Impulsivity–anxiety-related behavior and profiles of morphine-induced analgesia in heterozygous reeler mice

Abstract

Reelin is an extracellular matrix protein, secreted by GABAergic interneurons, that provides a signal for neural plasticity. A downregulation of reelin may be a factor to be considered in the study of major psychiatric disorders. The heterozygous reeler mouse model, thus, may be important to reveal those alterations in behavioral phenotype produced by reduced neural plasticity. Heterozygous (HZ) and wild-type (WT) mice were tested for anxiety-related behavior, motor impulsivity, and morphine-induced analgesia. Heterozygous mice showed significantly lower levels of anxiety- and risk-assessment-related behaviors in the elevated plus-maze during adolescence, in the absence of basal changes in general locomotion. Adult mice were assessed for profiles of impulsive behavior in operant chambers, and HZ mice exhibited elevated levels of motor impulsivity. When mice were assessed in nociception tests, a genotype difference in morphine-induced analgesia was found, and these results were confirmed by measurement of μ-receptors in the midbrain. The basal behavioral profile of the HZ genotype reveals important differences, consistent with decreased behavioral inhibition and emotionality, which can be revealed as early as in adolescence, together with slight increment of impulsive behavior and altered pain threshold and at the adult age. The HZ genotype can thus represent a useful animal model for the study of behavioral disorders consequent to reduced neural plasticity.

Introduction

Studies of molecular biology revealed that Reelin, a protein of the extracellular matrix, plays a function of control in the migration and positioning of central nervous system cells in the embryonic period. Reelin seems to have a role also in adulthood, in synaptogenesis, and in providing a signal for neural plasticity (Costa et al., 2001, Costa et al., 2002). These evidences are confirmed by in vitro studies on the homozygous reeler mouse. Interestingly, a 50% downregulation of reelin expression (mRNA and protein), exhibited by heterozygous reeler mice (HZ), compared to wild-type (WT) littermates, may inhibit dendritic spine maturation and may lead to atrophy of postsynaptic spines (D'Arcangelo and Curran, 1998, D'Arcangelo et al., 1995, Tueting et al., 1999). These neuroanatomical characteristics, together with decreased neuropil and GAD₆₇ expression, are quite similar to the brain abnormalities found postmortem in schizophrenic patients (Carboni et al., 2004, Guidotti et al., 2000). Indeed, among the possible causes of schizophrenia, and other major psychiatric diseases, recent research included a decrease of reelin gene expression (Fatemi et al., 2001, Laviola et al., 2006, Ognibene et al., submitted for publication). Due to similarity in neurochemical features and behavioral impairment, HZ reeler mice were proposed by Costa and colleagues (2002) as a useful model for the above mentioned diseases.

The reeler mouse model may be important to reveal alterations in behavioral phenotype produced by a reduction of neural plasticity due to reelin deficiency (Gainetdinov et al., 2001, Impagniatello et al., 1998). The present study attempts to deeper characterize the behavior of the heterozygous reeler mice, in comparison with the wild-type ones, and to confirm the neurodevelopmental hypotheses. According to these theories, in fact, an abnormal brain development leads to an abnormal behavior (Bartlett et al., 2005, Costa et al., 2001). Although the HZ reeler mice have been previously assessed in several behavioral paradigms by Salinger and colleagues (2003), there are still some aspects that need to be investigated.

The behavioral alterations caused by schizophrenia have their onset during pubertal period (Woods, 1998), for this reason our experimental design started when mice were about 35 days old, during the adolescence period (Laviola et al., 1999, Laviola et al., 2003). Mice of the two genotypes were assessed in the open field, with a novelty paradigm, and levels of anxiety-related and risk assessment behaviors were also measured in the elevated plus-maze apparatus. These parameters have been already measured by Salinger and colleagues (2003), and more recently by Qiu and colleagues (2006), who reported no differences between HZ and WT mice. Nevertheless, since both authors have conducted testing on adult animals, our aim was to investigate the same behaviors in non fully mature animals (Macrì et al., 2002). Indeed, genotype-dependent differences in behavioral profile during adolescence could represent an index for a precocious onset of neurobehavioral anomalies in the HZ mice.

When adult, mice were assessed for levels of motor impulsivity. Many different aspects of impulsivity have been studied with operant behavior paradigms in laboratory settings since impulsivity can be defined in several ways (Evenden, 1999). Two main categories of impulsivity are (a) the failure to resist an impulse, drive, or temptation (“motor impulsivity”), and (b) decision making without consideration of alternatives and/or consequences (“impulsive choice”). In the present study, we investigated motor impulsivity with a one-choice paradigm, requiring animals to refrain from responding during a signaled delay interval preceding the reward delivery (Adriani et al., 2006). Such kind of tests, where inappropriate responding during a waiting interval is punished (Brockel and Cory-Slechta, 1998), allows evaluating motor impulsivity. It is well known that self-control and inhibition of behavior are dependent on integrity of 5-HT and DA pathways (Linnoila et al., 1983, Soubriè, 1986), and brain dopaminergic changes in HZ mice, compared to WT have been reported by Ballmaier and colleagues (2002). As a consequence, we hypothesized that the monoaminergic alterations present in the heterozygous reeler mouse could affect their ability to inhibit behavioral actions. Impulsivity, indeed, is common in schizophrenic patients, and it could be a valuable tool to assess the impulsive-related profile of HZ reeler mice since they have been proposed as a model of psychosis vulnerability.

One further contribution of the present study was the evaluation of basal differences in nociception between HZ reeler mouse and WT controls. Pain perception is a complex process, influenced by a variety of environmental and genetic factors (Linnoila et al., 1983, Diatchenko et al., 2005). We investigated if the genotype was able to influence the nociceptive threshold of mice using thermal nociceptive stimuli (Podhorna and Didriksen, 2003, Qiu et al., 2006). In this case, as it will be discussed later, our aim was to investigate whether there was a difference also in spinal nociception using the tail flick test, and if pharmacological stimulation with morphine could unmask dissimilar responses in the two genotypes.