Research report
Altered behavioural response to acute stress in mice lacking cellular prion protein

https://doi.org/10.1016/j.bbr.2005.02.003Get rights and content

Abstract

Although many studies have investigated the function of cellular prion protein (PrPc), its physiologic role remains elusive. PrPc null mice have been reported to develop normally and to show normal performance in most behavioural tests. In the present study we investigated whether this also holds true after episodes of acute stress. PrPc gene ablated (Prnp0/0) and wild-type mice were subjected to restraint stress, electric foot shock, or swimming and compared with non-stressed animals. Immediately after the stressful situation, the anxiety levels and locomotion of the animals were measured using plus-maze and open-field tests. Among non-stressed animals, there was no significant difference in performance between Prnp0/0 and wild type animals in either test. However, after acute stress provoked by a foot shock or a swimming trial, Prnp0/0 animals showed a significant decrease in anxiety levels when compared with control animals. Moreover, after the swimming test, knockout mice presented decreased locomotion when compared to wild-type mice. Because of this observation, we also assessed both types of mice in a forced swimming test with the objective of better evaluating muscle function and found that Prnp0/0 animals presented reduced forced swimming capacity when compared to controls. As far as we know, this is the first report suggesting that cellular prion protein is involved in modulation of anxiety or muscular activity after acute psychic or physical stress.

Introduction

Cellular prion protein (PrPc) is a glycoprotein anchored to the outer cell surface which has been highly conserved during evolution. It is expressed predominantly in neurons and, to a lesser extent, in other types of cells [1], [2], [3], [4], [5], [6]. An alteration in its secondary structure leads to accumulation of an abnormal protein PrPsc and causes an uncommon disease known as spongiform encephalopathy [7], [8], [9].

The physiological role of PrPc, however, remains elusive. The first observations of PrPc gene ablated (Prnp0/0) mice suggested that these animals developed normally and did not show major behavioural abnormalities at baseline [10]. However, during the last 10 years, some differences in behaviour were noticed between some PrPc gene ablated mice and their respective wild-type controls [11], [12], [13], [14], [15], [16]. Interestingly, many of these differences were observed only if the animals were subjected to conditions which were different from baseline ones, such as during provoked seizures [16] or after head trauma [17].

The differences between diverse PrPc null mice strains and their controls have been better studied at the cellular or subcellular level. It has been demonstrated, for example, that PrPc-deficient cells show an altered response to oxidative stress [18], [19], [20], [21], present increased levels of oxidative stress markers [20], and show imbalances in antioxidant defence [22]. Absence of PrPc also leads to altered calcium [23], zinc, [24] and copper [25] homeostasis, which might explain some of the differences in oxidative stress. Furthermore, lack of PrPc expression results in minor morphological alterations such as differences in mitochondrial numbers and morphology [26] and in hippocampal mossy fibers [27].

Based on these observations, some authors proposed physiological roles for PrPc in the regulation of oxidative stress, as well as in cell survival, signal transduction and neural excitability [1], [28], [29], [30], [31], [32]. Because of its location, anchored to the outer surface of the neurons, some authors have also suggested that PrPc may influence synaptic function as well [28], [31]. Common to several of the above cited observations is the concept that PrPc may become physiologically important when levels of functioning are different from baseline conditions. In other words, when normal homeostasis is altered, the deficits caused by the absence of PrPc may start to appear. In order to test if this might be true for behavioural tests, we evaluated the effect of acute stress on the levels of anxiety of Prnp0/0 mice. Because of additional observations during the experiments, we also evaluated the effect of physical stress on locomotor function among wild-type and Prnp0/0 mice.

Section snippets

Animals

A total of 111 adult male knockout mice (3-month-old, weighing 20–40 g) homozygous for disrupted PrPc gene, PrnP (designated Prnp0/0 mice) produced as previously described [10] were used and 118 male wild-type (Prnp+/+) mice of the same age and weight were used as controls. The Prnp0/0 mice used were descendants of Zrch I animals [10]. Wild-type controls were generated by crossing F1 descendants from a 129/Sv × C57BL/6J mating. To confirm the genotype of the animals we conduced PCR procedures with

Statistical analysis

Data for the plus-maze test (experiment 1) are presented as mean (±S.E.M.) number of entries into the open arms, number of entries into the closed arms, total number of entries, time of permanence in the open arms, time of permanence in the closed arms, and time of permanence in the center of the maze. Data for the open-field test (experiment 2) are presented as mean (±S.E.M.) number of crossings, latency to start locomotion, and latency to reach the corner. The results of the plus-maze and

Experiment 1

The first experiment was designed to determine if PrPc is involved in anxiety modulation after acute behavioural stress. First, we randomly assigned four groups of Prnp0/0 mice (n = 57) and their respective Prnp+/+ controls (n = 58) to four types of acute stress: no stress, restraint stress, foot-shock, and swimming (n = 12–15 mice/group). Immediately after the acute stress, the animals were placed in the plus-maze apparatus for the evaluation of anxiety levels (Diagram 1).

Discussion

In this study, we observed that under baseline conditions Prnp0/0 mice presented similar anxiety levels and locomotion when compared to Prnp+/+ mice. However, after either electric footshock or swimming, Prnp0/0 mice spent significantly less time in the closed arms and more time in open places in the plus-maze test than controls. This suggests that the anxiety-related behavioural reactions after acute stress observed in controls are impaired in Prnp0/0 mice. Furthermore, we observed that

Acknowledgments

Research supported by FAPESP (2003/13189-2). B. Lobão-Soares is supported by a CAPES fellowship. M.M. Bianchin is supported by FAPESP (02/03743-0). We would like to thank Dr. Charles Weissman for kindly providing the parental knockout mice to set up our breeding colony.

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