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The authors observed brief, directed movement to a familiar enclosure in rats to determine whether this behavior is part of a rat's defensive repertoire when exposed to a conditional-fear stimulus. In Experiment 1, upon exposure to the compound conditional-fear stimulus of tone and light, only rats that received paired presentations of the conditional stimuli and shock fled into a small, familiar enclosure where they then froze. Rats that had received unpaired presentations did not enter the enclosure in significant amounts when later tested. In Experiment 2, the authors observed rats' freezing and use of either a familiar or an unfamiliar enclosure when tested with a conditional-fear stimulus. Rats tested with a familiar enclosure entered it more quickly than did rats without prior exposure to the enclosure. Freezing was greatest when both training and testing environments were similar with respect to access to the enclosure. The results of these 2 experiments support the idea that brief, directed flight in rats is a component of the postencounter stage of predatory imminence (M. S. Fanselow & L. S. Lester, 1988) and is compatible with freezing.

Background: Enhancement of the function of gamma-aminobutyric acid type A receptors containing the alpha1 subunit may underlie a portion of inhaled anesthetic action. To test this, the authors created gene knock-in mice harboring mutations that render the receptors insensitive to isoflurane while preserving sensitivity to halothane., Methods: The authors recorded miniature inhibitory synaptic currents in hippocampal neurons from hippocampal slices from knock-in and wild-type mice. They also determined the minimum alveolar concentration (MAC), and the concentration at which 50% of animals lost their righting reflexes and which suppressed pavlovian fear conditioning to tone and context in both genotypes., Results: Miniature inhibitory postsynaptic currents decayed more rapidly in interneurons and CA1 pyramidal cells from the knock-in mice compared with wild-type animals. Isoflurane (0.5-1 MAC) prolonged the decay phase of miniature inhibitory postsynaptic currents in neurons of the wild-type mice, but this effect was significantly reduced in neurons from knock-in mice. Halothane (1 MAC) slowed the decay of miniature inhibitory postsynaptic current in both genotypes. The homozygous knock-in mice were more resistant than wild-type controls to loss of righting reflexes induced by isoflurane and enflurane, but not to halothane. The MAC for isoflurane, desflurane, and halothane did not differ between knock-in and wild-type mice. The knock-in mice and wild-type mice did not differ in their sensitivity to isoflurane for fear conditioning., Conclusions: gamma-Aminobutyric acid type A receptors containing the alpha1 subunit participate in the inhibition of the righting reflexes by isoflurane and enflurane. They are not, however, involved in the amnestic effect of isoflurane or immobilizing actions of inhaled agents.

Signaling by the Ca(2+)/calmodulin kinase (CaMK) cascade has been implicated in neuronal gene transcription, synaptic plasticity, and long-term memory consolidation. The CaM kinase kinase alpha (CaMKKalpha) isoform is an upstream component of the CaMK cascade whose function in different behavioral and learning and memory paradigms was analyzed by targeted gene disruption in mice. CaMKKalpha mutants exhibited normal long-term spatial memory formation and cued fear conditioning but showed deficits in context fear during both conditioning and long-term follow-up testing. They also exhibited impaired activation of the downstream kinase CaMKIV/Gr and its substrate, the transcription factor cyclic AMP-responsive element binding protein (CREB) upon fear conditioning. Unlike CaMKIV/Gr-deficient mice, the CaMKKalpha mutants exhibited normal long-term potentiation and normal levels of anxiety-like behavior. These results demonstrate a selective role for CaMKKalpha in contextual fear memory and suggest that different combinations of upstream and downstream components of the CaMK cascade may serve distinct physiological functions.

Morphine analgesic tolerance is heritable in both humans and rodents, with some individuals and strains exhibiting little and others exhibiting robust tolerance. 129S6/SvEv and 129P3/J mice reportedly do not demonstrate tolerance to morphine analgesia. Using our laboratory's standard morphine tolerance regimen and a between-subjects design, tolerance developed in the hot plate and tail withdrawal assays as indicated by a change in analgesic efficacy following a morphine challenge dose. Furthermore, the non-competitive NMDA receptor antagonist MK-801 (dizocilipine) blocked morphine tolerance in 129S6/SvEv and CD-1 mice in the hot plate assay. As previously reported, when a within-subjects design and cumulative dosing was employed, no tolerance was observed in the 129P3/J strain. However, using the same morphine regimen and a between-subjects design, comparable tolerance developed between 129P3/J and C57BL/6J strains following a single challenge dose of morphine. Spontaneous hyperalgesia was observed in the tail withdrawal assay following chronic morphine in C57BL/6J, but not 129P3/J mice. Additionally, morphine-tolerant C57BL/6J mice, but not 129P3/J mice, exhibited a large increase in the frequency of tail flicks during the first second following the baseline nociceptive response which may facilitate detection of the response during the tolerant state. We conclude that the method of tolerance assessment affects the ability to detect tolerance and thus may affect the degree and pattern of heritability of this trait and this could have implications for gene mapping studies.

Multiple studies demonstrate that coadministration of N-methyl-D-aspartate (NMDA) receptor antagonists with the opioid agonist morphine attenuates the development of analgesic tolerance. Sex differences in the effects of noncompetitive, but not competitive NMDA receptor antagonists on acute morphine analgesia, have been reported in mice, yet the role of sex in modulation of morphine tolerance by NMDA receptor antagonists has yet to be addressed. Therefore, we tested whether there is a sex difference in the effect of NMDA receptor antagonists on the development of morphine analgesic tolerance in C57BL/6J mice. Acutely, at a dose required to affect morphine tolerance in male mice, the noncompetitive NMDA receptor antagonist dizocilpine (MK-801) prolonged morphine analgesia similarly in both sexes in the hot plate and tail withdrawal assays. In the hot plate assay, coadministration of MK-801 or the competitive antagonist 3-(2-carboxpiperazin-4-yl)propyl-1-phosphanoic acid (CPP) with morphine attenuated the development of tolerance in male mice, while having no effect in females. Like normal and sham females, ovariectomized mice were similarly insensitive to the attenuation of morphine tolerance by MK-801 in the hot plate assay. Surprisingly, in the tail withdrawal assay, MK-801 facilitated the development of morphine-induced hyperalgesia and tolerance in males but not females. The results demonstrate that male mice are more sensitive to modulation of nociception and morphine analgesia after repeated coadministration of NMDA receptor antagonists. Furthermore, the underlying mechanisms are likely to be different from those mediating the sex difference in the modulation of acute morphine analgesia that has previously been reported.

Pavlovian contextual fear conditioning occurs when an aversive unconditional stimulus (US), such as a footshock, is presented to a rat shortly after it is placed in an experimental context. Contextual fear conditioning does not occur when the shock is presented immediately upon placement of the rat in the novel chamber. In the present study, the authors report that increasing either the number of immediate shock sessions (Experiment 1) or the immediate shock duration (Experiment 2) did not reverse this deficit. However, immediate shock seems to sensitize subsequent context conditioning (Experiment 3). These findings suggest that the associative deficit produced by immediate shock is not related to the rat's ability to process the footshock US., (Copyright 2006 APA, all rights reserved.)

Lesions of the rodent hippocampus invariably abolish context fear memories formed in the recent past but do not always prevent new learning. To better understand this discrepancy, we thoroughly examined the acquisition of context fear in rats with pretraining excitotoxic lesions of the dorsal hippocampus. In the first experiment, animals received a shock immediately after placement in the context or after variable delays. Immediate shock produced no context fear learning in lesioned rats or controls. In contrast, delayed shock produced robust context fear learning in both groups. The absence of fear with immediate shock occurs because animals need time to form a representation of the context before shock is presented. The fact that it occurs in both sham and lesioned rats suggests that they learn about the context in a similar manner. However, despite learning about the context in the delay condition, lesioned rats did not acquire as much fear as controls. The second experiment showed that this lesion-induced deficit could be overcome by increasing the number of conditioning trials. Lesioned animals learned normally after multiple shocks, regardless of freezing level or trial spacing. The last experiment showed that animals with complete hippocampus lesions could also learn about the context, although the same lesions produced devastating retrograde amnesia. These results demonstrate that alternative systems can acquire context fear but do so less efficiently than the hippocampus.

Rats confronted with the onset of a light gradient display a transient increase in locomotion called the activity response (AR) and a dark preference (Godsil & Fanselow, 2004). These experiments demonstrate that the magnitude of the AR can be blunted with Pavlovian fear-conditioning procedures via associative and nonassociative fear. Although manifested in decreased locomotion, the blunted AR effect was not due to increased freezing or immobility behaviors. Instead, rats displayed reduced rearing and an increase in a class of behaviors called stationary activity. These results suggest that the lighting differential supplied by the cue influences the topography of defensive behavior and reduces the expression of freezing. This procedure provides a means by which to examine learned and unlearned defensive responses to the same stimulus.

The hippocampus has been implicated in anxiety, novelty detection, spatial- contextual processing, and hyperactivity. Accordingly, the authors contrasted the role of the dorsal hippocampus (DH) and the basolateral amygdala complex (BLA) in an open field task that presents the onset and termination of a bright light gradient. In the dark, DH rats demonstrated impaired habituation of locomotion behavior and hyperactivity, whereas in bright light their behaviors were normal. DH rats responded differentially to the onset and termination of the light stimulus, which indicates they have normal novelty detection. BLA lesion rats responded normally to bright light. These results demonstrate that a mild fear stimulus, such as bright light, can suppress DH lesion-evoked hyperactivity, and this hyperactivity results from impaired contextual processing.

Placing a "trace" interval between a warning signal and an aversive shock makes consolidation of the memory for trace conditioning hippocampus dependent. To determine the trace at which memory consolidation requires the hippocampus, mice were trained with 0-s, 1-s, 3-s, or 20-s trace intervals and tested for freezing to context and tone. Posttraining dorsal hippocampus (DH) lesions decreased context conditioning regardless of trace interval. However, DH lesions attenuated only the 20-s trace tone freezing. Like eyeblink conditioning, the DH is necessary for trace fear conditioning only at long trace intervals, but the time scale for the effective interval in fear conditioning is about 40 times longer. Manipulations that alter trace fear conditioning with short trace intervals probably do not reflect altered DH function. Given this difference in time scale along with the use of posttraining DH lesions, hippocampus dependency of trace conditioning is not related to a bridging function or response timing.

Inhaled anesthetics are believed to produce anesthesia by their actions on ion channels. Because inhaled anesthetics robustly enhance GABA A receptor (GABA(A)-R) responses to GABA, these receptors are considered prime targets of anesthetic action. However, the importance of GABA(A)-Rs and individual GABA(A)-R subunits to specific anesthetic-induced behavioral effects in the intact animal is unknown. We hypothesized that inhaled anesthetics produce amnesia, as assessed by loss of fear conditioning, by acting on the forebrain GABA(A)-Rs that harbor the alpha1 subunit. To test this, we used global knockout mice that completely lack the alpha1 subunit and forebrain-specific, conditional knockout mice that lack the alpha1 subunit only in the hippocampus, cortex, and amygdala. Both knockout mice were 75 to 145% less sensitive to the amnestic effects of the inhaled anesthetic isoflurane. These results indicate that alpha1-containing GABA(A)-Rs in the hippocampus, amygdala, and/or cortex influence the amnestic effects of inhaled anesthetics and may be an important molecular target of the drug isoflurane.

The mu opioid receptor may constitute a critical component of a negative feedback system that regulates Pavlovian fear conditioning. We investigated context fear conditioning acquisition and expression in mu opioid receptor knockout mice (on an inbred, C57 genetic background). We discovered that the mu receptor knockout results in an unexpected and significant deficit in context fear acquisition. Mice lacking the mu receptor showed normal fear acquisition when subjected to a 1-day fear conditioning protocol but evinced deficient fear learning when acquisition was conducted across 5 days. The knockout mice showed normal reactivity to footshock in both fear conditioning protocols. Finally, we confirmed the effectiveness of the receptor deletion in the C57 strain by subjecting the mice to a test of morphine analgesia in the hot-plate assay. As has been seen with mixed genetic background, the receptor deletion resulted in a complete lack of analgesic response to 10 mg/kg morphine. Surprisingly, mice with a single copy of the mu receptor gene (heterozygous knockouts) showed intact sensitivity to morphine but a significant deficit in Pavlovian fear conditioning. The results indicate that deletion of the mu receptor gene impairs fear conditioning and that the conditioning and analgesia effects of heterozygous deletion are dissociable. The conditioning deficit seen in this line of mice may be related to impairment in hippocampus function.

Several studies show that the hippocampus is critical for the memories mediating trace and contextual fear conditioning. This study investigates whether N-methyl-D-aspartate-induced lesions of the dorsal hippocampus made prior to training affect context fear conditioning and trace fear conditioning measured with the fear-potentiated startle. Pretraining excitotoxic lesions of the dorsal hippocampus blocked acquisition of trace fear conditioning to a tone stimulus but did not affect context fear conditioning. These data indicate that without a dorsal hippocampus rats are unable to acquire trace conditioning but can acquire contextual fear when fear is measured by potentiation of the startle response., ((c) 2005 APA, all rights reserved.)

Previous reports suggest that the administration of epinephrine increases learning during deep barbiturate-chloral hydrate anesthesia in rats but not during anesthesia with 0.4% isoflurane in rabbits. We revisited this issue, using fear conditioning to a tone in rats as our experimental model for learning and memory and isoflurane and desflurane as our anesthetics. Expressed as a fraction of the minimum alveolar anesthetic concentration (MAC) preventing movement in 50% of rats, the amnestic 50% effective dose (ED(50)) for fear to tone in control rats inhaling isoflurane and injected with saline intraperitoneally (i.p.) was 0.32 +/- 0.03 MAC (mean +/- se) compared with 0.37 +/- 0.06 MAC in rats injected with 0.01 mg/kg of epinephrine i.p. and 0.38 +/- 0.03 MAC in rats injected with 0.1 mg/kg of epinephrine i.p. For desflurane, the amnestic ED(50) were 0.32 +/- 0.05 MAC in control rats receiving a saline injection i.p. versus 0.36 +/- 0.04 MAC in rats injected with 0.1 mg/kg of epinephrine i.p. We conclude that exogenous epinephrine does not decrease amnesia produced by inhaled isoflurane or desflurane, as assessed by fear conditioning to a tone in rats.

The delta subunit of the GABA(A) receptor (GABA(A)R) is highly expressed in the dentate gyrus of the hippocampus. Genetic deletion of this subunit reduces synaptic and extrasynaptic inhibition and decreases sensitivity to neurosteroids. This paper examines the effect of these changes on hippocampus-dependent trace fear conditioning. Compared to controls, delta knockout mice exhibited enhanced acquisition of tone and context fear. Hippocampus-independent delay conditioning was normal in these animals. These results suggest that reduced inhibition in the dentate gyrus facilitates the acquisition of trace fear conditioning. However, the enhancement in trace conditioning was only observed in female knockout mice. The sex-specificity of this effect may be a result of neuroactive steroids. These compounds vary during the estrus cycle, can increase GABAergic inhibition, and have been shown to impair hippocampus-dependent learning. We propose that activation of GABA(A)Rs by neuroactive steroids inhibits learning processes in the hippocampus. Knockouts are immune to this effect because of the reduced neurosteroid sensitivity that accompanies deletion of the delta subunit. Relationships between neurosteroids, hippocampal excitability, and memory are discussed.

Fear is an adaptive response that initiates defensive behavior to protect animals and humans from danger. However, anxiety disorders, such as Posttraumatic Stress Disorder (PTSD), can occur when fear is inappropriately regulated. Fear conditioning can be used to study aspects of PTSD, and we have developed a model in which pre-exposure to a stressor of repeated footshock enhances conditional fear responding to a single context-shock pairing. The experiments in this chapter address interpretations of this effect including generalization and summation or fear, inflation, and altered pain sensitivity. The results of these experiments lead to the conclusion that pre-exposure to shock sensitizes conditional fear responding to similar less intense stressors. This sensitization effect resists exposure therapy (extinction) and amnestic (NMDA antagonist) treatment. The pattern predicts why in PTSD patients, mild stressors cause reactions more appropriate for the original traumatic stressor and why new fears are so readily formed in these patients. This model can facilitate the study of neurobiological mechanisms underlying sensitization of responses observed in PTSD.

The dorsal hippocampus (DH) is critically involved in the acquisition and expression of trace and contextual fear conditioning. NMDA/glutamate receptor-mediated transmission is thought to be one mechanism mediating the plastic changes that support long-term memories in the DH. However, their precise involvement in acquisition and expression processes has not been defined. To examine this issue, the NMDA receptor antagonist, D,L-2-amino-5-phosphonovaleric acid (APV; 10 microg/microl; 0.5 microl), was infused into the DH prior to conditioning and/or testing, using a trace fear conditioning procedure. All rats were tested for freezing to both tone and context in separate, counterbalanced sessions. The three sessions (1 training and 2 test) were separated by approximately 24 h. Using this design, it was possible to assess the role for DH NMDA receptors in the acquisition versus expression of trace and contextual fear conditioning. APV disrupted acquisition, but not expression, of contextual fear conditioning. By contrast, APV attenuated both acquisition and expression of trace fear memories. Thus, DH NMDA receptors appear to contribute to retrieval of some, but not all, fear memories., (Copyright 2005 Wiley-Liss, Inc.)

Mammalian associative learning is organized into separate anatomically defined functional systems. We illustrate the organization of two of these systems, Pavlovian fear conditioning and Pavlovian eyeblink conditioning, by describing studies using mutant mice, brain stimulation and recording, brain lesions and direct pharmacological manipulations of specific brain regions. The amygdala serves as the neuroanatomical hub of the former, whereas the cerebellum is the hub of the latter. Pathways that carry information about signals for biologically important events arrive at these hubs by circuitry that depends on stimulus modality and complexity. Within the amygdala and cerebellum, neural plasticity occurs because of convergence of these stimuli and the biologically important information they predict. This neural plasticity is the physical basis of associative memory formation, and although the intracellular mechanisms of plasticity within these structures share some similarities, they differ significantly. The last Annual Review of Psychology article to specifically tackle the question of mammalian associative learning ( Lavond et al. 1993 ) persuasively argued that identifiable "essential" circuits encode memories formed during associative learning. The next dozen years saw breathtaking progress not only in detailing those essential circuits but also in identifying the essential processes occurring at the synapses (e.g., Bi & Poo 2001, Martinez & Derrick 1996 ) and within the neurons (e.g., Malinow & Malenka 2002, Murthy & De Camilli 2003 ) that make up those circuits. In this chapter, we describe the orientation that the neuroscience of learning has taken and review some of the progress made within that orientation.

Defensive responses to a cat were observed in rats given excitotoxic lesions of the central nucleus of the amygdala (ACe), dorsolateral periaqueductal gray (dlPAG), ventral periaqueductal gray (vPAG), or sham lesions. Rats were placed adjacent to a compartment containing a cat. Sham-lesioned rats avoided the area nearest the cat and preferred the area furthest away from the cat. They also exhibited numerous defensive responses including, climbing, escape from the apparatus, and freezing. Rats with lesions of the ACe reacted like the sham lesioned rats by preferring the area of the apparatus furthest from the cat, however they climbed and escaped significantly less than sham lesioned rats. Avoidance of the area adjacent to the cat was attenuated in rats with lesions of the vPAG. Climbing along the walls of the apparatus was also attenuated in rats with lesions of the vPAG. Escapes from the apparatus were not significantly reduced by lesions of the vPAG and dlPAG. Thus, ACe lesions attenuated climbing and eliminated escapes, but did not impair locomotion of the rat away from the cat.

Bright light may be a danger signal for rats because they are more vulnerable to predators in bright environments. We examined the fear-evoking properties of bright light with a novel open-field procedure that confronts a rat with the sudden onset or termination of a bright light gradient. The rats did not freeze but exhibited a transient increase in locomotion to light onset and termination, which we call the activity response (AR). This finding suggests that the AR is an exploratory response geared at investigating stimulus change. The rats also displayed a preference for dark to the lighting differential that was not due to the novelty or slight heating differential supplied by the lamps. These experiments demonstrate that the sudden onset of bright light engages preencounter defensive behavior, as described by the predatory imminence model (Fanselow & Lester, 1988). This task is amenable to studying light-evoked defensive responses.

The basolateral amygdala (BLA) is intimately involved in the development of conditional fear. Converging lines of evidence support a role for this region in the storage of fear memory but do not rule out a time-limited role in the memory consolidation. To examine this issue, we assessed the stability of BLA contribution to fear memories acquired across the adult lifetime of rats. Fear conditioning consisted of 10 tone-shock pairings in one context (remote memory), followed 16 months later by 10 additional tone-shock pairings with a novel tone in a novel context (recent memory). Twenty-four hours after recent training, rats were given NMDA or sham lesions of the BLA. Contextual and tone freezing were independently assessed in individual test sessions. Sham-lesioned rats showed high and comparable levels of freezing across all context and tone tests. In contrast, BLA-lesioned rats displayed robust freezing deficits across both recent and remote tests. Subsequent open-field testing revealed no effects of BLA lesions on activity patterns in a dark open field or during bright light exposure. Lesioned rats were able to reacquire normal levels of context-specific freezing after an overtraining procedure (76 unsignaled shocks). Together, these findings indicate that BLA lesions do not disrupt freezing behavior by producing hyperactivity, an inability to suppress behavior, or an inability to freeze. Rather, the consistent pattern of freezing deficits at both training-to-lesion intervals supports a role for the BLA in the permanent storage of fear memory.

Rats exposed to a footshock show conditional fear when reexposed to the shock context. Immediate presentation of shock after placement in the context significantly reduces this fear. Preexposure to the context in the absence of shock, coupled with a minimum preshock interval during training, overcomes this immediate shock deficit. Because rats learn about the context during preexposure and express that learning after being reinforced, the context preexposure effect is an aversive analogue of latent learning. The authors examined the effect of the N-methyl-D-aspartate (NMDA) receptor antagonist D,L-2-amino-5-phosphovalerate (APV) on the facilitatory effect of context preexposure. Rats were preexposed to a chamber after APV administration. The next day they were placed in the same chamber without drug and received shock 35 s later. APV blocked the facilitatory effect of preexposure. Therefore NMDA receptors are important for contextual latent learning.

Higher cognitive functions such as attention have been difficult to model in genetically tractable organisms. In humans, attention-distracting stimuli interfere with trace but not delay conditioning, two forms of associative learning. Attention has also been correlated with activation of anterior cingulate cortex (ACC), but its functional significance is unclear. Here we show that a visual distractor interferes selectively with trace but not delay auditory fear conditioning in mice. Trace conditioning is associated with increased neuronal activity in ACC, as assayed by relative levels of c-fos expression, and is selectively impaired by lesions of this structure. The effects of the ACC lesions are unlikely to be caused by indirect impairment of the hippocampus, which is required for mnemonic aspects of trace conditioning. These data suggest that trace conditioning may be useful for studying neural substrates of attention in mice, and implicate the ACC as one such substrate.

Ca(2+)-regulated gene transcription is essential to diverse physiological processes, including the adaptive plasticity associated with learning. We found that basal synaptic input activates the NF-kappa B transcription factor by a pathway requiring the Ca(2+)/calmodulin-dependent kinase CaMKII and local submembranous Ca(2+) elevation. The p65:p50 NF-kappa B form is selectively localized at synapses; p65-deficient mice have no detectable synaptic NF-kappa B. Activated NF-kappa B moves to the nucleus and could directly transmute synaptic signals into altered gene expression. Mice lacking p65 show a selective learning deficit in the spatial version of the radial arm maze. These observations suggest that long-term changes to adult neuronal function caused by synaptic stimulation can be regulated by NF-kappa B nuclear translocation and gene activation.

These experiments explore parallels between the neurobiological substrates of spatial and context learning. Male Long-Evans rats were employed in a context fear conditioning protocol that involved sequential acquisition and testing in two distinct contexts. Rats received three unsignaled footshocks in one context and were tested for context fear, measured as freezing, the next day. Four days later, the procedure was repeated in a different, distinct context. Rats received a hippocampal infusion of either the NMDA receptor antagonist 5-amino-phosphonovaleric acid (APV, 10 microg) or vehicle prior to training in each context. NMDA receptor blockade was effective in impairing context learning only in the first context. Context fear acquisition was not impaired by APV in the second context, indicating that pre-training (in the first context) mitigated the effects of APV. These data agree with those seen previously in the water maze, where pre-training prevented learning deficits produced by NMDA receptor blockade. The data thus suggest that the neurobiological substrates of context learning and place learning overlap.

Unlabelled: In animals, the conventional inhaled anesthetic, isoflurane, impairs learning fear to context and fear to tone, doing so at concentrations that produce amnesia in humans. Nonimmobilizers are inhaled compounds that do not produce immobility in response to noxious stimulation, nor do they decrease the requirement for conventional inhaled anesthetics. Like isoflurane, the nonimmobilizer 1,2-dichlorohexafluorocyclobutane (2N) impairs learning at concentrations less than those predicted from its lipophilicity to produce anesthesia. The capacity of the nonimmobilizer di-(2,2,2,-trifluoroethyl) ether (flurothyl) to affect learning and memory has not been studied. Both nonimmobilizers can cause convulsions. We hypothesized that if isoflurane, 2N, and flurothyl act by the same mechanism to impair learning and memory, their effects should be additive. We found that isoflurane, 2N, and flurothyl (each, alone) impaired learning fear to context and fear to tone in rats, with the nonimmobilizers doing so at concentrations less than those that cause convulsions. (Fear was defined by freezing [volitional immobility] in the presence of the conditioned stimulus [context or tone].) However, the combination of isoflurane and 2N or flurothyl produced an antagonistic rather than an additive effect on learning, a finding in conflict with our hypothesis. And flurothyl was no less potent than 2N (at least no less potent relative to the concentration of each that produced convulsions) in its capacity to impair learning. We conclude that conventional inhaled anesthetics and nonimmobilizers impair learning and memory by different mechanisms. The basis for this impairment remains unknown., Implications: Conventional inhaled anesthetics and nonimmobilizers are antagonistic in their effects on learning and memory, and this finding suggests that they impair learning and memory, at least in part, by different mechanisms.

Lesions of the frontotemporal region of the amygdala, which includes lateral and basal nuclei, cause a loss of conditional fear responses, such as freezing, even when the lesions are made over a year and a half from the original training. These amygdala-damaged animals are not hyperactive and show normal reactivity to strong stimuli such as bright lights. After receiving tone-mild shock pairings rats normally display an appropriately weak response when exposed to the tone. Rats' fear of the tone can be inflated by giving them exposure to strong shocks in the absence of the tone between training and testing. This inflation of fear memory is abolished if the frontotemporal amygdala is inactivated by muscimol only during the inflation treatment with strong shocks. Based on such findings we suggest that the frontotemporal amygdala permanently encodes a memory for the hedonic value of the aversive stimulus used to condition fear.

Pavlovian fear conditioning is a phenomenon amenable to laboratory analysis of the neurobiology of fear and the investigation of neural mechanisms of learning and memory. Investigators have made much progress in delineating the neurocircuitry and neurochemistry of fear conditioning. The place of the hippocampus in context fear remains a controversial issue. In this review, we examine the evidence that the hippocampus plays a role in fear conditioning. We then critically examine hypotheses concerning its exact role in learning and memory for cued and context fear conditioning.

Rats were conditioned across 2 consecutive days where a single unsignaled footshock was presented in the presence of specific contextual cues. Rats were tested with contexts that had additional stimulus components either added or subtracted. Using freezing as a measure of conditioning, removal but not addition of a cue from the training context produced significant generalization decrement. The results are discussed in relation to the R. A. Rescorla and A. R. Wagner (1972), J. M. Pearce (1994), and A. R. Wagner and S. E. Brandon (2001) accounts of generalization. Although the present data are most consistent with elemental models such as Rescorla and Wagner, a slight modification of the Wagner-Brandon replaced-elements model that can account for differences in the pattern of generalization obtained with contexts and discrete conditional stimuli is proposed.

Background: Production of retrograde amnesia by anesthetics would indicate that these drugs can disrupt mechanisms that stabilize memory. Such disruption would allow suppression of memory of previous untoward events. The authors examined whether isoflurane provides retrograde amnesia for classic (Pavlovian) fear conditioning., Methods: Rats were trained to fear tone by applying three (three-trial) or one (one-trial) tone-shock pairs while breathing various constant concentrations of isoflurane. Immediately after training, isoflurane administration was either discontinued, maintained unchanged, or rapidly increased to 1.0 minimum alveolar concentration for 1 h longer. Groups of rats were similarly trained to fear context while breathing isoflurane by applying shocks (without tones) in a distinctive environment. The next day, memory for the conditioned stimuli was determined by presenting the tone or context (without shock) and measuring the proportion of time each rat froze (appeared immobile). For each conditioning procedure, the effects of the three posttraining isoflurane treatments were compared., Results: Rapid increases in posttraining isoflurane administration did not suppress conditioned fear for any of the training procedures. In contrast, isoflurane administration during conditioning dose-dependently suppressed conditioning (P < 0.05). Training to tone was more resistant to the effects of isoflurane than training to context (P < 0.05), and the three-trial learning procedure was more was more resistant than the one-trial procedure (P < 0.05)., Conclusions: Isoflurane provided intense dose-dependent anterograde but not retrograde amnesia for classic fear conditioning. Isoflurane appears to disrupt memory processes that occur at or within a few minutes of the conditioning procedure.

Unlabelled: The nonimmobilizer 1,2-dichlorohexafluorocyclobutane (2N, also termed F6) does not suppress movement to noxious stimuli but does suppress learning of fear-potentiated startle. The mechanism whereby 2N suppresses this learning is unknown. Herein, we report the effect of 2N on suppression of two other forms of learning, fear conditioning to context and to tone. Because 2N does not cause sedation, we could study the effect of 2N on short-term memory (memory for fear conditioning measured during or immediately after training) as well as on long-term memory (measured 24 h after training). The EC(50) for suppression of long-term memory (the concentration decreasing memory by 50%) of fear conditioning to context was 2.00% plus/minus 0.01% (mean plus/minus SEM), and for fear conditioning to tone was 3.45% plus/minus 0.26%, (P < 0.05). The EC(50) for suppression of short-term memory of fear conditioning to context was 2.59% plus/minus 0.21% (P < 0.05, compared with long-term memory of context conditioning), whereas short-term memory of fear conditioning to tone was not suppressed by 3.5%, the largest concentration studied. Thus, short-term memory resists the depressant effect of 2N more than long-term memory, fear conditioning to tone is less vulnerable to the effect of 2N than fear conditioning to context, and 3.5% 2N does not preclude transmission of tone and shock signals to the site where tone-shock associations are formed., Implications: The nonimmobilizer 1,2-dichlorohexafluorocyclobutane has a greater depressant effect on long-term memory than short-term memory, suggesting that it impairs the processes responsible for the retention of memory more than for the formation of memory itself.

The present study sought to determine whether post-training excitotoxic lesions of the dorsal hippocampus would disrupt retention of fear conditioned using a trace procedure. Rats were trained using one of six procedures. Forward trace conditioning consisted of 10 trials in which a 16-s tone conditional stimulus (CS) was followed by a 28-s stimulus-free trace interval and then a mild footshock unconditional stimulus (US). We used two forms of delay conditioning where the tone and footshock co-terminated. Short delay used a 16-s tone and long delay used a 46-s tone. Backward trace conditioning was the same as forward trace, except that the order of the CS and US was reversed. CS-only and US-only were similar to forward trace except that the footshock or tone, respectively, was eliminated. One day later, animals received either an N-methyl-D-aspartate (NMDA)-induced lesion of the dorsal hippocampus or sham surgery. One week later, the rats were tested for freezing to the tone in a novel context. The next day, they were tested for freezing to the original training context. Hippocampal lesioned trace conditioned rats showed significantly less freezing during the tone compared with their sham lesioned controls. The lesion did not affect freezing during the tone in delay conditioning, nor in the other training conditions. During the 1-min period after tone offset, there was a trend in all hippocampal lesioned animals toward a deficit in freezing, compared with their corresponding sham lesioned controls, although only short delay, forward and backward trace groups showed a significant deficit. Hippocampal lesions also attenuated contextual conditioning. Thus, the hippocampus is critical for the consolidation and/or expression of a trace fear conditioned stimulus.

Background: Recent reports suggest that one type of learning, fear conditioning to context, requires more neural processing than a related type, fear conditioning to tone. To determine whether these types of learning were differentially affected by anesthesia, the authors applied isoflurane during the training phases of fear conditioning paradigms for freezing to context and freezing to tone., Methods: The authors trained seven groups of eight rats to fear tone by administering a tone (conditioned stimulus) while breathing various concentrations of isoflurane from 0.00 to 0.75 minimum alveolar concentration (MAC; one concentration per group) separated by 0.12-MAC steps. On the succeeding day, and in the absence of isoflurane, the authors presented the tone (without shock) in a different context (different cage shape and odor) and measured the time each rat froze (became immobile). Six other groups of eight rats were trained to fear context by applying the shock in the absence of a tone but in the presence of environmental cues such as cage shape, texture, and odor. Fear to context was determined the succeeding day by returning the rat to the training cage (without shock) and measuring duration of freezing. Control groups (16 per group) received 0.75 MAC isoflurane but no foot shocks. Group scores were compared using analysis of variance, and the ED50 values for quantal responses of individual rats were calculated using logistic regression., Results: Conditioning to context occurred at 0.00 and 0.13 MAC (P < 0.05 compared with unshocked control) but not 0.25 MAC; the ED50 was 0.25 +/- 0.03 MAC (mean +/- SEM). In contrast, conditioning to tone occurred at 0.48 MAC (P < 0.05) but not 0.62 MAC; the ED50 was 0.47 +/- 0.02 MAC (P < 0.01 for the difference between ED50 values)., Conclusions: Suppression of fear conditioning to tone required approximately twice the isoflurane concentration that suppressed fear conditioning to context. Thus, the concentration of anesthetic required to suppress learning may depend on the neural substrates of learning. Our results suggest that isoflurane concentrations greater than 0.5 MAC may be needed to suppress both forms of fear conditioning.

The acquisition of context fear in rats is affected by variables such as the sex of the animal, the placement to shock interval (PSI), and preexposure to the context. The current experiments assessed the effects of these variables on context conditioning in mice (C57BL/6). In Experiment 1, mice were placed in a chamber and received a single shock 5s, 20 s, 40s, 60s, 180s, or 720s later. Increasing the PSI produced corresponding increases in conditional freezing during the context test. In addition, male mice acquired more context conditioning than female mice did but only at intermediate PSIs. In Experiment 2, preexposure to the context before training alleviated the sex difference found with an intermediate PSI. The results are discussed in terms of configural learning theory and are argued to be contrary to the predictions of scalar expectancy theory.

Dorsal hippocampal (DH) lesions produce a severe deficit in recently, but not remotely, acquired contextual fear without impairing memory of discrete training stimuli, i.e., DH lesions produce an anterograde and time-limited retrograde amnesia specific to contextual memory. These data are consistent with the standard model which posits temporary involvement of the hippocampus in recent memory maintenance. However, three recent controversies apparently weaken the case for a selective mnemonic role for the hippocampus in contextual fear. First, although retrograde amnesia (from posttraining lesions) is severe, anterograde amnesia (from pretraining lesions) may be mild or nonexistent. Second, a performance, rather than mnemonic, account of contextual freezing deficits in hippocampal-lesioned animals has been offered. Third, damage to the entire hippocampus, including the ventral hippocampus, can produce a dramatic and temporally stable disruption of context and tone fear. These data are reviewed and explanations are offered as to why they do not necessarily challenge the standard model of hippocampal memory function in contextual fear. Finally, a more complete description of the hippocampus' proposed role in contextual fear is offered, along with new data supporting this view. In summary, the data support a specific mnemonic role for the DH in the acquisition and consolidation of contextual representations.

Cholinergic neurotransmission has been implicated in the acquisition of a variety of tasks, including Pavlovian fear conditioning. To more precisely define the role of cholinergic modulation in this process, the effect of site-specific cholinergic antagonism was assessed. Male Long-Evans rats were implanted with chronic, bilateral cannulae aimed at the dorsal hippocampus. Infusions of scopolamine hydrobromide (50 microg bilaterally) or phosphate-buffered saline (PBS) were made immediately prior to a signaled Pavlovian fear conditioning procedure. On consecutive days following training, all rats were given independent tests assessing freezing to both the training context and the tone conditional stimulus (CS). Relative to PBS infused controls, rats that received intrahippocampal infusions of scopolamine showed a significant attenuation of contextual freezing but comparable levels of freezing to the tone CS. Neither shock sensitivity nor general activity levels differed between rats infused with scopolamine or PBS. These findings suggest that fear conditioning to context, but not discrete CS, requires intact cholinergic neurotransmission in the hippocampus.

On six days rats were exposed to each of two contexts. They received an electric shock in one context and nothing in the other. Rats were tested later in each environment without shock. The rats froze and defecated more often in the shock-paired environment; they also exhibited a significantly larger elevation in rectal temperature in that environment. The rats discriminated between each context, and we suggest that the elevation in temperature is the consequence of associative learning. Thus, body temperature can be used as a conditional response measure in Pavlovian fear conditioning experiments that use footshock as the unconditional stimulus.

This review examines the relationship between exploration and contextual fear conditioning. The fear acquired to places or contexts associated with aversive events is a form of Pavlovian conditioning. However, an initial period of exploration is necessary to allow the animal to form an integrated memory of the features of the context before conditioning can take place. The hippocampal formation plays a critical role in this process. Cells within the dorsal hippocampus are involved in the formation, storage and consolidation of this integrated representation of context. Projections from the subiculum to the nucleus accumbens regulate the exploration necessary for the acquisition of information about the features of the context. This model explains why electrolytic but not excitotoxic lesions of the dorsal hippocampus cause enhanced exploratory activity but both cause deficits in contextual fear. It also explains why retrograde amnesia of contextual fear is greater than anterograde amnesia.

The Sanfilippo syndrome type B is an autosomal recessive disorder caused by mutation in the gene (NAGLU) encoding alpha-N-acetylglucosaminidase, a lysosomal enzyme required for the stepwise degradation of heparan sulfate. The most serious manifestations are profound mental retardation, intractable behavior problems, and death in the second decade. To generate a model for studies of pathophysiology and of potential therapy, we disrupted exon 6 of Naglu, the homologous mouse gene. Naglu-/- mice were healthy and fertile while young and could survive for 8-12 mo. They were totally deficient in alpha-N-acetylglucosaminidase and had massive accumulation of heparan sulfate in liver and kidney as well as secondary changes in activity of several other lysosomal enzymes in liver and brain and elevation of gangliosides G(M2) and G(M3) in brain. Vacuolation was seen in many cells, including macrophages, epithelial cells, and neurons, and became more prominent with age. Although most vacuoles contained finely granular material characteristic of glycosaminoglycan accumulation, large pleiomorphic inclusions were seen in some neurons and pericytes in the brain. Abnormal hypoactive behavior was manifested by 4.5-mo-old Naglu-/- mice in an open field test; the hyperactivity that is characteristic of affected children was not observed even in younger mice. In a Pavlovian fear conditioning test, the 4.5-mo-old mutant mice showed normal response to context, indicating intact hippocampal-dependent learning, but reduced response to a conditioning tone, perhaps attributable to hearing impairment. The phenotype of the alpha-N-acetylglucosaminidase-deficient mice is sufficiently similar to that of patients with the Sanfilippo syndrome type B to make these mice a good model for study of pathophysiology and for development of therapy.

Muscarinic-cholinergic antagonism produces learning and memory deficits in a wide variety of hippocampal-dependent tasks. Hippocampal lesions produce both acquisition deficits and retrograde amnesia of contextual fear (fear of the place of conditioning), but do not impact fear conditioning to discrete cues (such as a tone). In order to examine the effects of muscarinic antagonism in this paradigm, rats were given 0.01 to 100 mg/kg of scopolamine (or methylscopolamine) either before or after a fear conditioning session in which tones were paired with aversive footshocks. Fear to the context and the tone were assessed by measuring freezing in separate tests. It was found that pretraining, but not post-training, scopolamine severely impaired fear conditioning; methylscopolamine was ineffective in disrupting conditioning. Although contextual fear conditioning was more sensitive to cholinergic disruption, high doses of scopolamine also disrupted tone conditioning. Scopolamine did not affect footshock reactivity, but did produce high levels of activity. However, hyperactivity was not directly responsible for deficits in conditioning. It was concluded that scopolamine disrupts CS-US association formation or CS processing, perhaps through an attenuation of hippocampal theta rhythm.

gamma-Aminobutyric acid (GABA) type A receptors mediate fast inhibitory synaptic transmission and have been implicated in responses to sedative/hypnotic agents (including neuroactive steroids), anxiety, and learning and memory. Using gene targeting technology, we generated a strain of mice deficient in the delta subunit of the GABA type A receptors. In vivo testing of various behavioral responses revealed a strikingly selective attenuation of responses to neuroactive steroids, but not to other modulatory drugs. Electrophysiological recordings from hippocampal slices revealed a significantly faster miniature inhibitory postsynaptic current decay time in null mice, with no change in miniature inhibitory postsynaptic current amplitude or frequency. Learning and memory assessed with fear conditioning were normal. These results begin to illuminate the novel contributions of the delta subunit to GABA pharmacology and sedative/hypnotic responses and behavior and provide insights into the physiology of neurosteroids.

After a few pairings of a threatening stimulus with a formerly neutral cue, animals and humans will experience a state of conditioned fear when only the cue is present. Conditioned fear provides a critical survival-related function in the face of threat by activating a range of protective behaviors. The present review summarizes and compares the results of different laboratories investigating the neuroanatomical and neurochemical basis of conditioned fear, focusing primarily on the behavioral models of freezing and fear-potentiated startle in rats. On the basis of these studies, we describe the pathways mediating and modulating fear. We identify several key unanswered questions and discuss possible implications for the understanding of human anxiety disorders.

We have shown previously that electrolytic lesions of the dorsal hippocampus (DH) produce a severe deficit in contextual fear if made 1 d, but not 28 d, after fear conditioning (). As such, the hippocampus seems to play a time-limited role in the consolidation of contextual fear conditioning. Here, we examine retrograde amnesia of contextual fear produced by DH lesions in a within-subjects design. Unlike our previous reports, rats had both a remote and recent memory at the time of the lesion. Rats were given 10 tone-shock pairings in one context (remote memory) and 10 tone-shock pairings in a distinct context (with a different tone) 50 d later (recent memory), followed by DH or sham lesions 1 d later. Relative to controls, DH-lesioned rats exhibited no deficit in remote contextual fear, but recent contextual fear memory was severely impaired. They also did not exhibit deficits in tone freezing. This highly specific deficit in recent contextual memory demonstrated in a within-subjects design favors mnemonic over performance accounts of hippocampal involvement in fear. These findings also provide further support for a time-limited role of the hippocampus in memory storage.

Angelman syndrome (AS) is a severe neurodevelopmental disorder resulting from a deletion/mutation in maternal chromosome 15q11-13. The genes in 15q11-13 contributing to the full array of the clinical phenotype are not fully identified. This study examines whether a loss or reduction in the GABAA receptor beta3 subunit (GABRB3) gene, contained within the AS deletion region, may contribute to the overall severity of AS. Disrupting the gabrb3 gene in mice produces electroencephalographic abnormalities, seizures, and behavior that parallel those seen in AS. The seizures that are observed in these mice showed a pharmacological response profile to antiepileptic medications similar to that observed in AS. Additionally, these mice exhibited learning and memory deficits, poor motor skills on a repetitive task, hyperactivity, and a disturbed rest-activity cycle, features all common to AS. The loss of the single gene, gabrb3, in these mice is sufficient to cause phenotypic traits that have marked similarities to the clinical features of AS, indicating that impaired expression of the GABRB3 gene in humans probably contributes to the overall phenotype of Angelman syndrome. At least one other gene, the E6-associated protein ubiquitin-protein ligase (UBE3A) gene, has been implicated in AS, so the relative contribution of the GABRB3 gene alone or in combination with other genes remains to be established.

Central neurokinin-1 (NK-1) receptors are thought to modulate aversion, whereas the periaqueductal gray matter (PAG) is a common pathway for the integration of fear behaviors. The authors determined whether injection of an NK-1 agonist (GR73632) into subregions of the PAG would alter fear-related behaviors. Behavioral inactivity was increased by GR73632 injected into the caudodorsal PAG or the dorsal raphe. Flight behavior induced by stimulation of the dorsal PAG or by a footshock was decreased after injection of GR73632 into the dorsal PAG. Rats that had 6 pairings of a tone with a footshock after injection of GR73632 into the dorsal PAG displayed more freezing behavior than controls at the beginning of the session. However, there was no change in the shock- or the tone-induced freezing because some GR73632-treated rats, but no controls, froze during the baseline period. It is concluded that NK-1 receptors in the dorsal PAG modulate the unconditional but not the mnemonic aspects of fear behaviors.