this projection system were exposed to a single 30-min footshock session and sacrificed 2 hr later. Despite depletion of the aminergic innervation on the ipsilateral side, shock-induced up-regulation of Fos protein and corticotropin-releasing factor mRNA were comparable in strength and distribution in the PVH on both sides of the brain. This lesion did, however, result in a substantial reduction of Fos expression in medullary aminergic neurons on the ipsilateral side. These results contrast diametrically with those obtained in a systemic cytokine (interleukin 1) challenge paradigm, where similar cuts ablated the Fos response in the ipsilateral PVH but left intact the induction seen in the ipsilateral medulla. We conclude that (i) footshock-induced activation of medullary aminergic neurons is a secondary consequence of stress, mediated via a descending projection transected by our ablation, (ii) stress-induced activation of medullary aminergic neurons is not necessarily predictive of an involvement of these cell groups in driving hypothalamic visceromotor responses to a given stressor, and (iii) despite striking similarities in the complement of hypothalamic effector neurons and their afferents that may be activated by stresses of different types, distinct mechanisms may underlie adaptive hypothalamic responses in each.

The ontogeny of corticotropin-releasing hormone (CRH) receptor messenger ribonucleic acid (mRNA) in rat brain, using in situ hybridization, is the focus of this study. The developmental profile of CRH receptor using binding assays and receptor autoradiography has been reported, but may be confounded by the presence of a binding protein. The recent cloning of the rat CRH receptor gene has permitted the use of in situ hybridization histochemistry to map the distribution of cells expressing CRH receptor mRNA in the developing brain. We used antisense 35S-labeled oligodeoxynucleotide probes for the two reported splice-variants of the CRH receptor mRNA, which yielded essentially identical localization patterns. CRH receptor mRNA was clearly detectable in infant brain starting on the second postnatal day. Signal in hippocampal CA1, CA2 and CA3a increased to 300-600% of adult levels by postnatal day 6 with a subsequent gradual decline. In the amygdala, in contrast, CRH receptor mRNA abundance increased steadily between the second and the ninth postnatal days, to levels twice higher than those in the adult. In the cortex, CRH receptor mRNA levels were high on postnatal day 2 and decreased to adult levels by day 12. Transient signal over the hypothalamic paraventricular nucleus, observed on the second postnatal day, was not evident at older ages. These results demonstrate robust synthesis of CRH receptor as early as on the second postnatal day and unique region-specific developmental profiles for CRH receptor gene expression.

We investigated the effect of exogenous corticotropin-releasing factor on plasma levels of ACTH and cortisol in 13 patients with ACTH-secreting pituitary adenomas (Cushing's disease) and in 9 patients with other forms of Cushing's syndrome. In all patients with Cushing's disease, ovine corticotropin-releasing factor, given intravenously as a bolus injection (1 microgram per kilogram of body weight), caused a further increase in the already elevated levels of ACTH and cortisol. Successful transphenoidal adenomectomy was followed as early as one week after surgery by normalization or near-normalization of the ACTH and cortisol responses to corticotropin-releasing factor. On the other hand, patients with the ectopic ACTH syndrome, who also had high basal plasma concentrations of ACTH and cortisol, had no ACTH or cortisol responses to corticotropin-releasing factor. This difference in responsiveness between these two patient groups cannot be explained on the basis of different metabolic clearance rates of exogenous corticotropin-releasing factor, as shown by similar disappearance curves of immunoreactive corticotropin-releasing factor from plasma. Patients with Cushing's syndrome of adrenal origin who were hypercortisolemic during testing had undetectable plasma levels of ACTH and no ACTH or cortisol responses to corticotropin-releasing factor. We conclude that stimulation of the pituitary-adrenal axis with corticotropin-releasing factor may be useful in differentiating pituitary from ectopic causes of Cushing's syndrome.

Chandelier neurons are a unique subclass of cortical nonpyramidal neurons. The axons of these neurons terminate in distinctive vertically arrayed cartridges that synapse on the axon initial segment of pyramidal neurons. In this study, the rapid Golgi method and immunohistochemical techniques were used to characterize the morphology, regional distribution, laminar location, and biochemical content of chandelier neurons in the prefrontal and occipital cortices of three monkey species. As in our previous studies of visual areas V1 and V2 (Lund: Journal of Comparative Neurology 257:60-92, 1987; Lund et al.: Journal of Comparative Neurology 202:19-45, 1981, 276:1-29, 1988), Golgi impregnations of areas 46 and 9 of macaque prefrontal cortex show chandelier neurons to be present in layers 2 through superficial 5. The vertical arrays of terminal boutons (axon cartridges) typical of this neuron class are also present in layers 2-6 of the prefrontal cortex, but are not found in layer 1 or the subcortical white matter. In immunohistochemical studies, a calcium-binding protein, parvalbumin, and a neuropeptide, corticotropin-releasing factor (CRF), identify rod-like structures that are morphologically similar to the axon cartridges of chandelier neurons seen in the Golgi material. In addition, both parvalbumin- and CRF-immunoreactive cartridges are located below the somata of unlabeled pyramidal neurons and appear to outline the axon initial segment of these neurons. However, we find that parvalbumin and CRF are present in only subpopulations of chandelier axon cartridges. For example, in adult primary visual cortex, parvalbumin-labeled cartridges are present in very low numbers only in layers 2-3, whereas in prefrontal and occipital association cortices these cartridges are a very prominent component of layers 2-superficial 3 and are present in much lower density in the deeper cortical layers. In contrast to these findings in adult macaque monkeys, prefrontal and occipital association cortices of infant macaque monkeys contain a very high density of parvalbumin-labeled cartridges in layer 4 and relatively few in the superficial cortical layers. Furthermore, in adult squirrel monkey prefrontal cortex, CRF-labeled cartridges are predominately present in layer 4, but these CRF-immunoreactive structures have not been observed in the homologous regions of infant or adult macaque monkeys. These findings indicate that even for neurons of such distinctive morphology and presumably constant functional role as chandelier neurons, factors such as regional and laminar location, age, and primate species are associated with differences in the biochemical content of subpopulations of these neurons.

Inadequate pharmacological tools, until recently, hindered the understanding of the roles of corticotropin-releasing factor (CRF) receptor subtypes in appetite regulation and gastrocolonic motor function. Now, novel ligands that are selective for CRF(1) or CRF(2) receptors are helping to uncover the specific functions of CRF receptor subtypes. Central or peripheral CRF(2) receptor activation suppresses feeding independently of CRF(1) receptors. In the rat, central administration of CRF(2) receptor agonists promotes satiation without eliciting the malaise, behavioral arousal or anxiogenesis associated with CRF(1) receptor agonists. Conversely, central administration of CRF(1) receptor agonists elicits short-onset anorexia independently of CRF(2) receptor activation. With respect to gastrointestinal motor function, stress inhibits gastric motility through CRF(2) receptor-dependent central autonomic and peripheral myenteric systems. By contrast, stress stimulates colonic motility via CRF(1) receptor-dependent sacral parasympathetic and colonic myenteric mechanisms. These findings have important physiological implications and suggest targeted approaches for the pharmacotherapy of obesity and stress-related functional gastrointestinal and eating disorders.

The present studies were designed to evaluate the competitive binding properties and functional effects of a novel nonpeptide CRF1 receptor antagonist, R121919. R121919 administered in doses of 0.63 to 20 mg/kg p.o. 60 min pretest in Wistar rats dose dependently attenuated the swim stress-induced anxiogenic-like behavior in the elevated plus-maze model of anxiety. Moreover, receptor autoradiography revealed that R121919 dose-dependently occupied brain CRF1 receptors in subjects tested in the plus-maze experiment. Orally administered doses of up to 20 mg/kg R121919 also blunted basal and swim stress-induced pituitary-adrenocortical activation, produced additional anxiolytic-like behavioral actions in the defensive withdrawal and defensive burying paradigms, and functionally antagonized the locomotor stimulatory properties of exogenously administered CRF. Taken together, these results suggest that the anxiolytic-like efficacy of R121919 in attenuating the stress-, novelty-, shock-, and CRF-induced increases in behavioral arousal is correlated with competitive blockade of central CRF1 receptors.

BACKGROUND

Negative feedback regulation of the hypothalamic-pituitary-adrenal axis occurs through a dual-receptor system of mineralocorticoid receptors (MR) and glucocorticoid receptors (GR). Their affinity for cortisol and their distribution in the brain differ. Studies using an MR antagonist have demonstrated that MR is active throughout the circadian rhythm. Because major depression is accompanied by increased glucocorticoid secretion and insensitivity to glucocorticoid feedback, and because glucocorticoids are capable of down-regulating MR and GR, we expected that major depression would be accompanied by decreased MR activity.

METHODS

To test this hypothesis, we administered spironolactone, an MR antagonist, to individuals with major depression and matched control subjects and assessed levels of corticotropin and cortisol secretion in response to this acute challenge. Studies were conducted in the morning, the time of peak activation of the hypothalamic-pituitary-adrenal axis. All patients were currently depressed and free of all medications. All controls were free of all psychiatric diagnoses and of all medications.

RESULTS

Spironolactone treatment resulted in a significant increase in cortisol secretion levels in both groups. Depressed patients demonstrated higher cortisol secretion levels than control subjects. In addition, depressed patients demonstrated a different pattern of increase in cortisol secretion levels after spironolactone administration. Furthermore, a significant effect of spironolactone treatment on corticotropin secretion levels can be observed in depressed patients, whereas controls show no such effect.

CONCLUSIONS

Despite high baseline cortisol levels, patients with major depression show high functional activity of the MR system. Paired with the body of evidence regarding decreased sensitivity to GR agonists, these data suggest an imbalance in the MR/GR ratio. The balance of MR and GR is known to affect brain serotonin systems and may play an etiologic role in serotonin receptor changes observed in patients with major depression.

Stress is known to precipitate or worsen a number of disorders, such as migraines, in which mast cells are suspected of being involved by releasing vasoactive, nociceptive, and proinflammatory mediators. However, no functional association has been demonstrated yet between a migraine trigger and brain mast cell activation. Nontraumatic immobilization (restrain) stress has been shown to stimulate the hypothalamic-pituitary-adrenal axis and to cause redistribution of immune cells. Here, restrain stress caused degranulation in 70% of rat dura mast cells within 30 min, as shown both by light and electron microscopy. These morphologic findings were accompanied by cerebrospinal fluid elevation of rat mast cell protease I, but not II, indicating secretion from connective tissue type mast cells. Mast cell activation due to stress was abolished in animals that had been treated neonatally with capsaicin, indicating that neuropeptides in sensory nerve endings are involved in this response. Complete inhibition was also achieved by pretreating the animals ip with polyclonal antiserum to CRH. Mast cells in the dura were localized close to nerve processes containing substance P, but no CRH-positive fibers were identified even though these were found close to mast cells in the median eminence. This is the first time that stress is shown to activate intracranial mast cells; apparently through the sequential action of CRH and sensory neuropeptides. These findings may have implications for the pathophysiology and possible therapy of neuroinflammatory disorders such as migraines, which are induced or exacerbated by stress.

Addiction and stress are linked at multiple levels. Drug abuse is often initiated as a maladaptive mechanism for coping with stress. It is maintained in part by negative reinforcement to prevent the aversive consequences of stress associated with abstinence. Finally, stress is a major factor leading to relapse in subjects in which drug seeking behavior has extinguished. These associations imply overlapping or converging neural circuits and substrates that underlie the processes of addiction and the expression of the stress response. Here we discuss the major brain serotonin (5-HT) system, the dorsal raphe nucleus (DRN)-5-HT system as a point of convergence that links these processes and how the stress-related neuropeptide, corticotropin-releasing factor (CRF) directs this by a bimodal regulation of DRN neuronal activity. The review begins by describing a structural basis for CRF regulation of the DRN-5-HT system. This is followed by a review of the effects of CRF and stress on DRN function based on electrophysiological and microdialysis studies. The concept that multiple CRF receptor subtypes in the DRN facilitate distinct coping behaviors is reviewed with recent evidence for a unique cellular mechanism by which stress history can determine the type of coping behavior. Finally, work on CRF regulation of the DRN-5-HT system is integrated with literature on the role of 5-HT-dopamine interactions in addiction.

Dysregulation of the hypothalamic-pituitary-adrenal (HPA) axis in adults with major depressive disorder is among the most consistent and robust biological findings in psychiatry. Given the importance of the adolescent transition to the development and recurrence of depressive phenomena over the lifespan, it is important to have an integrative perspective on research investigating the various components of HPA axis functioning among depressed young people. The present narrative review synthesizes evidence from the following five categories of studies conducted with children and adolescents: (1) those examining the HPA system's response to the dexamethasone suppression test (DST); (2) those assessing basal HPA axis functioning; (3) those administering corticotropin-releasing hormone (CRH) challenge; (4) those incorporating psychological probes of the HPA axis; and (5) those examining HPA axis functioning in children of depressed mothers. Evidence is generally consistent with models of developmental psychopathology that hypothesize that atypical HPA axis functioning precedes the emergence of clinical levels of depression and that the HPA axis becomes increasingly dysregulated from child to adult manifestations of depression. Multidisciplinary approaches and longitudinal research designs that extend across development are needed to more clearly and usefully elucidate the role of the HPA axis in depression.