As noted above, the experience of potent stressors alters how the organism reacts to subsequent stressors. The issues to be explored here are whether, and by what mechanism(s), the behavioral controllability of the initial stressor alters reactions to subsequent stressors. The recent work from our laboratory directed at these questions derives from older reports indicating that a session of ES prevented a later exposure to IS from producing shuttlebox escape deficits (Seligman & Maier, 1967
), even if the two experiences occurred in different environments (Williams & Maier, 1977
). This was demonstrated to be a stressor controllability effect as an initial experience with yoked IS did not have this blunting effect. This immunizing effect of prior ES has proved to be quite general, with demonstrated blockade of the effects of later IS on reducing social interaction (Christianson et al., 2009
) as well as drug reward (Rozeske et al., 2008
). Furthermore, the proactive blunting of the behavioral consequences of IS has been shown to be quite enduring, persisting for at least 2 months between ES and later IS (unpublished results).
Role of the vmPFC in the Production of Stressor Resistance
As reviewed above, activation of the vmPFC during ES has proven necessary for the presence of control to blunt the impact of the ongoing stressor. Thus, it is reasonable to suppose that engagement of the vmPFC during ES might also be necessary for ES to produce immunization with regard to the effects of future uncontrollable stressors. To examine this possibility, the vmPFC was inactivated with muscimol during the initial ES experience, and now ES no longer blocked either the DRN 5-HT activation or the behavioral consequences of IS administered 7 days later (Amat et al., 2006
The experiment described above indicates that immunization requires the vmPFC at the time of the initial controllable stressor. Perhaps immunization occurs because the experience of ES alters the vmPFC in such a way that even uncontrollable aversive events then activate it, thereby inhibiting the DRN and perhaps other stress-responsive structures. If this is so, then the vmPFC would be needed not only to acquire information about control but also to use this information to regulate the DRN at the time of later uncontrollable aversive stimulation. Thus, the vmPFC was inactivated at the time of later IS rather than earlier ES, and this manipulation also prevented immunization (Amat et al., 2006
). Now, IS both activated the DRN and produced behavioral consequences such as shuttlebox escape failure, even though the subjects had experienced prior ES. These data suggest that the vmPFC is a site of neural plasticity within the stressor controllability paradigm. It is often argued that lasting neural plasticity requires the synthesis of new proteins, and so Amat et al. (2006)
microinjected the protein synthesis inhibitor anisomycin in the vmPFC during ES, and this prevented ES from blocking the neurochemical and behavioral changes produced by later IS.
There are various ways to conceptualize the implications of the data above, but one idea is that perhaps the activation of vmPFC output neurons during ES “ties” or associates vmPFC neuronal activation to some aspect of the ES experience, to tailshock for example, so that after exposure to ES even IS now activates vmPFC output, thereby leading to the inhibition of the DRN and the blockade of behavioral changes that depend on DRN activation/sensitization. If the joint occurrence of aversive stimulation and ES-induced vmPFC activation is sufficient to tie the two together, then perhaps simply activating the vmPFC during stressor exposure would be sufficient to lead to stressor immunization. That is, perhaps control is not needed at all, just vmPFC activity during a stressor. To test this idea, the vmPFC was activated by picrotoxin during ES, IS, or control treatment (Amat et al., 2008
). IS administered 7 days after IS+vmPFC activation failed to activate the DRN or produce typical IS-induced behavioral changes. Thus, even IS produces immunization if the vmPFC is activated during its administration. Interestingly, vmPFC activation with picrotoxin by itself, in the absence of tailshock, failed to lead to immunization. Immunization required the conjoint presence of the stressor and the activation of the vmPFC, consistent with the argument that they become associated.
Social Defeat and Fear Conditioning
The research reviewed above suggests that the experience of control over tailshock blunts the DRN response to later uncontrollable tailshock and the behavioral changes that result from DRN 5-HT activation. It also suggests that this resistance to the effects of IS is mediated by ES-induced alterations in the vmPFC. Several questions suggest themselves. Perhaps the most obvious concerns whether the resistance to later stress effects produced by exposure to control over tailshock is restricted to later tailshock and very similar stimuli, or whether the resistance is more general.
To begin to explore this issue we chose to study the effects of ES on neurochemical and behavioral responses to a stressor that has as few external stimuli in common with tailshock and the wheel-turn apparatus as possible. Social defeat (SD) seemed to fit this requirement. In SD paradigms a target subject is exposed to a larger aggressive resident animal, and the outcome is almost always that the resident attacks the target, with the target subject rapidly adopting species-specific defeat postures. In the procedure that we adopted, the target rat was introduced into the cage of a resident male for 45 min. Thus, the SD procedure involves no shock or other discrete aversive stimuli, no apparatus similar to a wheel turn box, the presence of a conspecific, and in our procedure even occurs on another floor of the building so as to minimize the presence of common odor cues.
We first had to determine behavioral changes that might result from SD so that it could be determined whether prior ES would block them. As SD might be uncontrollable from the subjects’ perspective as defeat of the target is essentially inevitable, we determined whether SD would produce behavioral sequelae similar to those of IS. Indeed, SD produced shuttlebox escape failure and reduced juvenile social investigation 24 hr later (Paul et al., 2008
). Gardner et al. (2005)
had found a somewhat similar SD procedure to induce Fos in the DRN, so we determined whether SD would increase 5-HT in the DRN, and it did so. The important result was that ES occurring 7 days before SD blocked both the DRN activation and the behavioral consequences (poor escape and reduced social investigation) of SD. Yoked IS did not reduce the effects of later SD, and indeed, exacerbated them. Thus, it was control over tailshock that was responsible for the blunting of the impact of later SD (Maier, unpublished data).
These data clearly show that the experience of behavioral control over a potent aversive event alters the organism in a way that is much more general than how it responds to the same stimulus over which it had had control. It has yet to be determined whether alterations in the vmPFC are responsible for this generalized resistance produced by control over tailshock, but one interpretation would be that during ES vmPFC activation becomes associated with something much more general than tailshock, something common to a broad range of stressors, something such as fear/anxiety (see below). Determining how this generalized resilience operates will be a challenge.
A second question concerns whether control activates vmPFC projections to stress-responsive structures other than the DRN, thereby regulating behaviors controlled by these other structures. The amygdala is an obvious candidate. The role of the amygdala in fear and fear conditioning is well known (LeDoux, 2007
). The view has emerged that the association between neutral stimuli such as tones and aversive stimuli such as footshocks form in the basal nucleus, and from there input is provided to the central nucleus (CE), either directly or via the lateral nucleus. The CE, in turn, projects to the proximate mediators of fear responses, to the periaqueductal gray for example, resulting in freezing behavior. Thus, a tone or a context that has been paired with footshock produces fear responses because the CE is activated.
Interestingly, the vmPFC sends glutamatergic projections to a number of regions within the amygdaloid complex (Vertes, 2006
). The prelimbic region (PL) sends somewhat complex projections to the basolateral region. The infralimbic region (IL) sends a clear projection to a region of the amygdala called the intercalated cell region (ITC). These cells are almost entirely GABAergic, and in turn project to the CE (CE). As would be expected, activation of the IL with picrotoxin activates these ITC GABAergic cells, and stimulation of the IL results in the inhibition of CE output neurons and fear behaviors (Berretta et al., 2005
Given this anatomy, Baratta et al. (2007)
gave rats either ES, yoked IS, or control treatment. 7 days later all subjects received fear conditioning in which a tone was paired with footshock, followed 1 day later by fear testing to determine how much conditioning had occurred. Freezing was measured both to the conditioning context and to the tone (presented in a novel context) that had been paired with shock. The wheel turn apparatus and the conditioning boxes were sufficiently different that fear behavior did not generalize from the ES/IS treatment to the conditioning situation. That is, there was no freezing in the boxes before footshock, an outcome that would confound differences in conditioning. In keeping with prior reports (Rau et al., 2005
), IS potentiated fear conditioning. More importantly for the present paper, ES retarded fear conditioning—less fear was conditioned to both the context and the tone.
Because there has been a great deal of recent interest in the extinction of conditioned fear, Baratta et al. (2007)
determined whether experience with ES might not facilitate fear extinction. The logic of this question required a design in which ES and IS were administered after fear conditioning. This is because it is necessary that the amount of fear conditioning be equal in the various groups, otherwise, any apparent extinction differences could be attributed to acquisition differences. Subjects thus first received fear conditioning. For simplicity, there was no tone and only fear conditioning to the experimental context was assessed. Then, 1 day later, the rats were exposed to either ES, yoked, IS, or control treatment. 7 days later, all subjects began fear extinction consisting of daily 5 min exposures to the conditioning context, without the occurrence of footshock, that continued until a criterion of extinction had been reached. Remarkably, ES that occurred after fear conditioning hastened extinction of conditioned fear. This is remarkable because ES is not negatively fearful or stressful, it is quite aversive, yet it actually reduced fear responding relative to no exposure to a stressor at all.
Although the natural interpretation was that ES facilitated extinction learning, a more detailed observation questioned this possibility. The fear response in the conditioning context did disappear more rapidly in the ES subjects, but fear was diminished by the second minute of the very first extinction session. Clearly, this is too rapid for the subjects to have learned that footshock would no longer occur. Indeed, the first footshock did not occur on the conditioning day until the subjects had been in the context for 3 min.
If ES did not facilitate extinction learning, then how could it have produced the more rapid disappearance of fear? Another possibility would be that the experience of ES reduces the expression of fear behavior. That is, the stimulus context might still have high associative strength for the footshock UCS in ES subjects, but the behavior that such an association would normally produce is inhibited.
Perhaps a consideration of the amygdala circuitry briefly reviewed above would make this distinction more intelligible. The association between CS and UCS occurs in basolateral regions of the amygdala, but the IL does not project to this portion of the amygdala. Instead, the IL inhibits the CE projections to regions that produce fear responses; that is, the expression of fear.
Thus, it is possible that ES alters the IL in such a way that it is later activated by fear, thereby reducing fear expression. This could occur because the IL is sensitized by ES or because IL activity becomes associated with fear, or something that is part of fear. However, by either process, the argument is that the experience of ES does not alter the subsequent fear conditioning or extinction process, but rather reduces the expression of the fear behavior that is elicited by stimuli that signal footshock. The possibility that ES exposure reduces fear expression can be tested by selectively inactivating the IL during each of the phases of the experimental procedure in which an initial experience with ES was shown to interfere with fear conditioning. Recall that in this experiment schematized in Fig. 3, bottom row, animals first received either ES or IS (Phase 1), then 7 days later fear conditioning (Phase 2), and then 1 day later testing for the amount of fear that was conditioned (Phase 3).
Consider the predictions to be made by the hypotheses that ES interferes with fear expression:
If ES activates the IL, thereby altering it in such a way that it is later activated by fear, then inactivation of the IL during ES should prevent ES-induced reduction of fear measured in Phase 3. This is because the IL would now not be activated during fear, and therefore not either sensitized or “tied” to fear.
In contrast to inactivation during Phase 1, inactivation of the IL during the fear conditioning should have no effect, that is, prior ES should still retard fear conditioning. This is because the association between CS and UCS occurs in basolateral regions of the amygdala not in the ITC-CE pathway, the target of IL projections. Thus, the association should form normally even if the IL-ITC-CE pathway is active during the conditioning session.
The most interesting question concerns the prediction to be made if the IL is inactivated only during the last stage of the experiment, the test for fear. Since the IL was not inactivated during Phase 1 it should either become sensitized or tied to fear. The critical aspect of the fear expression hypothesis is that the experience of ES does not later fear learning, and so this should proceed normally in Phase 2. The hypothesis supposes that in the test phase the elicitation of fear activates the IL in subjects that had experienced ES, and so fear behavior is reduced because the ITC is activated, the ITC inhibiting CE output. Thus, inactivation of the IL during the test phase should lead to normal expression of fear behavior. That is, IL inactivation should reveal the fear conditioning that was there all along in ES subjects, an unmasking of the learning that was present, but whose output in behavior was inhibited. Clearly, the idea that exposure to ES interferes with the actual fear conditioning process would predict that IL inactivation of Phase 3 would be without impact as from this view Es would have undermined the conditioning in Phase 2. If conditioning was reduced, then fear should still be less in Phase 3 even if the IL was inhibited.
Baratta et al. (2008)
conducted precisely the experiment just outlined. Not surprisingly, muscimol microinjected into the IL during ES blocked the reduction in later fear conditioning produced by ES. Also perhaps not surprisingly, intra-IL muscimol administered before the fear conditioning did influence the ES-induced reduction in fear conditioning, nor did it have any effect on fear conditioning in controls. The critical question was what intra-IL muscimol before fear testing would do. In vehicle-injected controls ES in Phase 1 reduced the fear measured in Phase 3 testing. However, IL inactivation during testing eliminated the reduction in fear behavior in ES subjects and revealed the fear that had indeed been conditioned. Thus, it would appear that experiencing control over an aversive stimulus alters the IL in such a way that it is later activated under fear conditions, leading to the inhibition of fear behavior. Clearly, ES reduced the expression of fear, not the development of fear conditioning.
In sum, the experience of control over tailshock does not alter only how the organism reacts to later shock and regulates not only how the DRN responds to later events. The experience of ES had a substantial effect on how the subjects reacted to SD, an event quite different than tailshock. Indeed, SD is sufficiently different from tailshock that it is more than possible that experiencing control over a potent aversive stimulus alters the organism in a profound manner. In coping terms, perhaps it shifts how the organism attempts to deal with future threat. This is clearly mediated, at least in part, by changes in the vmPFC and its regulation of other structures such as the DRN. The experiments involving fear conditioning suggest that this regulation is not limited to the DRN.