Corticotropin-releasing factor modulates defensive-withdrawal and exploratory behavior in rats

The role of corticotropin-releasing factor (CRF), an endogenous neuropeptide, in modulating species-typical responses was examined in an unfamiliar open field containing a small chamber. Rats placed in this small chamber spent most of their time withdrawn in it. However, rats given an intracerebroventricular injection (20 micrograms) of alpha-helical CRF(9-41), a CRF receptor antagonist, emerged from the chamber and explored the unfamiliar open field. Additional studies showed that after 1 exposure to the test environment, vehicle-treated rats increased their time spent in the open field and returned intermittently to the chamber. This result suggests that reexposure reduces the threatening impact of an unfamiliar open field. Importantly, CRF (300 ng) injected centrally, but not peripherally, before reexposure to the test environment significantly reduced exploration in the open field and increased a pattern of defensive-withdrawal into the chamber. Data suggest that whether defensive-withdrawal or exploratory behavior is exhibited may depend on CRF actions in brain systems that mediate the perception of threat in the environment.     

Rat's responses to blood and body odors of stressed and non-stressed conspecifics

Male rats were tested in a Y-maze for exploratory responses to odors from stressed and non-stressed rats, and from the blood of stressed and non-stressed rats. Significant differences in activity but not preference were obtained in rats receiving odors from stressed and non-stressed rats. Odors from rats given 5 random, signalled, 2 sec electric shocks over 15 min, significantly increased the activity of receivers, compared to responses elicited by odors from non-stressed animals (Experiment 1). Odors from rats given 12 random, signalled, 2 sec electric shocks over 35 min significantly reduced receivers' activity (Experiment 2). Interestingly, responses of rats to odors from the blood of stressed and non-stressed rats were similar to their responses to odors from stressed and non-stressed rats. It is concluded that responses to these odors depend upon the stressor applied to the odor producing rats. Rats subjected to different stress conditions may change the quality or concentration of the odors they emit and thereby alter the responses of rats receiving the odors.     

Corticotropin-releasing factor receptor antagonist infused into the locus coeruleus attenuates immobilization stress-induced defensive withdrawal in rats

It has been proposed that corticotropin-releasing factor (CRF) released during stress in the region of the locus coeruleus (LC) induces changes in behavior that are typical indices of anxiety. The experiments tested the ability of a CRF antagonist, hCRF_9–41, to attenuate stress-induced defensive withdrawal in rats. 1 μg of hCRF in 300 nl was infused bilaterally in the LC of rats 10 min prior to 30 min immobilization. The apparatus consisted of a small chamber set on one side of a 1 m open field, into which the rat was placed to start the test. Restraint induced defensive withdrawal in rats familiar with the apparatus and significantly increased latency time to emerge from the chamber, total time and mean time spent in the chamber. Infusion of hCRF into the LC prior to restraint significantly decreased total and mean time spent in the chamber comparing to stressed animals. These results are consistent with anatomical, electrophysiological and neurochemical evidence that CRF receptors located in, or close to, the LC region influence behaviors induced by stress.     

The effects of odors from stressed mice on conspecific behavior

Four experiments correlate conspecific reactions to odors from stressed (foot shocked) BALB/cJ mice with the frequency of specific motor activities and taste avoidance. Where behavior was restricted to forward and backward movement in a tube, animals tended to avoid the side where the odors from stressed animals entered. In a more socially complex home cage (3 recipients) a wide variety of behaviors were affected by odors from stressed conspecifics. Animals were alerted by the odor, searched out the source of the odor and showed increases in general activity, rearing, and air sampling. Many of these behaviors habituated with continuous exposure. The major response to odors from stressed animals was to increase "vigilance." A restriction of behavioral opportunities will lead to odor avoidance; however, when the environment permits, the behavioral reaction to odors becomes more complex. These odors failed to produce conditioned taste aversion, suggesting a sensory specificity in the use of these odors.     

Intracerebroventricular administration of corticotropin-releasing factor antagonist attenuates arousal response accompanied by yawning behavior in rats

We have reported that an arousal response accompanied by yawning behavior can be evoked by electrical and chemical stimulation of the hypothalamic paraventricular nucleus (PVN) in rats, although the mechanism responsible for the arousal response accompanied by yawning evoked by PVN stimulation is still unknown. In the present study, we examined the involvement of corticotropin-releasing factor (CRF) in the arousal response during yawning induced by electrical stimulation of the PVN in anesthetized, spontaneous breathing rats using intracerebroventricular (icv) injection of alpha-helical CRF, a CRF antagonist (4.2 microg, lateral ventricle). The electrocorticogram (ECoG) was recorded to evaluate arousal responses during yawning. Fast Fourier transform was used to obtain the power spectrum in delta (0.5-4 Hz), theta (4-8 Hz), alpha (8-13 Hz), and beta (13-20 Hz) bands. We also recorded the intercostal electromyogram as an index of inspiratory activity and blood pressure (BP) as an index of autonomic function to evaluate yawning response. PVN stimulation induced significant increases in relative powers of theta, alpha, and beta bands, but not delta band, concurrent with yawning events regardless of icv injection, though the relative powers after icv injection of alpha-helical CRF were significantly lower than those after saline injection. These findings suggest that CRF neurons in the PVN are primarily responsible for the arousal response accompanied by yawning behavior.     

Discrimination of odors from stressed rats by non-stressed rats

Comparison of earlier reports of rat stress odors is complicated by the many differences in experimental parameters and responses measured. To evaluate whether these stress odors provide a special signal, rats were subjected to different levels of stressful foot-shock in one half of a simple two-compartment test box whilst the other half was clean and unoccupied. The results show that whilst test subjects preferred the half containing odors from non-stressed rats, this preference was decreased by the presence of stress odors to an extent concordant with the level of stressor applied to the odor donors. There were no differences in plasma corticosterone among the odor donors indicating that this hormone is probably not the source of stress odors. Plasma corticosterone levels of the subjects were similar to each other and to the odor donors. Compared to odors from non-stressed rats, stress odors increased the activity of the subjects. The evidence strongly suggests a special signalling function for stress odors although responses to this signal are not stereotyped.     

The sources of odors from stressed rats

Recent studies [8,9] have shown that odors from stressed Norway rats act as signals to which other rats respond primarily by overall changes in activity and exploration. At present the source of these odors is unknown. In this study odors from urine, feces and the bodies of stressed rats were delivered along a runway in which the subjects had been previously trained to run for a water reward in the presence of odors from non-stressed rats. The results indicate that odors are released from the body surface and in the urine but not the feces of stressed rats.     

Corticotropin-releasing factor antagonist reduces activation of noradrenalin and serotonin neurons in the locus coeruleus and dorsal raphe in the arousal response accompanied by yawning behavior in rats

We previously reported that intracerebroventricular (icv) administration of corticotropin-releasing factor (CRF) antagonist attenuates the arousal response during yawning behavior in rats. However, the CRF-related pathway involved in the arousal response during yawning is still unclear. In the present study, we assessed the involvement of the CRF-containing pathway from the hypothalamic paraventricular nucleus (PVN) to the locus coeruleus (LC) and the dorsal raphe nucleus (DRN) in the arousal response during frequent spontaneous yawning, which was induced by several microinjections of l-glutamate into the PVN in anesthetized rats, using c-Fos immunohistochemistry. The PVN stimulation showed significant increases in activation of PVN CRF neurons, LC noradrenalin (NA) neurons and DRN serotonin (5-HT) neurons as well as arousal response during yawning. But icv administration of a CRF receptor antagonist, α-helical CRF (9-41), significantly inhibited the activation of both LC NA neurons and DRN 5-HT neurons except the activation of CRF neurons in the PVN, and significantly suppressed the arousal response during yawning. These results suggest that the CRF-containing pathway from PVN CRF neurons to LC NA neurons and DRN 5-HT neurons can be involved in the arousal response during yawning behavior.     

Psychophysiological treatment outcomes: Corticotropin-releasing factor type 1 receptor antagonist increases inhibition of fear-potentiated startle in PTSD patients

After exposure to a traumatic event, a subset of people develop post-traumatic stress disorder (PTSD). One of the key deficits in PTSD is regulation of fear, and impaired inhibition of fear-potentiated startle (FPS) has been identified as a potential physiological biomarker specific to PTSD. As part of a larger clinical trial, this study investigated the effects of a CRF receptor 1 antagonist, GSK561679, on inhibition of fear-potentiated startle during a conditional discrimination fear-conditioning paradigm, termed AX+/BX−. Prior research using this paradigm has demonstrated deficits in inhibition of conditioned fear in several PTSD populations. The randomized, double-blind, placebo-controlled clinical trial compared fear inhibition between female PTSD participants taking 350 mg/day GSK561679 (n = 47 pre- and 29 post-treatment) and patients taking a placebo pill (n = 52 pre- and 30 post-treatment) daily for 6 weeks. There was no significant difference between the two groups in their acquisition of fear or discrimination between threat and safety cues, and no pre–post-treatment effect on these measures. However, there was a significant effect of treatment on inhibition of FPS during the AB trials in the AX+/BX− transfer test (p < 0.05). While all PTSD participants showed typical impairments in fear inhibition prior to treatment, GSK561679 enhanced fear inhibition post-treatment, independent of clinical effects. The current study suggests that CRF receptor 1 antagonism may have specific effects within neural circuitry mediating fear inhibition responses, but not overall symptom presentation, in PTSD.     

Attraction of newborn rabbits to abdominal odors of adult conspecifics differing in sex and physiological state

The present study aimed at assessing the ability of newborn rabbits (1-3 days) to detect and discriminate abdominal odors emitted by adult conspecifics varying in sex and physiological state. Pups were submitted to a two-choice test exclusively based on olfaction, which permitted exposure to conspecifics either during successive or simultaneous presentations. Their orientation toward either of the two stimuli was timed. These behavioral assays revealed that (a) pups display attraction to odors of the abdomen of nonlactating, nonpregnant females and males when pitted against a control stimulus; (b) pups preferred the odor of nonlactating, nonpregnant females when simultaneously presented with the odor of a male; (c) all sexually mature females (virgin, pregnant, or lactating) were attractive to pups; (d) the odor of lactating females was preferred to the odor of either nonlactating, nonpregnant or pregnant females. Thus, from the first day after birth, newborn rabbits are able to olfactorily discriminate conspecifics from cues emitted at the abdominal level and display the strongest attraction to lactating females.     

The NMDA receptor antagonist MK-801 alters lipoprivic eating elicited by 2-mercaptoacetate

Eating behavior is controlled, at least in part, by levels of circulating metabolic fuels such as glucose and free fatty acids, and drugs that interfere with the availability of these fuels can elicit eating. One such drug is 2-mercaptoacetate (2MA), an inhibitor of fatty acid oxidation. Evidence also suggests that NMDA receptors may mediate some aspects of normal eating and satiety. The present study was conducted in order to determine whether NMDA receptors may play a role in feeding elicited by 2MA. Rats received intraperitoneal injections of either saline, 2MA, the non-competitive NMDA receptor antagonist MK-801 or a combined injection of 2MA and MK-801, and subsequent intake of a fat-enriched, mash diet was measured at 1, 2, 3 and 4 h post-injection. Results showed that cumulative food intake was significantly increased by 2MA alone, as compared to saline controls, with most of the 2MA-elicited eating occurring during the first hour post-injection. While MK-801 alone did not alter food intake, it did have a biphasic effect on feeding elicited by 2MA. MK-801 initially suppressed and later enhanced eating elicited by 2MA. Although it is unclear whether MK-801 is acting centrally, peripherally or both to alter 2MA-induced eating, these results implicate NMDA receptors and the neurotransmitter glutamate in the regulation of lipid-associated eating and satiety.     

Changes in the feeding behavior of rats elicited by histamine infusion

In this study, we examined the effect of a putative neurotransmitter or a neuromodulator histamine (HA) on the feeding behavior to elucidate its physiological function in the central nervous system. Rats were implanted with a cannula into the suprachiasmatic nucleus through which HA was continuously infused for 200 hours with an Alzet osmotic minipump. The food intake was recorded automatically. This infusion resulted in decrease in food intake during the dark period and increase in it during the light period which contributed to the decrease in total food intake and increase in the percentage of food intake during the light period. Percentage of food intake during the light period is a good index of the amplitude of the circadian rhythm. Presumably, HA is concerned not only in the meal size, but also in the chronological aspect of the feeding behavior. The administration of H1-antagonist, pyrilamine, antagonized the HA induced increase in food intake during the light period. These findings suggest that continuous infusion of HA affected the feeding behavior which is possibly mediated through the H1-receptors in rat brain.     

Inactivation of the periaqueductal gray attenuates antinociception elicited by stimulation of the rat medial preoptic area

The medial preoptic area (MPOA) is a sexually dimorphic structure that plays key roles in gonado-steroidal regulation and thermoregulation. The MPOA may be involved in sex-based differences in nociceptive processing and steroid hormones effect on pain thresholds. Consistent with this, there is evidence that MPOA can produce antinociception or hyperalgesia. MPOA stimulation inhibits spinal cord or trigeminal neuronal responses to noxious stimuli or produces analgesia, yet most of these studies utilized electrical stimulation which antidromically activates periaqueductal gray (PAG) and rostroventromedial medulla (RVM) neurons involved in descending modulation of nociception. Effects of selective activation of MPOA neurons on behavioral indices of antinociception and the site-specificity of such responses are unknown. To address these questions, we examined the influence of MPOA microinjections of d,l homocysteate (DLH) on hindlimb and tail nocifensive reflexes in lightly anesthetized rats. DLH, but not saline, microinjections into several MPOA subregions markedly increased withdrawal response latencies to noxious thermal stimuli. Antinociceptive effects of MPOA activation were abolished by microinjection of lidocaine into PAG. These results suggest that activation of MPOA neurons produces antinociception that is at least partly mediated by projections to PAG.     

Early behavior of optokinetic responses elicited by transparent motion stimuli during depth-based attention

When two visual patterns moving in different directions are superimposed on the same depth plane (transparent motion stimulus), observers perceive transparent surfaces sliding over each other on different depth planes. Simultaneously, an optokinetic response (OKR) occurs so that one of the visual patterns is stabilized on the retina. In this study, we investigated the early behavior of the OKR elicited by transparent motion stimuli while subjects focused their attention on either the near or far surface. Two random dot patterns were superimposed and moved in orthogonal or opposite directions. Subjects were instructed to report the motion direction of the surface on which their attention was focused. The mean latency of initiation of OKR in the case of motion in opposite directions (150 ms) was significantly longer than that in the case of motion in orthogonal directions (100 ms). In the case of motion in orthogonal directions, the distribution of directions of OKR during the initial period, from 100 to 150 ms, was biased toward the mean direction of the two stimulus motions. After 160 ms, the eyes started to pursue a particular motion pattern of which the direction agreed with the far-perceived motion regardless of depth-based attention. Depth-based attention changed the direction of eye movements after 200 ms and eventually made the eyes follow a pattern on which the attention was focused. These results suggest that pursuit eye movement immediately after 160 ms may determine perceptual depth order through change of retinal image motion, because the slow-moving retinal image may be perceived in the far depth plane. Following this process of determination of perceptual depth order, depth-based attention starts to affect OKR.     

Corticotropin-releasing factor in the dorsal raphe elicits temporally distinct serotonergic responses in the limbic system in relation to fear behavior

The neurotransmitters serotonin and corticotrophin-releasing factor are thought to play an important role in fear and anxiety behaviors. This study aimed to determine the relationship between corticotrophin-releasing factor-evoked changes in serotonin levels within discrete regions of the limbic system and the expression of fear behavior in rats. The effects of corticotrophin-releasing factor administration to the serotonin cell body regions of the dorsal raphe nucleus on fear behavior, behavioral activity, and extracellular serotonin levels were assessed in freely moving rats with microdialysis probes implanted into the central nucleus of the amygdala and the medial prefrontal cortex. Infusion of corticotrophin-releasing factor (0.5 microg) into the dorsal raphe rapidly induced freezing behavior, which was positively correlated with an immediate increase in serotonin release in the central nucleus of the amygdala. In contrast, cessation of freezing behavior correlated with a delayed and prolonged increase in serotonin release within the medial prefrontal cortex. Our findings suggest that corticotrophin-releasing factor-induced freezing behavior is associated with regionally and temporally distinct serotonergic responses in the limbic system that may reflect differing roles for these regions in the expression of fear/anxiety behavior.     

Feeding elicited by injection of the cholecystokinin antagonist dibutyryl cyclic GMP into the cerebral ventricles of sheep

Dibutyryl cyclic GMP (Bt₂cGMP) is a specific competitive antagonist of the action of cholecystokinin (CCK) on dispersed pancreatic acini and on gall bladder smooth muscle, and it exerts its effects at the CCK receptor level. CCK peptides in brain are proposed to have a role in satiety, and because (1) brain CCK receptors have characteristics similar to those of peripheral CCK receptors, and (2) injections of CCK-octapeptide (CCK-OP) into the cerebrospinal fluid (CSF) of sheep decreased feeding, we tested the ability of Bt₂cGMP to inhibit satiety when injected into the cerebral ventricles of sheep. Feed intakes were increased by more than 150% during the first hour of a two hour continuous injection of 2.9 nmole/min Bt₂cGMP (N=20, p<0.05). Although 29 and 290 nmoles/min Bt₂cGMP caused agitation and no increase in feeding, only normal behaviors were observed with the 2.9 nmole/min dose. A lower dose (0.72 nmole/min) had no effect on behavior or feed intake. The increase in feed intake caused by injection of 2.9 nmole/min Bt₂cGMP into the cerebral ventricles suggests that this nucleotide, acting as an antagonist of CNS CCK receptors, inhibited satiety and thus elicited feeding. These results, in conjunction with those demonstrating an increase in feeding following the binding of CCK in CSF by CCK antibody, support the hypothesis that brain CCK is released during feeding and acts on CCK receptors which are involved in the control of feed intake.     

Corticotropin-releasing hormone receptor blockade fails to alter stress-evoked catecholamine release in prefrontal cortex of control or chronically stressed rats

Although it is well documented that stress can increase the activity of central dopamine and norepinephrine neurons, little is known about the role of other neurotransmitters in modulating this response. Previous studies have implicated corticotropin-releasing hormone in modulating stress-evoked changes in the activity of locus coeruleus neurons. The present study examines whether corticotropin-releasing hormone contributes to stress-evoked increases in extracellular norepinephrine and dopamine in rat medial prefrontal cortex, as monitored by in vivo microdialysis. As noted previously, 30 min of tail-shock increased extracellular levels of norepinephrine and dopamine in the medial prefrontal cortex of na?ve rats, and this was enhanced in rats previously exposed to chronic cold ( approximately 5 degrees C for 2-3 weeks). Previous intraventricular administration of a corticotropin-releasing hormone antagonist (D-Phe-corticotropin-releasing hormone; 3 and 9 microg) did not alter the tail-shock evoked in increase in extracellular levels of norepinephrine and dopamine in either na?ve or chronically cold-exposed rats. Intraventricular administration of 3 microg of D-Phe-corticotropin-releasing hormone attenuated the increase in extracellular norepinephrine induced by co-administration of 3 microg of corticotropin-releasing hormone, confirming the efficacy of this compound. Results of the present study suggest that endogenous corticotropin-releasing hormone does not play a role in modulating the release of norepinephrine and dopamine occurring in response to acute tail-shock or the expression of a potentiated response to tail-shock in rats exposed chronically to cold.     

An analysis of the behavior elicited by stimulation of the dorsal pons in rat

Dorsal pontine intracranial self-stimulation (ICSS) sites were electrically stimulated in rats. Only stimulation-induced oral behavior and not locomotor behavior was observed. The predominant elicited behavior was a fragmentary type of forepaw grooming. In some cases the stimulation-induced forepaw grooming was changed to drinking by the forced presentation of a drinking spout in the perioral region. Neither the dorsal pontine stimulation-induced oral behavior nor the ICSS was disrupted by six-hydroxydopamine lesions of ascending noradrenergic projections from locus coeruleus to widespread forebrain areas. It is suggested that the mesencephalic nucleus of the trigeminal nerve, with its connections to the motor trigeminal nucleus, may mediate dorsal pontine stimulation-induced behavior and ICSS.     

Broad-spectrum immunity against superantigens is elicited in mice protected from lethal shock by a superantigen antagonist peptide

Bypassing the restricted presentation of conventional antigens, superantigens trigger an excessive cellular immune response leading to toxic shock. Antagonist peptides that inhibit the induction of human Th1 cytokine gene expression by a variety of bacterial superantigens protect mice from lethal toxic shock. We show that the surviving mice rapidly develop a broad-spectrum, protective immunity against further lethal toxin challenges with the same superantigen and even with superantigen toxins that they have not encountered before. By blocking the induction of a cellular immune response leading to toxic shock, the antagonist peptide allows the superantigen to induce a vigorous humoral immune response directed against itself, resulting in anti-toxin IgM and IgG that are broadly protective. Adoptive transfer of these antibodies to na?ve mice rendered them resistant to lethal superantigen challenge. The appearance of these antibodies does not require immunization with an adjuvant and is not elicited by the antagonist peptide. Our results show that superantigens are potent immunogens when given the opportunity to induce a B cell response, in conditions where a deleterious Th1 response is prevented by the superantigen antagonist peptide.