Effects of REM deprivation and an NMDA agonist on the extinction of conditioned fear

Abstract

Rapid eye movement sleep (REM) has been implicated in a number of learning and memory tasks. Previous research has demonstrated that REM deprivation impairs the development of extinction of conditioned fear responses. However, the neurobiological mechanisms of this effect remain unclear. The present study investigated the effects of systemic administration of d-cycloserine (DCS), an NMDA agonist, on the extinction of a conditioned fear response following 6 h of REM deprivation. In experiment 1, rats were administered DCS between fear training and REM deprivation. In experiment 2, rats were administered DCS prior to extinction training. The results of experiment 1 indicated that both DCS alone and REM deprivation alone impaired extinction learning. Administration of DCS to REM deprived animals partially, but not completely, reversed the deficit in extinction. The results of experiment 2 indicated that regardless of prior REM deprivation history, DCS facilitated extinction learning. The results provide further evidence for a role of REM in the extinction of cued fear learning and indicate that this effect appears to be partially mediated by NMDA-dependent mechanisms.

Introduction

For many years researchers have investigated the possibility that one function of sleep is to facilitate learning and memory processes. While this area of research is not without controversy, a growing body of literature suggests that a relationship exists between sleep and learning. Both human and animal studies have shown that sleep deprivation affects certain types of learning, and that learning can change subsequent sleep patterns [1].

There are a number of ways to assess the role of sleep in learning and memory processes in animals. One is to record sleep patterns after acquisition of a learning task. Increases in rapid eye movement sleep (REM) have been seen after avoidance task training [2], [3] and spatial learning [4]. Decreases in REM are seen when sleep is recorded immediately following fear conditioning with inescapable shock [5], [6] and when sleep is recorded in the same context in which fear conditioning previously took place [7]. Therefore, changes in post-learning REM depend on a number of variables, including the type of learning task and the context in which sleep is recorded. Although it is difficult to draw simple conclusions due to the lack of uniformity in these findings, these results may illustrate the complexity of the relationship between sleep and learning and suggest that this relationship is deserving of further investigation to delineate the conditions under which learning can cause an increase or decrease in subsequent REM.

Another means by which researchers have investigated the relationship between sleep and learning is to measure learning and memory processes after a period of increased REM. One such study examined the effects of experimentally-induced REM enhancement on memory for a brightness discrimination task [8]. Rats were exposed to discrimination training, which was followed by one of three methods of REM enhancement: carbachol infusion into the pontine reticular formation, corticotropin-like intermediate lobe peptide infusion into the ventricles, or REM rebound produced by REM deprivation. All three methods of REM enhancement produced a significant increase in the retention of the discrimination task when tested 24 h later. Additionally, a significant positive correlation was observed between retention scores and the amount of time spent in REM after training.

A third way to assess this relationship is to deprive animals of sleep around the time of learning or recall of a task. REM deprivation either before or after training has been shown to impair the retention of a spatial learning task [4], [9], [10], [11], [12], [13]. Additionally, animals deprived of REM prior to either cued or contextual conditioning demonstrated impaired fear conditioning during a retention test [14].

REM deprivation has also been shown to impair the extinction of a previously learned conditioned response. In a recent investigation [15], rats were trained in a fear conditioning paradigm which was followed immediately by a single session of REM deprivation or a control condition in which REM was unaffected. Forty-eight hours later, all animals were presented with extinction trials. While memory for fear conditioning was unchanged by REM deprivation, extinction was impaired in REM deprived animals. Animals in the control condition demonstrated a reduction in freezing as the extinction session progressed, indicating normal extinction. However, REM deprived animals continued to freeze at high levels throughout the extinction session, indicating impaired extinction. During a second extinction session conducted 48 h later, REM deprived and control animals showed no differences in rates of freezing.

Extinction, typically defined as a reduction in the strength of the association between the conditioned stimulus (CS) and unconditioned stimulus (US), reflects the creation of a new memory. The occurrence of phenomena such as spontaneous recovery, renewal, and reinstatement, in which an extinguished conditioned response (CR) re-emerges, indicates that extinction training does not erase the memory of the initial CS–US association but instead creates a new memory (for review, see [16]). Therefore, following extinction training an organism has a memory for both conditioning and extinction. Following conditioning, presentation of the CS activates the amygdala, which controls expression of CRs such as freezing. Extinction training produces long-term potentiation-like changes in the medial prefrontal cortex (mPFC; [17], [18]), which in turn projects to the amygdala [19], [20]. The mPFC–amygdala projection is likely to be inhibitory in nature and may provide the neurobiological basis for the inhibition of CRs following extinction training [21].

A number of neurotransmitters have been implicated in extinction, including glutamate [22]. More specifically, activation of NMDA receptors seems to be integral to the development of extinction. Infusion of NMDA antagonists either peripherally or intra-amygdala impairs extinction [23], [24], [25], [26]. Most studies indicate that administration of an NMDA agonist facilitates extinction. Systemic or intra-amygdala administration of d-cycloserine (DCS), a partial agonist of the NMDA receptor complex at the glycine recognition site [27], [28], administered prior to extinction training enhanced extinction of a conditioned freezing response [29]. Post-extinction administration of DCS also enhanced extinction in a subsequent test, thereby implicating DCS in the consolidation of extinction learning [29]. Peripheral or intra-amygdala infusion of DCS also enhanced extinction of fear-potentiated startle [30].

In addition to their well-known role in learning and memory processes, NMDA receptors have a role in REM sleep. Single intracerebroventricular infusions of the NMDA antagonists MK-801 or AP5 reduced REM with no significant effect on slow-wave sleep or wakefulness [31], [32]. Systemic injections of the NMDA antagonist NPC12626 produced reduced time spent in REM at all doses tested, with effects on other sleep stages only at higher doses [33]. The effects of NMDA agonists such as DCS on sleep are currently unknown.

The present studies were designed to begin to elucidate the neurobiological mechanisms underlying the impairment of extinction due to short-term REM deprivation [15]. REM deprivation produces a variety of neurobiological effects that could be responsible for the previously observed extinction impairment. Therefore, it may be possible to reverse this impairment with a manipulation performed between training and REM deprivation. This strategy has proven successful in previous research, in which activation of the pontine-wave generator performed between training in an active avoidance task and 6 h of REM deprivation reversed the learning impairment that occurred without such activation [34]. Based on the demonstrated facilitation of extinction due to administration of NMDA agonists [29], [30] as well as the involvement of NMDA receptors in REM sleep [31], [32], [33], the NMDA agonist DCS was administered in an attempt to reverse the deficit in extinction seen following REM deprivation.