Ventral and dorsolateral regions of the midbrain periaqueductal gray (PAG) control different stages of defensive behavior: Dorsolateral PAG lesions enhance the defensive freezing produced by massed and immediate shock

Rats that receive nociceptive electric shock in an environment normally show the conditional fear-induced defensive response of freezing when returned to that environment. If several electric shocks are given in a massed manner they will condition less freezing than the same shocks given in a distributed manner. If a single shock is given immediately after placement in the chamber it does not support any conditioning, although the same shock given after a brief delay does. Electrolytic lesions of the dorsolateral periaqueductal gray (PAG), which damaged dorsomedial, dorsolateral, and lateral PAG, enhanced freezing under these conditions. Lesions of the ventral PAG, which caused extensive damage to the central gray below the aqueduct, reduced conditioning under the more optimal parameters (distributed or delayed shock). This was taken to indicate that both of these regions support different modes of defensive behavior and that when activated, the dorsolateral PAG inbits conditional fear-induced defensive behavior. © 1995 Wiley-Liss, Inc.     

The effects of naloxone administered into the periaqueductal gray on shock-elicited freezing behavior in the rat

Freezing behavior that occurs following footshock was found to increase in rats in which naloxone was injected into the ventrolateral region of the mesencephalic periaqueductal gray (PAG) area of the brain prior to footshock administration. Since naloxone administered into the ventrolateral region of the PAG induced minimal freezing in rats which did not receive footshock, the results suggest that the effect of naloxone on shock-induced freezing is not due to a nonspecific decrease in motor activity. Naloxone had no effect on freezing when injected into the dorsolateral region of the PAG. The data are consistent with the theory that conditioned fear induces opioid mediated analgesia, and that the ventrolateral region of the PAG is an important component of a pain-inhibitory system involved in this analgesia.     

Lesions of the ventral hippocampus, but not the dorsal hippocampus, impair conditioned fear expression and inhibitory avoidance on the elevated T-maze

The hippocampus has been suggested to be involved in spatial (or configural) memory and also in the inhibition of certain response or goal alternatives. An increasing number of anatomical, physiological, and behavioral studies indicate that the hippocampus is functionally heterogeneous along the dorsal-ventral axis. Identification of distinct behavioral roles for the dorsal (DH) and ventral (VH) hippocampus may resolve differences between the various theoretical accounts of hippocampal function. The present study examined the effects of electrolytic lesions restricted to the DH or VH on fear-conditioned freezing, passive avoidance on the elevated T-maze (ETM) test of anxiety, and general activity in male Sprague-Dawley (Charles-River derived) albino rats. We found that rats with lesions of the VH, but not DH showed reduced freezing to both context and tone conditioned stimuli (CS). Rats with VH lesions also showed a reduced latency to emerge from the enclosed arm on trials 2 and 3 of the ETM (indicating reduced anxious behavior), while having no effect on the latency to escape from the open arms on trial 4. There were no differences in activity between the groups. These results indicate that the VH and DH are differentially involved in passive avoidance on the ETM and conditioned freezing to context and tone CS. We suggest that the VH may be specifically involved in modulating goal-oriented, defensive behavior expression through hypothalamic and amygdaloid connections.     

One-trial conditioned suppression: effects of instructions on extinction

Two experimental groups of undergraduate volunteers received a single Pavlovian conditioning trial consisting of a paired presentation of a tone conditioned stimulus (CS) and a shock unconditioned stimulus (UCS). Control groups received either the CS alone or the UCS alone. Subjects from one experimental group were subsequently instructed that they would not receive further shocks, while the other experimental group received no such instructions. The CS alone was then presented once to all four groups while subjects were engaged in a button-pressing task maintained by slide reinforcement. During the latter phase, rate of button-pressing was measured. Classically conditioned suppression of button-pressing was obtained in the noninstructed experimental group but not in the instructed group. The results demonstrate that suppression can be a sensitive index of Pavlovian conditioning in humans but question the use of conditioned suppression as an adequate experimental analog of clinically observed anxiety-motivated behavior.     

US habituation,like CS extinction,produces a decrement in conditioned fear responding that is NMDA dependent and subject to renewal and reinstatement

Just as fear can be learned, it can also be inhibited. The most common way of reducing learned fear is through extinction, where the conditioned stimulus (CS) previously paired with an aversive unconditioned stimulus (US) is repeatedly presented on its own. Another, much less commonly studied, way to inhibit learned fear is by habituating, or devaluing, the US. In this procedure, fear responding to a CS is reduced by repeatedly presenting the US in the absence of the CS following the conditioning phase. The purpose of the present study was to directly compare the effects of US habituation and CS extinction on a learned fear response (freezing). Experiment 1 demonstrated that US habituation given either after (Experiment 1A) or before (Experiment 1B) fear conditioning reduced freezing to the CS at test. We then showed that the reduction in freezing resulting from either US habituation or CS extinction was context-specific (i.e., a change in context led to a renewal of the learned fear response; Experiment 2) and, furthermore, was attenuated when a pre-test shock was given (i.e., reinstatement of fear was observed in both cases; Experiment 3). Finally, Experiment 4 demonstrated that an injection of the NMDA antagonist MK-801 prior to US habituation impaired long-term retention of the learning that takes place during this procedure. Together, these results suggest that the decrement in conditioned fear responses produced by US habituation and CS extinction could rely on overlapping processes.     

Footshock intensity and generalization in contextual and auditory-cued fear conditioning in the rat

The relationship between US (footshock) intensity and the two conditioned freezing responses (to acoustic CS and to "context") was investigated in fear conditioning. Administered footshock intensity was 0.00, 0.15, 0.30, 0.60, 0.90, and 1.20 mA to six different groups of 70-day-old male Albino Wistar rats. To measure contextual freezing, the animals were again placed inside the conditioning apparatus without acoustic CS and US presentation. To measure acoustic CS freezing, the animals were placed in a totally different apparatus and only the acoustic CS was presented. The 0.15 mA footshock intensity was not sufficient to condition the animals, in fact no freezing was exhibited as in the non-shocked control group. The 0.30 mA footshock intensity was sufficient only to condition the animals to the acoustic CS, whereas the 0.60 mA was sufficient to condition the animals both to acoustic CS and to context. Footshock intensities (0.90 and 1.20 mA) did not elicit any significant increase in conditioned freezing for either acoustic CS or context but at the highest one the generalization phenomenon appeared (freezing in the different context before presentation of acoustic CS). Acoustic CS freezing to all over-threshold intensities was longer than that to context. In conclusion, freezing responses to acoustic CS and context after increasing footshock intensities follow distinct patterns, and intermediate footshock intensities (0.60 and 0.90 mA) appear to be the most useful for eliciting conditioned freezing responses to acoustic CS and to context without inducing a generalized fear status contamination.     

Transfer across unconditioned stimuli in serial feature discrimination training

The transfer of conditioned modulation across conditioned stimuli (CS) and unconditioned stimuli (US) was examined in 3 experiments that used Pavlovian appetitive training procedures with rats. In Experiment 1, after training in a positive patterning discrimination (X-->A+/X-/A-), X increased conditioned responding elicited by another trained-then-extinguished CS as long as that CS had been trained with the same US as was used in discrimination training. In Experiment 2, after training with a feature-negative discrimination (X-->A-/A+), X inhibited conditioned responding elicited by another trained-then-extinguished CS as long as that CS had been trained with the same US. Experiments 1 and 2 used a between-groups design, whereas Experiment 3 used a more powerful within-groups design. In Experiment 3, rats were trained in a feature-positive discrimination (X-->A+/A-). In transfer tests, X increased conditioned responding elicited by another CS trained then extinguished with the same US from training. This increase was greater than the X increased conditioned responding elicited by another CS trained then extinguished with a different US from training. The results supported the suggestion that features trained in serial discrimination tasks influence behavior indirectly by transiently raising or lowering the threshold for activation of the US representations by its target stimuli and by any other stimuli that may be associated with that US. Other interpretations of the findings were also considered.     

The benzodiazepine midazolam does not impair Pavlovian fear conditioning but regulates when and where fear is expressed.

Rats were injected with a benzodiazepine (midazolam) and shocked after presentation of an auditory conditioned stimulus (CS). They were then tested for fear reactions (freezing) to the CS in either the original context or a 2nd context after either a short (1-day) or long (21-day) retention interval. Rats tested in the original context froze less after 1 day than rats tested after that interval in the 2nd context or rats tested after 21 days. Moreover, rats tested after the long interval in the original context froze less than rats tested after that interval in the 2nd context. Therefore, midazolam does not impair the acquisition of conditioned fear but regulates when and where that fear is expressed. These effects of midazolam were interpreted as a contextually controlled deficit in the expression of conditioned fear that is similar to that associated with latent inhibition and extinction (M. E. Bouton, 1993).     

Malleability of conditioned associations: path dependence

Using conditioned suppression of barpressing to investigate the stability of a conditioned stimulus-unconditioned stimulus (CS-US) association, we gave water-deprived rats either a few pairings of the CS with a strong footshock US or many pairings with a weak footshock US so that barpress suppression in response to the CS was equated. Experiment 1 established training parameters that yielded this equivalence. Specifically, rapid acquisition to a preasymptotic level of responding with strong shock produced suppression comparable to the asymptotic level reached more slowly with weak shock. Experiment 2 showed that although equivalent performance was obtained from extensive conditioning with a weak shock or limited conditioning with strong shock, only extensive conditioning with weak shock resulted in retarded acquisition of an association between that same CS and a footshock level perceived as midway between the two initial training shock intensities as implied by asymptotic performance in Experiment 1. Experiment 3 demonstrated that the observed retardation in animals given many conditioning trials with weak shock was CS specific. Collectively, these findings indicate that the malleability of learned behavior is not simply a function of initial associative strength but is dependent on path during initial acquisition.     

Contextual control of inhibition with reinforcement: adaptation and timing mechanisms

Four experiments with rats studied the effects of switching the context after Pavlovian conditioning. In three conditioned suppression experiments, a large number of conditioning trials created "inhibition with reinforcement" (IWR), in which fear of the conditional stimulus (CS) reached a maximum and then declined despite continued CS-unconditional stimulus pairings. When IWR occurred, a context switch augmented fear of the CS; IWR and augmentation were highly correlated. Neither IWR nor augmentation resulted from inhibition of delay (IOD): In conditioned suppression, IWR and augmentation occurred without IOD (Experiment 3), and in appetitive conditioning (Experiment 4), IOD occurred without IWR or augmentation. IWR may occur in conditioned suppression because the animal adapts to fear of the CS in a context-specific manner. The authors discuss several implications.     

Social transmission of Pavlovian fear: fear-conditioning by-proxy in related female rats

Pairing a previously neutral conditioned stimulus (CS; e.g., a tone) to an aversive unconditioned stimulus (US; e.g., a foot-shock) leads to associative learning such that the tone alone will elicit a conditioned response (e.g., freezing). Individuals can also acquire fear from a social context, such as through observing the fear expression of a conspecific. In the current study, we examined the influence of kinship/familiarity on social transmission of fear in female rats. Rats were housed in triads with either sisters or non-related females. One rat from each cage was fear conditioned to a tone CS+ shock US. On day two, the conditioned rat was returned to the chamber accompanied by one of her cage mates. Both rats were allowed to behave freely, while the tone was played in the absence of the foot-shock. The previously untrained rat is referred to as the fear-conditioned by-proxy (FCbP) animal, as she would freeze based on observations of her cage-mate’s response rather than due to direct personal experience with the foot-shock. The third rat served as a cage-mate control. The third day, long-term memory tests to the CS were performed. Consistent with our previous application of this paradigm in male rats (Bruchey et al. in Behav Brain Res 214(1):80–84, 2010), our results revealed that social interactions between the fear conditioned and FCbP rats on day two contribute to freezing displayed by the FCbP rats on day three. In this experiment, prosocial behavior occurring at the termination of the cue on day two was significantly greater between sisters than their non-sister counterparts, and this behavior resulted in increased freezing on day three. Our results suggest that familiarity and/or kinship influences the social transmission of fear in female rats.     

A change of context can enhance performance of an aversive but not of an appetitive conditioned response

In four experiments rats received pairings of one auditory CS and mild shock in one context and of a second auditory CS and shock in a second context. When tested with one of these CSs in the context in which it had never been experienced, they consistently showed enhanced levels of suppression. In one of these experiments, suppression was measured over 15-sec intervals throughout a 90-sec long CS. A change of context resulted in an increased level of suppression at all points throughout the CS, a finding which does not encourage the belief that it was due to an increase in unconditioned suppression at the onset of the CS. Another experiment provided equally little support for an alternative account in terms of increased arousal. The results therefore suggest that a change of context can increase the level of conditioned suppression elicited by a CS paired with mild shock. Two final experiments employing the same CSs and contexts, however, found no evidence that a change of context had any effect at all on the performance of an appetitively conditioned response.     

Conditioned suppression under positive, negative, and no contingency between conditioned and unconditioned stimuli

Using a conditioned suppression procedure, the effects of three contingent relationships between conditioned (CS) and unconditioned (US) stimuli were investigated. A traditional positive (if CS—then US) contingency suppressed response rate during the CS relative to responding during stimulus-free minutes of the session. A negative (if CS—then no US) contingency resulted in suppressed responding during CS-off minutes, and rate increases during the CS. A no-contingency control procedure, during which CS and US were randomly related, almost totally suppressed responding throughout the session and showed no differential effects of the CS on response rate. An analysis of changes in response rate during the minute after US-offset revealed acceleration under the no-contingency condition and, to a somewhat lesser degree, under the negative contingency. Both conditioned suppression and non-suppression are analyzed in terms of the temporal relationship between CS and US.     

Recent exposure to a dangerous context impairs extinction and reinstates lost fear reactions

Rats were shocked in a context and then exposed to that context in the absence of shock. Shorter intervals between these extinction trials produced more long-term freezing than did longer ones, and shorter intervals between the final extinction trial and test produced more freezing than did longer ones. A short interval between a context extinction trial and test with an extinguished conditioned stimulus (CS) produced more freezing than did a longer one, and a short interval between a nonreinforced context exposure and an extinguished CS reinstated freezing when the CS was tested 24 hr later. The results suggest that recent fear acts to favor subsequent retrieval of the memory formed at conditioning rather than extinction and to render the retrieved memory more salient.     

Systemic blockade of D2-like dopamine receptors facilitates extinction of conditioned fear in mice

Extinction of conditioned fear in animals is the explicit model of behavior therapy for human anxiety disorders, including panic disorder, obsessive-compulsive disorder, and post-traumatic stress disorder. Based on previous data indicating that fear extinction in rats is blocked by quinpirole, an agonist of dopamine D2 receptors, we hypothesized that blockade of D2 receptors might facilitate extinction in mice, while agonists should block extinction, as they do in rats. One day after fear conditioning mice with three pairings of a white noise conditional stimulus (CS) with moderate footshock, we injected the D2 antagonist, sulpiride, the D2 agonist, quinpirole, or vehicle, just before repeated CS presentations to generate extinction. We assayed fear by measuring behavioral freezing during extinction presentations and then drug-free during CS presentations 1 d later. We found that sulpiride injections before extinction training facilitated extinction memory 24 h later, while quinpirole partially blocked extinction memory compared with vehicle-injected controls. Notably, sulpiride treatment yielded significant extinction after spaced CS presentations, which yield no extinction at all in vehicle-treated mice. These findings suggest that dopamine D2-mediated signaling contributes physiological inhibition of extinction, and that D2 antagonists may be useful adjuncts to behavior therapy of human anxiety disorders.     

Backward conditioning as an extinction procedure

In two conditioned suppression experiments, rats received Pavlovian forward defense conditioning in which tonal conditioned stimuli (CSs) terminated with the onset of scrambled grid shock unconditioned stimuli (USs). After this experience, the rats then received a Pavlovian backward conditioning procedure in which the same USs now terminated with the onset of the same CSs. Although the two experiments differed greatly in terms of CS and US parameters, number of forward and backward pairings, and in terms of the general techniques used to establish and measure the Pavlovian conditioned response (CR), the results of both experiments agreed in showing that backward conditioning can indeed weaken a CR based on forward pairings. The results also show that, under some conditions, the backward procedure can be at least as effective in weakening an established CR as the traditional CS-alone extinction procedure; but, under other conditions, the backward procedure is less effective and leads to more spontaneous recovery than the CS-alone procedure.     

Some notes on conditioned suppression and reinforcement schedules

Two pigeons were trained on an FR 150 schedule of reinforcement. An Estes-Skinner conditioned suppression procedure was then superimposed on this performance at varied intervals. If the CS occurred during the early stages of the ratio run, complete suppression resulted. If the CS occurred during the later stages of the run, the birds continued to respond until the reinforcement was obtained, which was then followed by complete suppression.     

The effects of lesions in the central nucleus of the amygdala on appetitive classical conditioning in rats

Abstract: Lesions in the central nucleus of the amygdala (cAMY) have been known to interfere with the acquisition of fear classical conditioning when footshock is used as an unconditioned stimulus (US). The present study examined whether or not a similar interference would occur with an appetitive US. Five rats with lesions in the cAMY (the cAMY group), and eight unoperated control rats were trained in an appetitive classical conditioning paradigm, which did not include elements of operant learning, using a visual conditioned stimulus (CS) (5 W of light for 10 s duration) paired with a food pellet US (45 mg, cheese flavor). The behavioral index of appetitive conditioning was an increase in rearing approach behavior to the CS after CS and US pairings. During CS and US pairings, the movement of the rat was limited so that this approach behavior could not occur. As a result, all control rats showed an increase in rearing, but the cAMY group did not. These results suggest that the cAMY is critical for appetitive as well as fear classical conditioning.     

An animal model of fetishism

An animal model of sexual fetishism was developed with male Japanese quail based on persistence of conditioned sexual responding during extinction to an inanimate object made of terrycloth (Experiments 1 and 3). This persistent responding occurred only in subjects that came to copulate with the terrycloth object, suggesting that the copulatory behavior served to maintain the fetishistic behavior. Sexual conditioning was carried out by pairing a conditioned stimulus (CS) with the opportunity to copulate with a female (the unconditioned stimulus or US). Copulation with the CS object and persistent responding did not develop if the CS was a light (Experiment 1) or if conditioning was carried out with a food US (Experiment 2). In addition, subjects that showed persistence in responding to the terrycloth CS did not persist in their responding to a light CS (Experiment 3). The results are consistent with the hypothesis that conditioned copulatory behavior creates a form of self-maintenance that leads to persistent responding to an inanimate object. The development of an animal model of such fetishistic behavior should facilitate experimental analysis of the phenomenon.     

Role of conditioned contextual stimuli in reinstatement of extinguished fear

If the unconditioned stimulus (US) is presented independently of the conditioned stimulus (CS) following extinction, the conditioned response may be reinstated to the CS. Three experiments are reported that suggest that reinstatement is mediated by conditioning to contextual stimuli that are present during both US presentation and testing. Shocks presented to rats following the extinction of conditioned suppression reliably reinstated suppression to the CS, but only when they were presented in the context in which testing was later to occur. Reinstatement was also reversed by extinguishing fear to the context through nonreinforced exposure to the context between shock presentation and testing. Reinstatement was obtained in these experiments in spite of procedures that have been used in the past to minimize the influence of context conditioning. Moreover, fear of the context was never detected directly by depressed bar-press rates in the absence of the CS. The results do not support the hypothesis that reinstatement results from an increment in the strength of a memory of the US that has been weakened during extinction. Problems inherent in controlling and detecting levels of context conditioning that may influence behavior toward nominal CSs are discussed.