Not all stressors are equal : behavioral and endocrine evidence for development of contextual fear conditioning after a single session of footshocks but not of immobilization

Exposure of animals to footshocks (FS) in absence of any specific cue results in the development of fear to the compartment where shocks were given (contextual fear conditioning), and this is usually evaluated by time spent freezing. However, the extent to which contextual fear conditioning always develops when animals are exposed to other stressors is not known. In the present work we firstly demonstrated, using freezing, that exposure of adult rats to a single session of FS resulted in short-term and long-term contextual fear conditioning (freezing) that was paralleled by increased hypothalamic-pituitary-adrenal (HPA) activation. In contrast, using a similar design, no HPA or behavioral evidence for such conditioning was found after exposure to immobilization on boards (IMO), despite this stressor being of similar severity as FS on the basis of standard physiological measures of stress, including HPA activation. In a final experiment we directly compared the exposure to the two stressors in the same type of context and tested for the development of conditioning to the context and to a specific cue for IMO (the board). We observed the expected high levels of freezing and the conditioned HPA activation after FS, but not after IMO, regardless of the presence of the board during testing. Therefore, it can be concluded that development of fear conditioning to context or particular cues, as evaluated by either behavioral or endocrine measures, appears to be dependent on the nature of the aversive stimuli, likely to be related to biologically preparedness to establish specific associations

Sex differences in the behavioural and hypothalamic-pituitary-adrenal response to contextual fear conditioning in rats

This is the author's version of a work that was accepted for publication in Hormones and behavior. Changes resulting from the publishing process, such as peer review, editing, corrections, structural formatting, and other quality control mechanisms may not be reflected in this document. Changes may have been made to this work since it was submitted for publication. A definitive version was subsequently published in Hormones and behavior, Vol. 66 No. 5 (Nov. 2014), p. 713-723In recent years, special attention is being paid to sex differences in susceptibility to disease. In this regard, there is evidence that male rats present higher levels of both cued and contextual fear conditioning than females. However, little is known about the concomitant hypothalamic-pituitary-adrenal (HPA) axis response to those situations which is critical in emotional memories. Here, we studied the behavioural and HPA responses of male and female Wistar rats to context fear conditioning using electric footshock as the aversive stimulus. Fear-conditioned rats showed a much greater ACTH and corticosterone response than those merely exposed to the fear conditioning chamber without receiving shocks. Moreover, males presented higher levels of freezing whereas HPA axis response was greater in females. Accordingly, during the fear extinction tests, female rats consistently showed less freezing and higher extinction rate, but greater HPA activation than males. Exposure to an open-field resulted in lower activity/exploration in fear-conditioned males, but not females, suggesting greater conditioned cognitive generalization in males than females. It can be concluded that important sex differences in fear conditioning are observed in both freezing and HPA activation, but the two set of variables are affected in the opposite direction: enhanced behavioural impact in males, but enhanced HPA responsiveness in females. Thus, the role of sex differences on fear-related stimuli may depend on the variables chosen to evaluate it, the greater responsiveness of the HPA axis in females perhaps being an important factor to be further explored

Stress-induced sensitization: the hypothalamic-pituitary-adrenal axis and beyond

Exposure to certain acute and chronic stressors results in an immediate behavioral and physiological response to the situation followed by a period of days when cross-sensitization to further novel stressors is observed. Cross-sensitization affects to different behavioral and physiological systems, more particularly to the hypothalamus-pituitary-adrenal (HPA) axis. It appears that the nature of the initial (triggering) stressor plays a major role, HPA cross-sensitization being more widely observed with systemic or high-intensity emotional stressors. Less important appears to be the nature of the novel (challenging) stressor, although HPA cross-sensitization is better observed with short duration (5-15 min) challenging stressors. In some studies with acute immune stressors, HPA sensitization appears to develop over time (incubation), but most results indicate a strong initial sensitization that progressively declines over the days. Sensitization can affect other physiological system (i.e. plasma catecholamines, brain monoamines), but it is not a general phenomenon. When studied concurrently, behavioral sensitization appears to persist longer than that of the HPA axis, a finding of interest regarding long-term consequences of traumatic stress. In many cases, behavioral and physiological consequences of prior stress can only be observed following imposition of a new stressor, suggesting long-term latent effects of the initial exposure

Condicionamiento de miedo al contexto: implicaciones para el estudio de modelos animales de Estrés Postraumático

La exposición a una sesión breve de choques eléctricos en un contexto determinado induce el desarrollo de miedo a dicho contexto, fenómeno conocido como condicionamiento de miedo al contexto. Además, produce a largo plazo una inhibición de la actividad en ambientes nuevos, efecto no necesariamente asociado a un aumento de la ansiedad, por lo que la interpretación de esta hipo-actividad se desconoce. Los resultados obtenidos indican que la aparición de la hipo-actividad inducida por la exposición previa a una sesión breve de choques eléctricos, sólo aparece si se ha podido desarrollar previamente un condicionamiento del miedo al contexto. Por otro lado, no observamos ninguna evidencia de un aumento de la conducta ansiosa. Este hallazgo sugiere que los animales que han podido desarrollar una generalización del miedo. Además, dicha hipo-actividad no iba asociada a un aumento de la respuesta del eje HPA y podía ser en gran medida revertida mediante la extinción del miedo condicionado. En el estudio de los efectos de la manipulación de la intensidad y el número de choques sobre los efectos condicionado, pudimos observar que desde un punto de vista conductual, la manipulación de los parámetros resultó poco efectiva, lo que contrasta con lo observado en la inducción de la hipoactividad, donde la variación de la intensidad si resultó relevante. Sin embargo, se observó una respuesta endocrina intensidad-dependiente delante del contexto condicionado. Por otro lado, la sensibilización hormonal observada tras la exposición a un ambiente nuevo está relacionada con la intensidad del choque recibido. Por último, el estudio de la IMO como estímulo estresante no mostró evidencias de condicionamiento ni a nivel conductual ni endocrino. Por lo tanto, concluimos que el desarrollo del condicionamiento del miedo, evaluado mediante medidas conductuales y endocrinas, no es una propiedad universal compartida por todos los estímulos estresantes, lo que ayudaría a explicar las divergencias en los efectos a largo plazo obtenidos con diferentes estímulos estresantes.Exposure to a single session of footshock in absence of any specific cues results in the development of fear to the shock chamber, this is called contextual fear conditioning, that it is usually evaluated by time spend freezing. Furthermore, this shock exposure induces long-lasting inhibition of activity in unknown environments, these effects are not necessarily associated to an enhanced anxiety and the interpretation of the hypoactivity remains unclear. However, the development of contextual fear conditioning using other stressors remains poorly studied. Our results indicated that hypoactivity and a certain level of generalization of fear to the unknown environment only appeared when animals are able to developed fear conditioning, but no evidence for enhanced anxiety was found. These results suggest that if animals are able to associate an aversive experience with unknown environments, they display more caution behaviors in any unknown environments. In addition, this hypoactivity was not associated with greater HPA response in any different environment and it could be partially reversed by an extinction protocol. Moreover, the effect of shock intensity and number of shocks on both conditioned and unconditioned consequences was also studied. From the behavioural point of view the manipulation of intensity and number of shocks does not appear to be very relevant. But, an intensity-dependent endocrine response is observed when animals were exposed to the conditioned context. On the other hand, the endocrine sensitization after novel environments exposure is related to the high response showed after shock exposure. Regarding the use of other stressors to induce contextual fear conditioning, despite shock and IMO showed similar severity, no behavioural or endocrine evidence of conditioning was found if IMO was used as a stressor. Therefore, it can be concluded that development of fear conditioning, as evaluated by either behavioural or endocrine measures, appears to be dependent on the nature of the aversive stimuli. These phenomena could help to explain the divergences in long-lasting effects of each stressor

Not all stressors are equal : behavioral and endocrine evidence for development of contextual fear conditioning after a single session of footshocks but not of immobilization

Exposure of animals to footshocks (FS) in absence of any specific cue results in the development of fear to the compartment where shocks were given (contextual fear conditioning), and this is usually evaluated by time spent freezing. However, the extent to which contextual fear conditioning always develops when animals are exposed to other stressors is not known. In the present work we firstly demonstrated, using freezing, that exposure of adult rats to a single session of FS resulted in short-term and long-term contextual fear conditioning (freezing) that was paralleled by increased hypothalamic-pituitary-adrenal (HPA) activation. In contrast, using a similar design, no HPA or behavioral evidence for such conditioning was found after exposure to immobilization on boards (IMO), despite this stressor being of similar severity as FS on the basis of standard physiological measures of stress, including HPA activation. In a final experiment we directly compared the exposure to the two stressors in the same type of context and tested for the development of conditioning to the context and to a specific cue for IMO (the board). We observed the expected high levels of freezing and the conditioned HPA activation after FS, but not after IMO, regardless of the presence of the board during testing. Therefore, it can be concluded that development of fear conditioning to context or particular cues, as evaluated by either behavioral or endocrine measures, appears to be dependent on the nature of the aversive stimuli, likely to be related to biologically preparedness to establish specific associations

Sex differences in the behavioural and hypothalamic-pituitary-adrenal response to contextual fear conditioning in rats

This is the author's version of a work that was accepted for publication in Hormones and behavior. Changes resulting from the publishing process, such as peer review, editing, corrections, structural formatting, and other quality control mechanisms may not be reflected in this document. Changes may have been made to this work since it was submitted for publication. A definitive version was subsequently published in Hormones and behavior, Vol. 66 No. 5 (Nov. 2014), p. 713-723In recent years, special attention is being paid to sex differences in susceptibility to disease. In this regard, there is evidence that male rats present higher levels of both cued and contextual fear conditioning than females. However, little is known about the concomitant hypothalamic-pituitary-adrenal (HPA) axis response to those situations which is critical in emotional memories. Here, we studied the behavioural and HPA responses of male and female Wistar rats to context fear conditioning using electric footshock as the aversive stimulus. Fear-conditioned rats showed a much greater ACTH and corticosterone response than those merely exposed to the fear conditioning chamber without receiving shocks. Moreover, males presented higher levels of freezing whereas HPA axis response was greater in females. Accordingly, during the fear extinction tests, female rats consistently showed less freezing and higher extinction rate, but greater HPA activation than males. Exposure to an open-field resulted in lower activity/exploration in fear-conditioned males, but not females, suggesting greater conditioned cognitive generalization in males than females. It can be concluded that important sex differences in fear conditioning are observed in both freezing and HPA activation, but the two set of variables are affected in the opposite direction: enhanced behavioural impact in males, but enhanced HPA responsiveness in females. Thus, the role of sex differences on fear-related stimuli may depend on the variables chosen to evaluate it, the greater responsiveness of the HPA axis in females perhaps being an important factor to be further explored

Stress-induced sensitization: the hypothalamic-pituitary-adrenal axis and beyond

Exposure to certain acute and chronic stressors results in an immediate behavioral and physiological response to the situation followed by a period of days when cross-sensitization to further novel stressors is observed. Cross-sensitization affects to different behavioral and physiological systems, more particularly to the hypothalamus-pituitary-adrenal (HPA) axis. It appears that the nature of the initial (triggering) stressor plays a major role, HPA cross-sensitization being more widely observed with systemic or high-intensity emotional stressors. Less important appears to be the nature of the novel (challenging) stressor, although HPA cross-sensitization is better observed with short duration (5-15 min) challenging stressors. In some studies with acute immune stressors, HPA sensitization appears to develop over time (incubation), but most results indicate a strong initial sensitization that progressively declines over the days. Sensitization can affect other physiological system (i.e. plasma catecholamines, brain monoamines), but it is not a general phenomenon. When studied concurrently, behavioral sensitization appears to persist longer than that of the HPA axis, a finding of interest regarding long-term consequences of traumatic stress. In many cases, behavioral and physiological consequences of prior stress can only be observed following imposition of a new stressor, suggesting long-term latent effects of the initial exposure

Behavioral and endocrine consequences of simultaneous exposure to two different stressors in rats : interaction or independence?

Although behavioral and endocrine consequences of acute exposure to stressors have been extensively studied, little is known about how simultaneous exposure to two different stressors interacts to induce short- and long-term effects. In the present experiment we studied this interaction in adult male rats exposed to cat fur odor (impregnated cloth) or immobilization on boards either separately or simultaneously. We reasoned that exposure to the odor of a potential predator while immobilized, may potentiate its negative consequences as compared to exposure to only one of the stressors. Exposure to cat odor elicited the expected reduction of activity and avoidance of the area where the impregnated cloth was located. The endocrine response (plasma levels of ACTH and corticosterone, as a measure of the hypothalamic-pituitary-adrenal axis, HPA) was markedly greater after immobilization than after cat fur odor and no additive effects were found by simultaneous exposure to both stressors. Cat odor, but not immobilization, increased anxiety-like behavior as evaluated in the elevated plus-maze 7 days after the stressors, with no evidence of enhanced HPA activation. In addition, cat odor exposure resulted in long-lasting (8 days later) fear conditioning to the box containing a clean cloth, which was reflected by hypoactivity, avoidance of the cloth area and enhanced HPA activation. All these effects were similarly observed in rats exposed simultaneously to cat odor and immobilization. In rats only exposed to immobilization, only some weak behavioral signs of fear conditioning were found, but HPA activation in response to the context paired to immobilization was enhanced to the same extent as in cat odor-exposed animals, supporting a certain degree of endocrine conditioning. The present results did not reveal important behavioral interactions between the two stressors when animals experienced both simultaneously, whereas some interactions were found regarding HPA activation. Theoretical implications are discussed

Sex-dependent effects of an early life treatment in rats that increases maternal care : vulnerability or resilience?

Altres ajuts: Del MICINN, el PTA2010-3472-I; FPU AP-2007-02841Early life stress (ELS) in rodents has profound long-term effects that are partially mediated by changes in maternal care. ELS not only induces "detrimental" effects in adulthood, increasing psychopathology, but also promotes resilience to further stressors. In Long-Evans rats, we evaluated a combination of two procedures as a model of ELS: restriction of bedding during the first post-natal days and exposure to a "substitute" mother. The maternal care of biological and "substitute" mothers was measured. The male and female offspring were evaluated during adulthood in several contexts. Anxiety was measured by the elevated plus-maze (EPM), acoustic startle response (ASR) and forced swim test (FST). In other group of animals, novelty-seeking was measured (activity in an inescapable novel environment, preference for novel environments and exploration of novel objects). Plasmatic ACTH and corticosterone in basal conditions and in response to stress were also measured. Cognitive impulsivity was assessed by a delay-discounting paradigm, and impulsive action, attention and compulsive-like behavior by a five choice serial reaction time task (5CSRTT). ELS decreased pup body weight and increased the care of the biological mother; however, the "substitute" mother did not exhibit overt maltreatment. A mixture of "detrimental" and "beneficial" effects was shown. In the 5CSRTT, attention was impaired in both genders, and in females, ELS increased compulsive-like behavior. Novel object exploration was only increased by ELS in males, but the preference for novel spaces decreased in both genders. Baseline anxiety (EPM and ASR) and recognition memory were not affected. Unexpectedly, ELS decreased the ACTH response to novelty and swim stress and increased active coping in the FST in both genders. Cognitive impulsivity was decreased only in females, but impulsive action was not affected. The enhancement in maternal care may "buffer" the effects of ELS in a context-dependent manner

Condicionamiento de miedo al contexto : implicaciones para el estudio de modelos animales de estrés postraumático /

La exposición a una sesión breve de choques eléctricos en un contexto determinado induce el desarrollo de miedo a dicho contexto, fenómeno conocido como condicionamiento de miedo al contexto. Además, produce a largo plazo una inhibición de la actividad en ambientes nuevos, efecto no necesariamente asociado a un aumento de la ansiedad, por lo que la interpretación de esta hipo-actividad se desconoce. Los resultados obtenidos indican que la aparición de la hipo-actividad inducida por la exposición previa a una sesión breve de choques eléctricos, sólo aparece si se ha podido desarrollar previamente un condicionamiento del miedo al contexto. Por otro lado, no observamos ninguna evidencia de un aumento de la conducta ansiosa. Este hallazgo sugiere que los animales que han podido desarrollar una generalización del miedo. Además, dicha hipo-actividad no iba asociada a un aumento de la respuesta del eje HPA y podía ser en gran medida revertida mediante la extinción del miedo condicionado. En el estudio de los efectos de la manipulación de la intensidad y el número de choques sobre los efectos condicionado, pudimos observar que desde un punto de vista conductual, la manipulación de los parámetros resultó poco efectiva, lo que contrasta con lo observado en la inducción de la hipoactividad, donde la variación de la intensidad si resultó relevante. Sin embargo, se observó una respuesta endocrina intensidad-dependiente delante del contexto condicionado. Por otro lado, la sensibilización hormonal observada tras la exposición a un ambiente nuevo está relacionada con la intensidad del choque recibido. Por último, el estudio de la IMO como estímulo estresante no mostró evidencias de condicionamiento ni a nivel conductual ni endocrino. Por lo tanto, concluimos que el desarrollo del condicionamiento del miedo, evaluado mediante medidas conductuales y endocrinas, no es una propiedad universal compartida por todos los estímulos estresantes, lo que ayudaría a explicar las divergencias en los efectos a largo plazo obtenidos con diferentes estímulos estresantes.Exposure to a single session of footshock in absence of any specific cues results in the development of fear to the shock chamber, this is called contextual fear conditioning, that it is usually evaluated by time spend freezing. Furthermore, this shock exposure induces long-lasting inhibition of activity in unknown environments, these effects are not necessarily associated to an enhanced anxiety and the interpretation of the hypoactivity remains unclear. However, the development of contextual fear conditioning using other stressors remains poorly studied. Our results indicated that hypoactivity and a certain level of generalization of fear to the unknown environment only appeared when animals are able to developed fear conditioning, but no evidence for enhanced anxiety was found. These results suggest that if animals are able to associate an aversive experience with unknown environments, they display more caution behaviors in any unknown environments. In addition, this hypoactivity was not associated with greater HPA response in any different environment and it could be partially reversed by an extinction protocol. Moreover, the effect of shock intensity and number of shocks on both conditioned and unconditioned consequences was also studied. From the behavioural point of view the manipulation of intensity and number of shocks does not appear to be very relevant. But, an intensity-dependent endocrine response is observed when animals were exposed to the conditioned context. On the other hand, the endocrine sensitization after novel environments exposure is related to the high response showed after shock exposure. Regarding the use of other stressors to induce contextual fear conditioning, despite shock and IMO showed similar severity, no behavioural or endocrine evidence of conditioning was found if IMO was used as a stressor. Therefore, it can be concluded that development of fear conditioning, as evaluated by either behavioural or endocrine measures, appears to be dependent on the nature of the aversive stimuli. These phenomena could help to explain the divergences in long-lasting effects of each stressor