John P. Christianson

5-Hydroxytryptamine 2C Receptors in the Basolateral Amygdala Are Involved in the Expression of Anxiety After Uncontrollable Traumatic Stress

BACKGROUND Exposure to uncontrollable stressors often increases anxiety-like behavior in both humans and rodents. In rat, this effect depends on stress-induced activity within the dorsal raphe nucleus (DRN). However, the role of serotonin in DRN projection regions is largely unknown. The goals of this study were to 1) assess the effect of uncontrollable stress on extracellular serotonin in the basolateral amygdala during the anxiety test, 2) determine whether DRN activity during a poststress anxiety test is involved in anxiety-like behavior, and 3) determine the role of the serotonin 2C receptor (5-HT(2C)) in uncontrollable stress-induced anxiety. METHOD Rats were exposed to tail shocks that were either controllable or uncontrollable. On the following day, anxiety-like behavior was assessed in a Juvenile Social Exploration (JSE) test. Basolateral amygdala (BLA) extracellular serotonin concentrations were assessed during JSE by in vivo microdialysis 24 hours after uncontrollable stress, controllable stress, or no stress. In separate experiments, drugs were administered before the JSE test to inhibit the DRN or to block 5-HT(2C) receptors. RESULTS Exposure to uncontrollable shock reduced later social exploration. Prior uncontrollable stress potentiated serotonin efflux in the BLA during social exploration, but controllable stress did not. Intra-DRN 8-OH-DPAT and systemic and intra-BLA 5-HT(2C) receptor antagonist SB 242,084 prevented the expression of potentiated anxiety in uncontrollably stressed rats. Intra-BLA injection of the 5-HT(2C) agonist CP 809,101 mimicked the effect of stress. CONCLUSIONS These results suggest that the anxiety-like behavior observed after uncontrollable stress is mediated by exaggerated 5-HT acting at BLA 5-HT(2C) receptors.

Controllable versus uncontrollable stressors bi-directionally modulate conditioned but not innate fear

Fear conditioning and fear extinction play key roles in the development and treatment of anxiety-related disorders, yet there is little information concerning experiential variables that modulate these processes. Here we examined the impact of exposure to a stressor in a different environment on subsequent fear conditioning and extinction, and whether the degree of behavioral control that the subject has over the stressor is of importance. Rats received a session of either escapable (controllable) tail shock (ES), yoked inescapable (uncontrollable) tail shock (IS), or control treatment (home cage, HC) 7 days before fear conditioning in which a tone and foot shock were paired. Conditioning was measured 24 h later. In a second experiment rats received ES, IS or HC 24 h after contextual fear conditioning. Extinction then occurred every day beginning 7 days later until a criterion was reached. Spontaneous recovery of fear was assessed 14 days after extinction. IS potentiated fear conditioning when given before fear conditioning, and potentiated fear responding during extinction when given after conditioning. Importantly, ES potently interfered with later fear conditioning, decreased fear responding during fear extinction, and prevented spontaneous recovery of fear. Additionally, we examined if the activation of the ventral medial prefrontal cortex (mPFCv) by ES is critical for the protective effects of ES on later fear conditioning. Inactivation of the mPFCv with muscimol at the time of the initial experience with control prevented ES-induced reductions in later contextual and auditory fear conditioning. Finally, we explored if the protective effects of ES extended to an unconditioned fear stimulus, ferret odor. Unlike conditioned fear, prior ES increased the fear response to ferret odor to the same degree as did IS.

The role of prior stressor controllability and the dorsal raphé nucleus in sucrose preference and social exploration

Research investigating how control over stressors affects behavior often utilizes freezing and shuttle escape learning as the behavioral endpoints. These endpoints have been argued to reflect anxious or depressed states, but these descriptions are problematic. The present study sought to determine the impact of stressor controllability and the dorsal raphé nucleus (DRN) on sucrose preference and juvenile social exploration, putative measures of anhedonia and anxiety that are commonly used in studies of stress per se. In Experiment 1 rats were exposed to escapable stress (ES) or yoked-inescapable stress (IS) tailshocks. In Experiment 2 ES or IS was given 7 days before all rats received IS. In Experiment 3 the DRN was inactivated during IS by microinjection of 8-OH-DPAT. Sucrose preference and social exploration were tested for several days after stress. A fourth experiment confirmed that juvenile social exploration is sensitive to traditional beta-carboline and benzodiazepine manipulations. Both ES and IS reduced sucrose preference, but only IS reduced social exploration. Prior treatment with ES prevented the effect of IS on social exploration but did not prevent the effect of IS on sucrose preference and inactivation of the DRN prevented the effect of IS on social exploration but did not change sucrose preference. The present results indicate that social exploration but not sucrose preference is sensitive to prior stressor controllability, and that DRN activation mediates the effect of IS on social exploration. We argue that DRN-5-HT activation mediates a state of generalized anxiety produced by uncontrollable stress and that juvenile social exploration is a useful behavioral endpoint in stressor controllability studies.

Inhibition of Fear by Learned Safety Signals: A Mini-Symposium Review

Safety signals are learned cues that predict the nonoccurrence of an aversive event. As such, safety signals are potent inhibitors of fear and stress responses. Investigations of safety signal learning have increased over the last few years due in part to the finding that traumatized persons are unable to use safety cues to inhibit fear, making it a clinically relevant phenotype. The goal of this review is to present recent advances relating to the neural and behavioral mechanisms of safety learning, and expression in rodents, nonhuman primates, and humans.

Medial prefrontal cortical activation modulates the impact of controllable and uncontrollable stressor exposure on a social exploration test of anxiety in the rat

The presence of behavioral control over a stressor can blunt many of the effects of the stressor. We have recently reported that uncontrollable stress (inescapable electric tailshock, IS) reduces later social exploration of a juvenile whereas controllable stress (escapable shock, ES) does not. Activation of the ventral medial prefrontal cortex (vmPFC) is crucial to blunting the effects of IS on later escape behavior (learned helplessness). The goal of the current study was to test the role of the vmPFC in modulating the effects of stressor controllability on anxiety in the social exploration test. Thus, adult male rats were implanted with cannula guides for drug microinjection into the vmPFC. In Experiment 1, temporary inactivation of the vmPFC with the GABAA agonist muscimol before exposure to ES prevented the protective effects of stress control, leading to reduced social exploration. In Experiment 2, excitation of the vmPFC prior to IS with the GABA-activated Cl( − ) channel antagonist picrotoxin mimicked the stress resistance produced by control and prevented IS-induced reduction in social exploration. These results are consistent with prior work and identify the vmPFC as a critical component of the neural circuitry mediating the effects of stressor control on later behaviors. The relationship between the vmPFC, dorsal raphé nucleus, and other structures mediating stress-induced anxiety are discussed.

The sensory insular cortex mediates the stress-buffering effects of safety signals but not behavioral control

Safety signals are learned cues that predict stress-free periods whereas behavioral control is the ability to modify a stressor by behavioral actions. Both serve to attenuate the effects of stressors such as uncontrollable shocks. Internal and external cues produced by a controlling behavior are followed by a stressor-free interval, and so it is possible that safety learning is fundamental to the effect of control. If this is the case then behavioral control and safety should recruit the same neural machinery. Interestingly, safety signals that prevented a behavioral outcome of stressor exposure that is also blocked by control (reduced social exploration) failed to inhibit activity in the dorsal raphé nucleus or use the ventromedial prefrontal cortex, the mechanisms by which behavioral control operates. However, bilateral lesions to a region of posterior insular cortex, termed the “sensory insula,” prevented the effect of safety but not of behavioral control, providing a double-dissociation. These results indicate that stressor-modulators can recruit distinct neural circuitry and imply a critical role of the sensory insula in safety learning.

The protective effects of voluntary exercise against the behavioral consequences of uncontrollable stress persist despite an increase in anxiety following forced cessation of exercise

Humans who exercise are less likely to suffer from stress-related mood disorders. Similarly, rats allowed voluntary access to running wheels have constrained corticosterone responses to mild stressors and are protected against several behavioral consequences of uncontrollable stress which resemble symptoms of human anxiety and depression, including exaggerated fear and deficits in shuttle box escape learning. Although exercise conveys clear stress resistance, the duration of time the protective effects of exercise against the behavioral consequences of uncontrollable stress persist following exercise cessation is unknown. The current studies investigated (1) whether exercise-induced stress resistance extends to social avoidance, another anxiety-like behavior elicited by uncontrollable stressor exposure, and (2) the duration of time the protective effects of exercise persist following forced cessation of exercise. Six weeks of wheel running constrained the increase in corticosterone elicited by social exploration testing, and prevented the reduction in social exploration, exaggerated shock-elicited fear, and deficits in escape learning produced by uncontrollable stress. The protective effect of voluntary exercise against stress-induced interference with escape learning persisted for 15 days, but was lost by 25 days, following cessation of exercise. An anxiogenic effect, as revealed by a reduction in social exploration and an increase in fear behavior immerged as a function of time following cessation of exercise. Results demonstrate that the protective effect of voluntary exercise against the behavioral consequences of uncontrollable stress extends to include social avoidance, and can persist for several days following exercise cessation despite an increase in anxiety produced by forced cessation of exercise.

Control over Stress, But Not Stress Per Se Increases Prefrontal Cortical Pyramidal Neuron Excitability

Behavioral control over a stressful event reduces the negative consequences of not only that event, but also future stressful events. Plasticity in the prelimbic (PL) medial prefrontal cortex is critical to this process, but the nature of the changes induced is unknown. We used patch-clamp recording to measure the intrinsic excitability of PL pyramidal neurons in acute slices from rats exposed to either escapable stress (ES), for which rats had behavioral control over tail-shock termination, or inescapable stress (IS) without control. Shortly after exposure (2 h) to tail-shock stress, neurons in the ES group had larger action potential (AP) amplitude and faster AP rise rate, larger postspike afterdepolarization, and reduced membrane time constant. No significant effects of IS were observed. We developed a conductance-based computer model using the simulation tool NEURON. The computer model simulated the observed changes in the ES group with increases in Na+ conductance (gNa) and T-type Ca2+ conductance (gCaT). The empirical and computational results indicate that behavioral control over stress, but not stress itself, increases PL pyramidal neuron excitability by increasing intrinsic membrane excitability. It is proposed that plasticity of excitability is important to the behavioral effects of controllable stressor exposure.

5-hydroxytryptamine 2C receptors in the dorsal striatum mediate stress-induced interference with negatively reinforced instrumental escape behavior.

Uncontrollable stress can interfere with instrumental learning and induce anxiety in humans and rodents. While evidence supports a role for serotonin (5-HT) and serotonin 2C receptors (5-HT(2C)R) in the behavioral consequences of uncontrollable stress, the specific sites of action are unknown. These experiments sought to delineate the role of 5-HT and 5-HT(2C)R in the dorsal striatum (DS) and the lateral/basolateral amygdala (BLA) in the expression of stress-induced instrumental escape deficits and exaggerated fear, as these structures are critical to instrumental learning and fear behaviors. Using in vivo microdialysis, we first demonstrated that prior uncontrollable, but not controllable, stress sensitizes extracellular 5-HT in the dorsal striatum, a result that parallels prior work in the BLA. Additionally, rats were implanted with bi-lateral cannula in either the DS or the BLA and exposed to uncontrollable tail shock stress. One day later, rats were injected with 5-HT(2C)R antagonist (SB242084) and fear and instrumental learning behaviors were assessed in a shuttle box. Separately, groups of non-stressed rats received an intra-DS or an intra-BLA injection of the 5-HT(2C)R agonist (CP809101) and behavior was observed. Intra-DS injections of the 5-HT(2C)R antagonist prior to fear/escape tests completely blocked the stress-induced interference with instrumental escape learning; a partial block was observed when injections were in the BLA. Antagonist administration in either region did not influence stress-induced fear behavior. In the absence of prior stress, intra-DS administration of the 5-HT(2C)R agonist was sufficient to interfere with escape behavior without enhancing fear, while intra-BLA administration of the 5-HT(2C)R agonist increased fear behavior but had no effect on escape learning. Results reveal a novel role of the 5-HT(2C)R in the DS in the expression of instrumental escape deficits produced by uncontrollable stress and demonstrate that the involvement of 5-HT(2C)R activation in stress-induced behaviors is regionally specific.

Stress-protective neural circuits: not all roads lead through the prefrontal cortex

Abstract Exposure to an uncontrollable stressor elicits a constellation of physiological and behavioral sequel in laboratory rats that often reflect aspects of anxiety and other emotional disruptions. We review evidence suggesting that plasticity within the serotonergic dorsal raphe nucleus (DRN) is critical to the expression of uncontrollable stressor-induced anxiety. Specifically, after uncontrollable stressor exposure subsequent anxiogenic stimuli evoke greater 5-HT release in DRN terminal regions including the amygdala and striatum; and pharmacological blockade of postsynaptic 5-HT2C receptors in these regions prevents expression of stressor-induced anxiety. Importantly, the controllability of stress, the presence of safety signals, and a history of exercise mitigate the expression of stressor-induced anxiety. These stress-protective factors appear to involve distinct neural substrates; with stressor controllability requiring the medial prefrontal cortex, safety signals the insular cortex and exercise affecting the 5-HT system directly. Knowledge of the distinct yet converging mechanisms underlying these stress-protective factors could provide insight into novel strategies for the treatment and prevention of stress-related psychiatric disorders.