Chronic stress alters behavior in the conditioned defensive burying test: role of the posterior paraventricular thalamus

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Abstract

In the present studies, we examined the effects of chronic restraint on behavior in the conditioned defensive burying paradigm, a well-validated test of anxiety. This test is based on the findings that rodents tend to cover or bury the source of a noxious or aversive stimulus. However, little is known about whether prior chronic stress exposure can alter this anxiety-related behavior. In the present study, we examined whether chronic restraint affects indices of behavior in the conditioned defensive burying paradigm. Furthermore, since the posterior division of the paraventricular thalamus (pPVTh) regulates neuroendocrine activity specifically in chronically stressed but not control rats, we hypothesized that the pPVTh may also regulate any chronic stress-induced changes in behavior observed in the defensive burying test. Chronically stressed rats (30-min restraint per day for seven consecutive days) exhibited decreased latency to bury compared to control rats regardless of the presence of lesions suggesting increased reactivity to the shock in these animals. Importantly, pPVTh-lesioned chronically stressed rats exhibited increased duration and height of burying compared to control rats with pPVTh lesions, whereas no differences existed between sham-lesioned control and chronically stressed rats. Since both burying height and duration of burying are considered indices of anxiety in the defensive burying test, the present results suggest that the intact pPVTh may be important in dampening behaviors related to anxiety in chronically stressed rats.

Introduction

Chronic or repeated exposure to stress exerts powerful influences on activity within the hypothalamic–pituitary–adrenal (HPA) axis, and on memory, synaptic plasticity and behavior Dallman et al., 2002, McEwen, 2002, Ottenweller et al., 1989. Chronic stress has different effects on subsequent responsivity of the HPA axis depending on whether the subsequent stress is novel or previously experienced (Dallman et al., 1992). Our recent findings pertaining to the neural circuitry that underlies chronic stress-induced HPA activity strongly suggest a critical role for the posterior paraventricular thalamus (pPVTh). Lesions of the pPVTh prevent habituation to repeated restraint (Bhatnagar et al., 2002) and augment the facilitated HPA responses seen in chronically stressed rats exposed to novel stress (Bhatnagar and Dallman, 1998). In both cases, pPVTh lesions did not alter HPA function in acutely stressed rats. These data suggest that, under normal conditions, the pPVTh inhibits HPA responses specifically in chronically stressed animals but has no functional effect in acutely stressed rats.

Given the importance of the pPVTh in regulation of chronic stress-induced HPA activity, we hypothesized that it would also play an important role in chronic stress-induced changes in physiology and behavior. Indeed, the pPVTh regulates circadian rhythms in body temperature in chronically stressed but not control rats (Bhatnagar and Dallman, 1999). In the present experiments, we asked whether the pPVTh regulates behavior in a test of anxiety differently in control vs. chronically stressed rats. The conditioned defensive burying paradigm is a well-validated test of anxiety Pinel and Treit, 1978, Treit et al., 1981. Burying behavior is conditioned to an aversive stimulus, an electrified probe, and is readily observable in a single trial. Anxiolytic drugs such as diazepam and chlordiazepoxide decrease duration and height of burying without producing overt motor impairments Pinel and Treit, 1983, Treit et al., 1981. Thus, increases in duration and height of buried bedding are thought to reflect increased anxiety. Defensive burying has been used to study animals of different genetic backgrounds (Pare, 1994) and following early environmental manipulations Horvath et al., 1999, Felszeghy et al., 1993. However, little is known about whether chronic exposure to stress can alter behavior in this test even though chronic or repeated exposure to stress is closely associated with anxiety-related disorders Korte, 2001, Haller, 2001, Tache et al., 2001.

Given the important role of the pPVTh in regulating chronic stress-induced changes in HPA activity and the lack of information about the effects of chronic stress on behavior in the defensive burying paradigm, we examined behavior in the defensive burying test of sham- and pPVTh-lesioned animals that were exposed to chronic restraint (for seven consecutive days) or to no restraint at all. Our previous work in chronically stressed animals has measured indices of neuroendocrine function and physiology on Day 8 after 7 days of stress Bhatnagar and Dallman, 1998, Bhatnagar et al., 2000, Bhatnagar et al., 2002. To allow us to make comparisons between the effects of chronic stress on neuroendocrine function, physiology and behavior, we wanted to measure behavior in the defensive burying paradigm on Day 8 after 7 days of chronic stress. However, testing in the burying apparatus requires habituation to the apparatus for 3–4 days immediately before testing so that exposure to environmental novelty does not interfere with latency to contact the probe or burying behavior (Treit et al., 1981). Use of such a habituation procedure would mean that chronically stressed rats would also be habituated during the last 4 days of chronic stress, whereas control rats would only be habituated to the apparatus. To determine whether we could avoid this potential confounding situation of pairing restraint with habituation to the apparatus in chronically stressed rats, we first determined whether habituation to the apparatus could be administered for 4 days before the 7 days when chronic stress would normally take place. In Experiment 1, we found that habituating naı̈ve (nonstressed) rats 7 days before testing did not alter behavior compared to rats habituated for the 4 days immediately before the testing day. Therefore, in Experiment 2, sham- and pPVTh-lesioned rats were exposed to 4 days of habituation. For the next 7 days, control rats were undisturbed whereas chronically stressed rats were restrained for 30 min each day. Both control and chronically stressed rats were then tested in the defensive burying test on Day 8.

Section snippets

Animals

All experiments used young adult male Sprague–Dawley rats supplied by Harlan Sprague Dawley (Indianapolis, IN). Body weights ranged from 200 to 225 g upon arrival at the animal housing facilities at the Department of Psychology, University of Michigan. Rats were individually housed in hanging metal cages, and were allowed ad libitum access to rat chow and water. They were maintained on a 12-h light/dark schedule (lights on at 07:00 h), and all experiments took place during the trough of the

Statistics

A one-way analysis of variance (ANOVA) was used to analyze data in Experiment 1. A two-way ANOVA [Stress (Control or Chronic)×Lesion (Sham or pPVTh Lesion)] was used to analyze data collected in Experiment 2 and significant effects were followed by Fisher's post hoc tests. The significance level was set at P≤.05 for all analyses.

Experiment 1

We examined behavior of rats habituated to the testing apparatus for 4 days before the day of testing or 7 days before the day of testing. Four out of seven rats exhibited measurable burying behavior in each group. We found that these two groups did not exhibit any differences in height of buried bedding, latency to first contact the probe and latency to bury the probe (Table 1). Based on these results, in Experiment 2, we habituated both control and chronically stressed rats for 4 days 1 week

Discussion

We examined whether chronic exposure to restraint stress alters behavior in the defensive burying test, a validated model for testing anxiety-related behaviors. Chronic restraint decreased the latency to bury compared to control rats regardless of whether either group of rats was lesioned or not. pPVTh lesions resulted in increased burying height and duration of burying in chronically stressed rats compared to controls with such lesions. Since duration of burying and burying height are

Acknowledgments

This work was supported by the National Science Foundation (IBN-01152).

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