Learning & memory (Cold Spring Harbor, N.Y.)

Published by Cold Spring Harbor Laboratory Press
Online ISSN: 1549-5485
Print ISSN: 1072-0502
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LTP induced by a short tetanus at hippocampal CA1 synapses of wild-type and mutant mice lacking the IP3R1. (A) Typical examples of waveforms and the time-course of S-EPSP. A short tetanus (10 pulses, 100 Hz) induced LTP in a slice from a mutant mouse (closed circles), but failed to do so in a slice from a wild-type mouse (open circles). (B) Summarized results for the time-course of LTP in the S-EPSP in wild-type mice (n = 6, open circles) or mutant mice (n = 6, closed circles). (C) Typical example showing that the effect of a short tetanus in a mutant mouse was blocked by AP5 (10 µM, horizontal bar; closed diamonds). The insert shows sample traces of the field EPSP at the times indicated. (D) Summarized results for the time-course of LTP in the S-EPSP in mutant mice in the presence of AP5 (n = 6). In this and the following figures, the plots show the mean ± SEM. **, significant (P < 0.01) difference at 60 min after tetanus between the slices from wild-type and mutant mice.  
DP in slices from wild-type and mutant mice. (A) Typical example of the time-course for S-EPSP in a slice from a wild-type (open circles) and a mutant (closed circles) mouse. A train of low-frequency stimuli (LFS, horizontal bar) given 60 min after the standard tetanus (arrow) reduced LTP close to the control level in a wild-type mouse but not in a mutant mouse. (B) Summarized time-course data for DP in S-EPSP for wild-type (n = 6, open circles) and mutant (n = 7, closed circles) mice. *, significant (P < 0.05) difference in the S-EPSP at 60 min after LFS between slices from wild-type and mutant mice. (C) Typical example of time-course of S-EPSP showing that the standard LFS applied to a slice from a wild-type mouse in the presence of AP5 (50 µM, horizontal bar) failed to induce DP (open diamonds). (D) Summarized time-course data for S-EPSP in wild-type mice (n = 4, open diamonds) when the LFS (upper horizontal bar) was delivered in the presence of 50 µM AP5 (lower horizontal bar). DP was blocked by AP5 applied during the standard LFS.  
LTD induced by a short LFS in slices from wild-type and mutant mice. (A) Typical example of the time-course of S-EPSP. LTD was induced by a short LFS train (200 pulses at 1 Hz) in slices from both a wild-type (open circles) and a mutant (closed circles) mouse. (B) Summarized time-course data for LTD induced in wild-type (n = 5, open circles) and mutant (n = 5, closed circles) mice. (C) Typical example of the time-course of S-EPSP showing that a short LFS (upper horizontal bar) given in the presence of 50 µM AP5 (lower horizontal bar) failed to induce LTD in a slice from a wild-type (open circles) and a mutant (closed circles) mouse. (D) Summarized time-course data for S-EPSP from wild-type (n = 5, open circles) and mutant (n = 5, closed circles) mice when the LFS (upper horizontal bar) was applied in the presence of AP5 (lower horizontal bar).  
LTP suppression by low-frequency stimulation (LFS, horizontal bar) delivered 60 min before a standard tetanus. (A) Typical time-course of LTP suppression in a slice from a wild-type (open circles) and a mutant (closed circles) mouse. In the wild-type mouse, LFS induced long-term depression (LTD) in S-EPSP but suppressed subsequent LTP induction. In the mutant mouse, LFS given before tetanus-induced LTD failed to suppress LTP induction. (B) Summarized time-course data for LTD and LTP suppression in slices from wild-type (n = 6, open circles) and mutant (n = 6, closed circles) mice. *, significant (P < 0.05) difference in the SEPSP at 60 min after tetanus between slices from wild-type and mutant mice.
LTP induced by a standard tetanus in slices from wildtype and mutant mice. (A) Typical example of the time-course for S-EPSP. A stable LTP was induced by a standard tetanus (100 pulses at 100 Hz) in slices from both a wild-type (open circles) and a mutant (closed circles) mouse. (B) Summarized time-course data for LTP induced in wild-type (n = 5, open circles) and mutant (n = 5, closed circles) mice.  
In hippocampal CA1 neurons of wild-type mice, delivery of a standard tetanus (100 pulses at 100 Hz) or a train of low-frequency stimuli (LFS; 1000 pulses at 1 Hz) to a naive input pathway induces, respectively, long-term potentiation (LTP) or long-term depression (LTD) of responses, and delivery of LFS 60 min after tetanus results in reversal of LTP (depotentiation, DP), while LFS applied 60 min before tetanus suppresses LTP induction (LTP suppression). To evaluate the role of the type 1 inositol-1,4,5-trisphosphate receptor (IP3R1) in hippocampal synaptic plasticity, we studied LTP, LTD, DP, and LTP suppression of the field excitatory postsynaptic potentials (EPSPs) in the CA1 neurons of mice lacking the IP3R1. No differences were seen between mutant and wild-type mice in terms of the mean magnitude of the LTP or LTD induced by a standard tetanus or LFS. However, the mean magnitude of the LTP induced by a short tetanus (10 pulses at 100 Hz) was significantly greater in mutant mice than in wild-type mice. In addition, DP or LTP suppression was attenuated in the mutant mice, the mean magnitude of the responses after delivery of LFS or tetanus being significantly greater than in wild-type mice. These results suggest that, in hippocampal CA1 neurons, the IP3R1 is involved in LTP, DP, and LTP suppression but is not essential for LTD. The facilitation of LTP induction and attenuation of DP and LTP suppression seen in mice lacking the IP3R1 indicates that this receptor plays an important role in blocking synaptic potentiation in hippocampal CA1 neurons.
 
To define the physiological role of IP(3)3-kinase(A) in vivo, we have generated a mouse strain with a null mutation of the IP(3)3-kinase(A) locus by gene targeting. Homozygous mutant mice were fully viable, fertile, apparently normal, and did not show any morphological anomaly in brain sections. In the mutant brain, the IP4 level was significantly decreased whereas the IP3 level did not change, demonstrating a major role of IP(3)3-kinase(A) in the generation of IP4. Nevertheless, no significant difference was detected in the hippocampal neuronal cells of the wild-type and the mutant mice in the kinetics of Ca2+ regulation after glutamate stimulation. Electrophysiological analyses carried out in hippocampal slices showed that the mutation significantly enhanced the LTP in the hippocampal CA1 region, but had no effect on the LTP in dentate gyrus (DG). No difference was noted, however, between the mutant and the wild-type mice in the Morris water maze task. Our results indicate that IP(3)3-kinase(A) may play an important role in the regulation of LTP in hippocampal CA1 region through the generation of IP4, but the enhanced LTP in the hippocampal CA1 does not affect spatial learning and memory.
 
A-type K+ channels are known to regulate neuronal firing, but their role in repetitive firing and learning in mammals is not well characterized. To determine the contribution of the auxiliary K+ channel subunit Kvbeta1.1 to A-type K+ currents and to study the physiological role of A-type K+ channels in repetitive firing and learning, we deleted the Kvbeta1.1 gene in mice. The loss of Kvbeta1.1 resulted in a reduced K+ current inactivation in hippocampal CA1 pyramidal neurons. Furthermore, in the mutant neurons, frequency-dependent spike broadening and the slow afterhyperpolarization (sAHP) were reduced. This suggests that Kvbeta1.1-dependent A-type K+ channels contribute to frequency-dependent spike broadening and may regulate the sAHP by controlling Ca2+ influx during action potentials. The Kvbeta1.1-deficient mice showed normal synaptic plasticity but were impaired in the learning of a water maze test and in the social transmission of food preference task, indicating that the Kvbeta1.1 subunit contributes to certain types of learning and memory.
 
Generation and characterization of Ca V 1.2 cKO mice. (A) Exon 2 of the Ca V 1.2 wild-type gene (Cacna1c) is flanked by 2 XbaI restriction sites that were eliminated in the targeting vector by the insertion of a loxP site (arrowhead) in the 5-flanking region of exon 2 and a neomycin cassette (Neo) that was flanked on both ends by loxP sites (arrowheads) introduced into the 3 adjacent intron (intron 2). The targeting construct also carried the MC1 thymidine kinase (MC1-TK) cassette, which was used for negative selection with ganciclovir. Successful homologous recombination resulted in a floxed allele in which both exon 2 and the neomycin cassette would be flanked by loxP sites and therefore eliminated when exposed to Cre-recombinase. (B) Verification of conditional deletion of Ca V 1.2 exon 2 by RT-PCR. RNA samples from Ca V 1.2 cKO mice and littermate controls were isolated from the cerebellum (Cb), cortex (Ctx; Left and Right hemispheres), and hippocampus (Hc). RNA was then reverse-transcribed, and the resulting cDNA was amplified by PCR (using primers P1 and P2 in A) and separated by electrophoresis on an agarose gel. Deletion of exon 2 in the Ca V 1.2 cKO mice (lower band; -Ex 2) was detected in the cortex and hippocampus, whereas no such deletion was seen in control littermates (upper band; +Ex 2). (C) Western blot analysis of membrane fractions isolated from hippocampus, cortex, and cerebellum using an anti-Ca V 1.2 antibody reveals dramatically decreased detection of Ca V 1.2 in the hippocampus and cortex of the Ca V 1.2 cKO mice with no detectable decrease in the cerebellum. Blots were also probed with anti-NrCAM to control for protein loading.  
Ca V 1.2 cKO mice exhibit disrupted remote spatial memories. (A) Mice were trained for two trials a day for 14 d. The average time to reach the hidden platform (latency in seconds) is plotted for each training day. The latency to reach the platform in both groups decreased as training progressed, reaching an average escape latency of 16.0 3.6 sec for the Ca V 1.2 cKO mice and 17.5 3.2 sec for the controls. There was a main effect of training day on the latency to find the hidden platform (F (13,325) = 11.7; p < 0.0001); however, there was no main effect of genotype on latency (F (1,25) = 0.24; p > 0.05) and no interaction between training day and genotype (F (13,325) = 0.86; p > 0.05). (B) A 60-sec probe trial completed before the start of training on day 13 reveals that both Ca V 1.2 cKO and control littermate mice spend a significant percentage of time during the trial searching in the quadrant where the platform was previously located (TQ; training quadrant) with the Ca V 1.2 cKO mice spending 57% 3.7% time in the TQ and control mice searching 57.6% 8.7% time in the TQ. However, there was no significant difference in the amount of time that the Ca V 1.2 cKO mice spent in the TQ compared to their control littermates (F (1,24) = 0.20; p > 0.05; One-Way ANOVA). The dashed line (25%) represents random or " chance " performance; (AR) adjacent right; (AL) adjacent left; (OP) opposite. (C) The percent time that the Ca V 1.2 cKO mice spent searching in the training quadrant (TQ) during each of the four probes completed during training (on days 4, 7, 10, and 13) was not significantly different when compared to control littermates. However, when tested 30 d later (remote memory probe), the Ca V 1.2 cKO mice exhibit a degraded search strategy when compare to their littermate controls. In contrast to the performance of the control mice during the remote memory probe (56.8% 2.7% time in TQ), which remained similar to that observed during the last memory probe administered during training, the Ca V 1.2 cKO mice spent significantly less time (41.7% 4.2%) in the TQ when compared to control littermates (F (1,23) = 9.4; P < 0.01; One-Way ANOVA). All data are presented as mean SEM.  
To determine whether L-type voltage-gated calcium channels (L-VGCCs) are required for remote memory consolidation, we generated conditional knockout mice in which the L-VGCC isoform Ca(V)1.2 was postnatally deleted in the hippocampus and cortex. In the Morris water maze, both Ca(V)1.2 conditional knockout mice (Ca(V)1.2(cKO)) and control littermates displayed a marked decrease in escape latencies and performed equally well on probe trials administered during training. In distinct contrast to their performance during training, Ca(V)1.2(cKO) mice exhibited significant impairments in spatial memory when examined 30 d after training, suggesting that Ca(V)1.2 plays a critical role in consolidation of remote spatial memories.
 
Percentage change in cholinesterase levels in red blood cells compared with baseline pretreatment levels for aged rabbits. Aged galantamine-administered (Aged/Gal) rabbits exhibited significantly reduced cholinesterase levels compared with aged saline-treated control (Aged/Veh) rabbits. Time points correspond to the first, tenth, and twen- tieth days of training. Aged/Gal and Aged/Veh groups include trace conditioned and pseudoconditioned rabbits. 
Mean conditioned response peak latency for each day of trace conditioning. Aged galantamine-administered (Aged/Gal) rabbits exhibited a significant change in peak latency timing across days, as well as significantly shorter mean peak latency time compared with aged saline control (Aged/Veh) rabbits. Aged/Gal and young rabbits did not differ significantly on the final day of training. No significant difference was observed between young galantamine-administered (Young/Gal) and young saline control (Young/Veh) rabbits. 
Mean conditioned response amplitude for each day of trace conditioning. No significant differences were observed within or between age groups over the course of training (Aged/Gal indicates aged galantamine-administered rabbits; Aged/Veh, aged saline control rabbits; Young/Gal, young galantamine-administered rabbits; and Young/Veh, young saline control rabbits). 
( A ) Mean tone-CS-elicited eyeblink responses by day during pseudoconditioning of aged rabbits. No significant differences were observed between aged galantamine-administered (Aged/Gal) and aged saline control (Aged/Veh) rabbits. ( B ) Mean tone-CS-elicited eyeblink responses by day during pseudoconditioning of young rabbits. No significant differences were observed between young galantamine- administered (Young/Gal) and young saline control (Young/Veh) rabbits. 
Cholinergic systems are critical to the neural mechanisms mediating learning. Reduced nicotinic cholinergic receptor (nAChR) binding is a hallmark of normal aging. These reductions are markedly more severe in some dementias, such as Alzheimer's disease. Pharmacological central nervous system therapies are a means to ameliorate the cognitive deficits associated with normal aging and aging-related dementias. Trace eyeblink conditioning (EBC), a hippocampus- and forebrain-dependent learning paradigm, is impaired in both aged rabbits and aged humans, attributable in part to cholinergic dysfunction. In the present study, we examined the effects of galantamine (3 mg/kg), a cholinesterase inhibitor and nAChR allosteric potentiating ligand, on the acquisition of trace EBC in aged (30-33 months) and young (2-3 months) female rabbits. Trace EBC involves the association of a conditioned stimulus (CS) with an unconditioned stimulus (US), separated by a stimulus-free trace interval. Repeated CS-US pairings results in the development of the conditioned eyeblink response (CR) prior to US onset. Aged rabbits receiving daily injections of galantamine (Aged/Gal) exhibited significant improvements compared with age-matched controls in trials to eight CRs in 10 trial block criterion (P = 0.0402) as well as performance across 20 d of training [F(1,21) = 5.114, P = 0.0345]. Mean onset and peak latency of CRs exhibited by Aged/Gal rabbits also differed significantly [F(1,21) = 6.120/6.582, P = 0.0220/0.0180, respectively] compared with age-matched controls, resembling more closely CR timing of young drug and control rabbits. Galantamine did not improve acquisition rates in young rabbits compared with age-matched controls. These data indicate that by enhancing nicotinic and muscarinic transmission, galantamine is effective in offsetting the learning deficits associated with decreased cholinergic transmission in the aging brain.
 
Representation of the maximal (outlined region) and minimal (filled region) basal amygdala lesions. The numbers in bold indicate the distance posterior to bregma.
Basal amygdala lesions do not interfere with the extinction of tone-elicited fear. Percent freezing to the tone is shown in blocks of two trials (except for first trial of conditioning and last trial of extinction in Day 3). Basal lesion (filled circles; n = 7) and Sham (open circles; n = 8). Freezing was assessed initially using an automated system (see Materials and Methods section), and then confirmed by observing the rat’s behavior stored on videotape. 
Fear extinction refers to the ability to adapt as situations change by learning to suppress a previously learned fear. This process involves a gradual reduction in the capacity of a fear-conditioned stimulus to elicit fear by presenting the conditioned stimulus repeatedly on its own. Fear extinction is context-dependent and is generally considered to involve the establishment of inhibitory control of the prefrontal cortex over amygdala-based fear processes. In this paper, we review research progress on the neural basis of fear extinction with a focus on the role of the amygdala and the prefrontal cortex. We evaluate two competing hypotheses for how the medial prefrontal cortex inhibits amygdala output. In addition, we present new findings showing that lesions of the basal amygdala do not affect fear extinction. Based on this result, we propose an updated model for integrating hippocampal-based contextual information with prefrontal-amygdala circuitry.
 
The present study investigated reinstatement of conditioned responses in humans by using a differential Pavlovian conditioning procedure. Evidence for reinstatement was established in a direct (fear rating) and in an indirect measure (secondary reaction time task) of conditioning. Moreover, the amount of reinstatement in the secondary reaction time task was significantly correlated with the difference in valence between the conditioned stimulus (CS)+ and the CS-after extinction. These data provide clear evidence for reinstatement and for the role of negative stimulus valence in the return of conditioned responding after extinction.
 
It is well established that the cerebellum and its associated circuitry are essential for classical conditioning of the eyeblink response and other discrete motor responses (e.g., limb flexion, head turn, etc.) learned with an aversive unconditioned stimulus (US). However, brain mechanisms underlying extinction of these responses are still relatively unclear. Behavioral studies have demonstrated extinction as an active learning process distinct from acquisition. Experimental data in eyeblink conditioning suggest that plastic changes specific to extinction may play an important role in this process. Both cerebellar and hippocampal systems may be involved in extinction of these memories. The nature of this phenomenon and identification of the neural substrates necessary for extinction of originally learned responses is the topic of this review.
 
The extent of damage caused by NMDA lesions. SHAM, sham operated controls; ADMS, anterior dorsomedial striatum; PDMS, posterior dorsomedial striatum; DLS, dorsolateral striatum. From top to bottom the distances from bregma in mm are: DLS and ADMS, 1.6, 1, 0.48, מ 0.26; PDMS, 0.2, 
Performance across 14 d of acquisition training. Abbreviations as in Figure 1.
The results from both probe tests. Abbreviations as in Figure 1. The numbers in the parentheses show how many rats from each lesion group made that particular choice (place or response).
The involvement of different subregions of the striatum in place and response learning was examined using a T-maze. Rats were given NMDA lesions of the dorsolateral striatum (DLS), anterior dorsomedial striatum (ADMS), posterior dorsomedial striatum (PDMS), or sham surgery. They were then trained to retrieve food from the west arm of the maze, starting from the south arm, by turning left at the choice point. After 7 d of training, with four trials a day, a probe test was given in which the starting arm is inserted as the north arm, at the opposite side of the maze. A left turn would indicate a "response" strategy; a right turn, a "place" strategy. The rats were then trained for 7 more days, followed by a second probe test. Unlike rats in the other groups, most of the rats in the PDMS group turned left, using the response strategy on both probe tests. These results suggest that the PDMS plays a role in spatially guided behavior.
 
(Experiment 1) Tg2576 mice do not differ from wild-type mice in the number of arm entries but exhibit a lower rate of spontaneous alternation. (A) Mean number SEM of arm entries and (B) mean percentage SEM of spontaneous alternation rate, during a 5-min session of exploration in the Y maze. (tg) Tg2576 mice; (wt) wild-type mice; (*) P < 0.05. 
(Experiment 2) Tg2576 mice show superior cross maze training performance. Mean number SEM of correct runs made during six daily four-run trials (top) and mean time SEM to complete each trial (bottom). (tg) Tg2576 mice; (wt) wild-type mice. 
Mutations in the amyloid precursor protein (APP) gene inducing abnormal processing and deposition of beta-amyloid protein in the brain have been implicated in the pathogenesis of Alzheimer's disease (AD). Although Tg2576 mice with the Swedish mutation (hAPPswe) exhibit age-related Abeta-plaque formation in brain regions like the hippocampus, the amygdala, and the cortex, these mice show a rather specific deficit in hippocampal-dependent learning and memory tasks. In view of recent findings showing that neural systems subserving different forms of learning are not simply independent but that depressing or enhancing one system affects learning in another system, we decided to investigate fronto-striatal synaptic plasticity and related procedural learning in these mutants. Fronto-striatal long-term depression (LTD) induced by tetanic stimulation of the cortico-striatal input was similar in Tg2576 and wild-type control mice. Behavioral data, however, pointed to an enhancement of procedural learning in the mutants that showed robust motor-based learning in the cross maze and higher active avoidance scores. Thus, in this mouse model of AD, an intact striatal function associated with an impaired hippocampal function seems to provide neural conditions favorable to procedural learning. Our results suggest that focusing on preserved or enhanced forms of learning in AD patients might be of interest to describe the functional reorganization of the brain when one memory system is selectively compromised by neurological disease.
 
Classical conditioning of the eyeblink reflex to a neutral stimulus that predicts an aversive stimulus is a basic form of associative learning. Acquisition and retention of this learned response require the cerebellum and associated sensory and motor pathways and engage several other brain regions including the hippocampus, neocortex, neostriatum, septum, and amygdala. The cerebellum and its associated circuitry form the essential neural system for delay eyeblink conditioning. Trace eyeblink conditioning, a learning paradigm in which the conditioned and unconditioned stimuli are noncontiguous, requires both the cerebellum and the hippocampus and exhibits striking parallels to declarative memory formation in humans. Identification of the neural structures critical to the development and maintenance of the conditioned eyeblink response is an essential precursor to the investigation of the mechanisms responsible for the formation of these associative memories. In this review, we describe the evidence used to identify the neural substrates of classical eyeblink conditioning and potential mechanisms of memory formation in critical regions of the hippocampus and cerebellum. Addressing a central goal of behavioral neuroscience, exploitation of this simple yet robust model of learning and memory has yielded one of the most comprehensive descriptions to date of the physical basis of a learned behavior in mammals.
 
This article provides a selective review and integration of the behavioral literature on Pavlovian extinction. The first part reviews evidence that extinction does not destroy the original learning, but instead generates new learning that is especially context-dependent. The second part examines insights provided by research on several related behavioral phenomena (the interference paradigms, conditioned inhibition, and inhibition despite reinforcement). The final part examines four potential causes of extinction: the discrimination of a new reinforcement rate, generalization decrement, response inhibition, and violation of a reinforcer expectation. The data are consistent with behavioral models that emphasize the role of generalization decrement and expectation violation, but would be more so if those models were expanded to better accommodate the finding that extinction involves a context-modulated form of inhibitory learning.
 
Yohimbine facilitates long-term cue fear extinction with massed CS exposures. Separate experiments examined the effects of vehicle, yohimbine, and propranolol on extinction with 0 (A), 5 (B), 10 (C), 20 (D), or 30 (E) nonreinforced CS presentations (seven to eight mice/group). The data shown represent freezing during a single 2-min test CS given 1 d after the extinction sessions (drug free). The final summary panel (F) shows the results of the five experiments with fear normalized to freezing in groups of mice that were injected with vehicle and not presented with any extinction CSs on Day 2 (retention control mice, RC). (*) p < 0.05 versus RC mice; (+) p < 0.05 versus vehicleextinction mice.  
Yohimbine facilitates long-term context fear extinction. Separate experiments examined the effects of vehicle, yohimbine, and propranolol on extinction with 0 (A), 20 (B), 40 (C), or 60 (D) min of context exposure (eight to 12 mice/group). The data shown represent freezing during a single 5-min context exposure given 1 d after the extinction sessions (drug free). The final summary panel (E) shows the results of the four experiments with fear normalized to freezing in groups of mice that were injected with vehicle and not exposed to the feared context on Day 2 (retention control mice, RC). (*) p < 0.05 versus RC mice; (+) p < 0.05 versus vehicle-extinction mice.  
Yohimbine facilitates extinction of cue and context fear acutely. CS-elicited freezing during the Day 2 extinction sessions from all of the experiments shown in Figures 1 and 2, 20 min after injection of vehicle, yohimbine, or propranolol (27–32 mice/group). Because mice were treated identically up to the fifth CS of the cue fear experiments (A) and the tenth minute of exposure in the context fear experiments (B), the data were combined for this analysis.  
Yohimbine has no effect on cue fear extinction when injected immediately after massed CS presentations. Separate experiments examined the effect of injecting vehicle, yohimbine, or propranolol after 10 (A) or 20 (B) massed CS presentations (eight mice/ group). The data shown represent freezing during a single 2-min CS given 1 d after the extinction sessions (drug free). Retention control (RC) mice received no CS presentations and were injected with vehicle on Day 2.  
Yohimbine leads to extinction, whereas propranolol leads to incubation, of cue fear when given in conjunction with spaced CS presentations. Separate experiments examined the effect of vehicle, yohimbine, or propranolol on the effect of spaced CS presentations after weak (A,B; see Materials and Methods) or strong (C,D; see Materials and Methods) cue fear conditioning (eight mice/group). In each experiment, 1 d after acquisition, mice were injected with vehicle, yohimbine, or propranolol and presented with seven CSs (20 min ITI). (A,C) Freezing during the spaced CS sessions 20 min after injections. (B,D) Freezing 1 d later during the final 5-min CS tests (drug free). (*) p < 0.05 versus RC mice; (+) p < 0.05 versus vehicleexposure mice.  
Extinction of classically conditioned fear, like its acquisition, is active learning, but little is known about its molecular mechanisms. We recently reported that temporal massing of conditional stimulus (CS) presentations improves extinction memory acquisition, and suggested that temporal spacing was less effective because individual CS exposures trigger two opposing processes: (1) fear extinction, which is favored by CS massing, and (2) fear incubation (increase), which is favored by spacing. We here report the effects of manipulating the adrenergic system during massed or spaced CS presentations in fear-conditioned mice. We administered yohimbine (5 mg/kg), an alpha(2)-receptor antagonist, or propranolol (10 mg/kg), a beta-receptor antagonist, systemically prior to CS presentation sessions and recorded both short- and long-term changes in conditional freezing. Yohimbine treatment facilitated extinction of both cue and context fear with massed protocols. When given before spaced CS presentations, propranolol led to a persistent incubation of cue fear, whereas yohimbine led to persistent extinction, compared with vehicle-treated animals, which showed no change in fear. These results suggest that norepinephrine positively modulates the formation of fear extinction memories in mice. They also provide clear evidence that spaced CS presentations trigger both fear-reducing (extinction) and fear-increasing (incubation) mechanisms.
 
Modulation of Gene and Protein Expression by Extinction Training in Cocaine Withdrawal.
Cocaine produces multiple neuroadaptations with chronic repeated use. Many of these neuroadaptations can be reversed or normalized by extinction training during withdrawal from chronic cocaine self-administration in rats. This article reviews our past and present studies on extinction-induced modulation of the neuroadaptive response to chronic cocaine in the mesolimbic dopamine system, and the role of this modulation in addictive behavior in rats. Extinction training normalizes tyrosine hydroxylase levels in the nucleus accumbens (NAc) shell, an effect that could help ameliorate dysphoria and depression associated with withdrawal from chronic cocaine use. Extinction training also increases levels of GluR1 and GluR2/3 AMPA receptor subunits, while normalizing deficits in NR1 NMDA receptor subunits, in a manner consistent with long-term potentiation of excitatory synapses in the NAc shell. Our results suggest that extinction-induced increases in AMPA and NMDA receptors may restore deficits in cortico-accumbal neurotransmission in the NAc shell and facilitate inhibitory control over cocaine-seeking behavior. Other changes identified by gene expression profiling, including up-regulation in the AMPA receptor aggregating protein Narp, suggest that extinction training induces extensive synaptic reorganization. These studies highlight potential benefits for extinction training procedures in the treatment of drug addiction.
 
The endogenous cannabinoid system has been shown recently to play a crucial role in the extinction of aversive memories. As the amygdala is presumably involved in this process, we investigated the effects of the cannabinoid receptor agonist WIN 55,212-2 (WIN-2) on synaptic transmission in the lateral amygdala (LA) of wild-type and cannabinoid receptor type 1 (CB1)-deficient mice. Extracellular field potential recordings and patch-clamp experiments were performed in an in vitro slice preparation. We found that WIN-2 reduces basal synaptic transmission and pharmacologically isolated AMPA receptor- and GABA(A) receptor-mediated postsynaptic currents in wild-type, but not in CB1-deficient mice. These results indicate that, in the LA, cannabinoids modulate both excitatory and inhibitory synaptic transmission via CB1. WIN-2-induced changes of paired-pulse ratio and of spontaneous and miniature postsynaptic currents suggest a presynaptic site of action. Inhibition of G(i/o) proteins and blockade of voltage-dependent and G protein-gated inwardly rectifying K(+) channels inhibited WIN-2 action on basal synaptic transmission. In contrast, modulation of the adenylyl cyclase-protein kinase A pathway, and blockade of presynaptic N- and P/Q- or of postsynaptic L- and R/T-type voltage-gated Ca(2+) channels did not affect WIN-2 effects. Our results indicate that the mechanisms underlying cannabinoid action in the LA partly resemble those observed in the nucleus accumbens and differ from those described for the hippocampus.
 
Systemic administration of WAY-100635 (WAY) blocks LTP induction in awake freely moving animals exploring novel environments. (A) Point plot of averaged medial perforant pathdentate field EPSP slopes (SEM) showing baseline responses, responses 25-30 min after application of drug, and 1 h following tetanus. Systemic WAY attenuated LTP in doses of 0.5 () and 0.01 mg/kg (), (P < 0.05). The 0.005 mg/kg dose ( ) did not differ significantly from controls receiving only the water vehicle (), (n = 4/group). Traces are representative responses collected 20-25 min following administration of either (a) the water vehicle or (b) 0.5 mg/kg of WAY, and responses collected 1 h after delivery of trains. Calibration: 1 mV, 5 msec. (B) Comparison of the time course of LTP over a 2-wk period induced following administration of the water vehicle () or 0.005 (•), 0.01 () or 0.5 mg/kg of WAY (), (n = 4/group). WAY in doses of 0.01 and 0.5 mg/kg attenuated significantly the duration of LTP as compared with LTP induced in the presence of the vehicle (P < 0.05).
Systemically administered WAY blocks LTP in novel, but not familiar, environments. (A) Comparison of the effects of the 0.5-mg/kg dose of WAY on LTP induced while animals explored either a novel (), (n = 4) or a familiar (), (n = 5) cage ( SEM). WAY blocked LTP induced in the novel environment, but did not alter LTP induced in the familiar home cage. Traces are representative responses collected 20-25 min after drug administration, and 1 h after delivery of trains in either familiar (a) or novel (b) conditions. Calibration: 1 mV, 5 msec. (B) Comparison of the time course of LTP induced after administration of 0.5 mg/kg WAY in either the novel (), (n = 4) or the familiar home environment (), (n = 4). LTP was observed 24 h later only when induced in a familiar environment. (C) Measures of total activity over the 5-min period of exploration in a novel or a familiar cage 25-30 min following systemic administration of 0.5 mg/kg WAY (n = 4/group). Activity measures reflect the total number of pixels detecting contrast changes in the caged area (1080 3250 pixels). WAY had no effect on the total activity of animals as measured in a novel environment (P > 0.05).
Numerous studies suggest roles for monoamines in modulating long-term potentiation (LTP). Previously, we reported that both induction and maintenance of perforant path-dentate gyrus LTP is enhanced when induced while animals explore novel environments. Here we investigate the contribution of serotonin and 5-HT1a receptors to the novelty-mediated enhancement of LTP. In freely moving animals, systemic administration of the selective 5-HT1a antagonist WAY-100635 (WAY) attenuated LTP in a dose-dependent manner when LTP was induced while animals explored novel cages. In contrast, LTP was completely unaffected by WAY when induced in familiar environments. LTP was also blocked in anesthetized animals by direct application of WAY to the dentate gyrus, but not to the median raphe nucleus (MRN), suggesting the effect of systemic WAY is mediated by a block of dentate 5-HT1a receptors. Paradoxically, systemic administration of the 5-HT1a agonist 8-OH-DPAT also attenuated LTP. This attenuation was mimicked in anesthetized animals following application of 8-OH-DPAT to the MRN, but not the dentate gyrus. In addition, application of a 5-HT1a agonist to the dentate gyrus reduced somatic GABAergic inhibition. Because serotonergic projections from the MRN terminate on dentate inhibitory interneurons, these data suggest 5-HT1a receptors contribute to LTP induction via inhibition of GABAergic interneurons. Moreover, activation of raphe 5-HT1a autoreceptors, which inhibits serotonin release, attenuated LTP induction even in familiar environments. This suggests that serotonin normally contributes to dentate LTP induction in a variety of behavioral states. Together, these data suggest that serotonin and dentate 5-HT1a receptors play a permissive role in dentate LTP induction, particularly in novel conditions, and presumably, during the encoding of novel, hippocampus-relevant information.
 
Cannulae placements in the ACC. The schematics depict the location of the injection cannulae tips () in the ACC for rats in Experiment 1 (A) and Experiment 2 (B). Plates are adaptations used with permission from Elsevier © 1986, Paxinos and Watson 1986. Numbers beside each plate correspond to millimeters anterior to bregma. (C) Nissl stain of a coronal section indicating the cannulae tip placements.
Effects of an intra-ACC D1 receptor blockade on effort-based decision making in a T-maze cost-benefit task. Mean ( ע SEM) percentage of high-rewarded arm choices (HR) per day is given. Testing block A was preoperatively without treatment, blocks B and C post-operatively after pre-trial infusions of SCH23390 (1 μg in 0.5 μL per side; n = 10; ᭞ ) or saline (0.5 μL; n = 11; ᭹ ) into the ACC. Each block consisted of three consecutive test days; on blocks A and B, a 30-cm barrier was placed in the HR; on block C, identical 30-cm barriers were placed into each goal arm, respectively. 
Effects of an intra-ACC D2 receptor blockade on effort-based decision making in a T-maze cost-benefit task. Mean ( ע SEM) percentage of high-rewarded arm choices (HR) per day is given. Testing block A was preoperatively without treatment, blocks B and C postoperatively after pre-trial infusions of eticlopride (1 μg in 0.5 μL per side; n = 10; छ ) or saline (0.5 μL; n = 12; ᭹ ) into the ACC. Each block consisted of three consecutive test days; on blocks A and B, a 30-cm barrier was placed in the HR; on block C, identical 30-cm barriers were placed into each goal arm, respectively. 
The anterior cingulate cortex (ACC) has been implicated in encoding whether or not an action is worth performing in view of the expected benefit and the cost of performing the action. Dopamine input to the ACC may be critical for this form of effort-based decision making; however, the role of distinct ACC dopamine receptors is yet unknown. Therefore, we examined in rats the effects of an intra-ACC D1 and D2 receptor blockade on effort-based decision making tested in a T-maze cost-benefit task. In this task, subjects could either choose to climb a barrier to obtain a high reward in one arm or a low reward in the other arm without a barrier. Unlike vehicle-treated rats, rats with intra-ACC infusion of the D1 receptor antagonist SCH23390 exhibited a reduced preference for the high-cost- high-reward response option when having the choice to obtain a low reward with little effort. In contrast, in rats with intra-ACC infusion of the D2 receptor antagonist eticlopride, the preference for the high-cost-high-reward response option was not altered relative to vehicle-treated rats. These data provide the first evidence that D1 receptors in the ACC regulate effort-based decision making.
 
Activation of ERK regulates histone H3 phosphorylation, acetylation, and phospho-acetylation in vitro. ( A ) Quantification of immunoblot densities for phospho-ERK and total ERK. Treatment of hippocampal slices with PDA (3 μM, n = 9) or FSK (50 μM with 100 μM Ro20- 1724, n = 9) for 1 h significantly increased phosphorylation of ERK2 in area CA1. Pre-incubation of slices with U0126 (20 μM, n = 5) for 10 min followed by PDA or FSK blocked the increase in ERK2 phosphorylation. Total ERK protein was unchanged in all treatments. Representative immunoblots for P-ERK and total ERK are shown for each treatment condition. Control (C) samples appear on the left , and experimental (E) samples appear on the right . ( B ) Quantification of immunoblot densities for phospho-histone H3, acetyl-histone H3, and phospho-acetyl-histone H3. Treatment of hippocampal slices with PDA (3 μM, n = 7) or FSK (50 μM with 100 μM Ro20-1724, n = 6) for 1 h significantly increased phosphorylation, acetylation, and phospho-acetylation of histone H3 in area CA1. Pre-incubation of slices with U0126 (20 μM, n = 5) for 10 min followed by PDA or FSK blocked these changes. Total histone H3 protein was unchanged. Representative immunoblots for P-H3 and total H3 are shown for each treatment condition. Control (C) samples appear on the left , and experimental (E) samples appear on the right . All drug-treated slices were compared with vehicle-treated controls. Error bars indicate standard error of the mean. Asterisks denote significant differences ( P < 0.05) as determined by Tukey ’ s multiple comparison test. 
Contextual fear conditioning regulates histone H3 phosphorylation, acetylation, and phospho-acetylation in vivo. (A) Quantification of immunoblot densities for phospho-histone H3, acetyl-histone H3, and phospho-acetyl-histone H3 at different time points following contextual fear conditioning. Histone H3 phosphorylation, acetylation, and phospho-acetylation in area CA1 were significantly increased at 1 h after training (FC 1 h, n = 8) before returning to baseline and remaining even after 24 h. Total histone H3 was unchanged. Representative immunoblots for P-H3 and total H3 are shown for each time point. Control (C) samples appear on the left, and experimental (E) samples appear on the right. (B) Quantification of immunoblot densities for phospho-histone H3, acetyl-histone H3, and phospho-acetyl-histone H3. The latent inhibition paradigm (lat inh, n = 4) significantly reduced histone H3 phosphorylation, acetylation, and phospho-acetylation in area CA1 compared with fear conditioning (FC 1 h, n = 8). Injection of animals with MK801 (300 µg/kg) prior to fear conditioning (FC + MK801, n = 3) significantly reduced histone H3 phosphorylation, acetylation, and phospho-acetylation in area CA1 compared with injection with saline (0.9% NaCl, 1.25 mL/kg) prior to fear conditioning (FC + saline, n = 3). Total histone H3 was unchanged. Representative immunoblots for P-H3 and total H3 are shown for each condition. Control (C) samples appear on the left, and experimental (E) samples appear on the right. Error bars indicate standard error of the mean. Asterisks denote significant differences (P < 0.05) as determined by Tukey's multiple comparison test.
MEK signaling through ERK contributes to long-term fear memory and contextual fear conditioning-induced changes in histone H3. ( A ) Quantification of freezing behavior 24 h following the training period. Animals that were fear conditioned and injected with DMSO (2.99 mL/kg, n = 4) displayed significantly greater freezing than either naive animals (naive, n = 4) or animals that were fear-conditioned and injected with SL327 (100 mg/kg, n = 7). Naive animals had not been previously exposed to either the training chamber or shocks. ( B ) Quantification of immunoblot densities for phospho-ERK and total ERK. Injection of animals with SL327 (100 mg/kg) following fear conditioning (FC + SL327, n = 9) significantly reduced ERK2 phosphorylation in area CA1 compared with injection 
Long-term memory formation is regulated by many distinct molecular mechanisms that control gene expression. An emerging model for effecting a stable, coordinated pattern of gene transcription involves epigenetic tagging through modifications of histones or DNA. In this study, we investigated the regulation of histone phosphorylation in the hippocampus by the ERK/MAPK (extracellular signal-regulated kinase/mitogen-activated protein kinase) pathway. We found that activation of ERK/MAPK in vitro significantly increased histone H3 phosphorylation in hippocampal area CA1. Furthermore, we found that contextual fear conditioning in vivo leads to a rapid time-dependent increase in histone H3 phosphorylation in area CA1. This increase paralleled the time course of contextual fear-dependent activation of ERK, and was inhibited in vivo by a latent inhibition paradigm as well as by injection of an N-methyl-d-aspartic acid receptor (NMDA-R) antagonist. Finally, injection of an inhibitor of MEK (MAP kinase/ERK kinase), the unique dual-specificity kinase upstream of ERK, blocked the increase in histone H3 phosphorylation seen after contextual fear conditioning. These results demonstrate that changes in histone phosphorylation in the hippocampus are regulated by ERK/MAPK following a behavioral fear conditioning paradigm.
 
Consolidation refers to item stabilization in long-term memory. Retrieval renders a consolidated memory sensitive, and a "reconsolidation" process has been hypothesized to keep the original memory persistent. Some authors could not detect this phenomenon. Here we show that retrieved contextual fear memory is vulnerable to amnesic treatments and that the amygdala is critically involved. Cholinergic and histaminergic systems seem to modulate only consolidation, whereas cannabinoids are involved in both consolidation and reactivation. The lability of retrieved memory affords opportunities to treat disorders such as phobias, post-traumatic stress, or chronic pain, and these results help searching for appropriate therapeutic targets.
 
Effects of SNAP, 8-br-cGMP, forskolin, DB-cAMP, and IBMX on LTM formation. At 20 min prior to single-trial conditioning, animals in two groups were each injected with 3 µL of saline (a) or saline containing 0.1% DMSO (b), and animals in another five groups were each injected with 3 µL of saline containing SNAP (200 µM) (c), 8-br-cGMP (200 µM) (d), forskolin (200 µM) (e), DB-cAMP (200 µM) (f), or IBMX (200 µM) and 0.1% DMSO (g). The PIs were measured before conditioning (white bars) and 24 h after conditioning (shaded bars) and are shown as means SE., and the results of statistical comparison between them are indicated. (***) P < 0.001, (NS) P > 0.05, WCX test. The number of animals tested is shown at each data point. The odor preferences at 24 h after conditioning of the groups shown in c-g did not significantly differ among each other (P > 0.05, K-W test).
Effects of various drugs on induction of LTM by an NO donor, cGMP analog, forskolin, or cAMP analog paired with single-trial conditioning. At 20 min prior to single-trial conditioning, animals were injected with 3 µL of saline containing various drugs. In A, animals in six groups were each coinjected with SNAP (200 µM) and cycloheximide (CHX, 10 mM) (a), 8-br-cGMP (200 µM) and CHX (10 mM) (b), DB-cAMP (200 µM) and CHX (10 mM) (c), SNAP (200 µM) and KT5720 (200 µM) (d), 8-br-cGMP (200 µM) and KT5720 (200 µM) (e), or DB-cAMP (200 µM) and KT5720 (200 µM) (f). In B, animals in four groups were each coinjected with SNAP (200 µM) and L-NAME (400 µM) (a), SNAP (200 µM) and ODQ (200 µM) (b), 8-br-cGMP (200 µM) and L-NAME (400 µM) (c), or 8-br-cGMP (200 µM) and ODQ (200 µM) (d). In C, animals in six groups were each coinjected with 8-br-cGMP (200 µM) and DDA (1 mM) (a), forskolin (200 µM) and ODQ (200 µM) (b), forskolin (200 µM) and DDA (1 mM) (c), DB-cAMP (200 µM) and L-NAME (400 µM) (d), DBcAMP (200 µM) and ODQ (200 µM) (e), or DB-cAMP (200 µM) and DDA (1 mM) (f). The PIs measured before (white bars) and 24 h after conditioning (shaded bars) are shown as means SE. The results of statistical comparison before and 24 h after conditioning are shown. (***) P < 0.001, (NS) P > 0.05, WCX test. The number of animals tested is shown at each data point. The odor preferences at 24 h after conditioning of the groups shown in a, c, and d in B and b, d, e, and f in C did not significantly differ among each other (P > 0.5, K-W test).
Investigation of the biochemical pathway intervening the NO-cGMP pathway and adenylyl cyclase-cAMP-PKA pathway. In A, animals in nine groups were each coinjected with 3 µL of saline containing 8-br-cGMP (200 µM) and KT5823 (200 µM) (a), 8-br-cGMP (200 µM) and L-DIL (1 mM) (b), forskolin (200 µM) and L-DIL (1 mM) (c), DB-cAMP (200 µM) and L-DIL (1 mM) (d), 8-br-cGMP (200 µM) and DCB (1 mM) (e), forskolin (200 µM) and DCB (1 mM) (f), 8-br-cGMP (200 µM) and W-7 (200 µM) (g), forskolin (200 µM) and W-7 (200 µM) (h), or DB-cAMP (200 µM) and W-7 (200 µM) (i) before single-trial conditioning. The PIs before (white bars) and at 24 h after conditioning (shaded bars) are shown as means SE. In B, animals in two groups were each injected with 3 µL saline (a,e) and animals in another six groups were each injected with 3 µL saline containing L-DIL (1 mM) (b,f), DCB (1 mM) (c,g), or W-7 (200 µM) (d,h) before multiple-trial conditioning. PIs were measured before (white bars) and at 30 min (hatched bars) or 1 d (shaded bars) after conditioning and are shown as mean SE. In C, animals in five groups were each injected with 3 µL saline containing A23187 (200 µM) (a), A23187 (200 µM) and ODQ (200 µM) (b), A23187 (200 µM) and L-DIL (1 mM) (c), A23187 (200 µM) and W7 (200 µM) (d), or A23187 (200 µM) and DDA (1 mM) (e). The PIs measured before (white bars) and 24 h (shaded bars) after conditioning are shown as means SE. The results of statistical comparison before and 24 h after conditioning are shown. (***) P < 0.001, (NS) P > 0.05, WCX test. The number of animals tested is shown at each data point. The odor preferences at 24 h after conditioning of the groups shown in a, c, d, f, h, and i in A and a,b,c in C did not significantly differ among each other (P > 0.05, K-W test).
Cyclic AMP pathway plays an essential role in formation of long-term memory (LTM). In some species, the nitric oxide (NO)-cyclic GMP pathway has been found to act in parallel and complementary to the cAMP pathway for LTM formation. Here we describe a new role of the NO-cGMP pathway, namely, stimulation of the cAMP pathway to induce LTM. We have studied the signaling cascade underlying LTM formation by systematically coinjecting various "LTM-inducing" and "LTM-blocking" drugs in crickets. Multiple-trial olfactory conditioning led to LTM that lasted for several days, while memory induced by single-trial conditioning decayed away within several hours. Injection of inhibitors of the enzyme forming NO, cGMP, or cAMP into the hemolymph prior to multiple-trial conditioning blocked LTM, whereas injection of an NO donor, cGMP analog, or cAMP analog prior to single-trial conditioning induced LTM. Induction of LTM by injection of an NO donor or cGMP analog paired with single-trial conditioning was blocked by inhibitors of the cAMP pathway, but induction of LTM by a cAMP analog was unaffected by inhibitors of the NO-cGMP pathway. Inhibitors of cyclic nucleotide-gated channel (CNG channel) or calmodulin-blocked induction of LTM by cGMP analog paired with single-trial conditioning, but they did not affect induction of LTM by cAMP analog. Our findings suggest that the cAMP pathway is a downstream target of the NO-cGMP pathway for the formation of LTM, and that the CNG channel and calcium-calmodulin intervene between the NO-cGMP pathway and the cAMP pathway.
 
The behavioral performance of auditory and visual 0-back, 1-back, and 2-back tasks in boys and girls in the three age groups of children. (A) The mean reaction times, (B) mean percentages of incorrect responses, and (C) mean percentages of multiple responses. (F ) Female; (M) male; (Vis) visual tasks; (Aud) auditory tasks. Vertical lines indicate SEM. (*) The difference between boys and girls is significant, P <0.05.
The experimental design. (A) The temporal order of the 0-back, 1-back, and 2-back tasks in the three consecutive blocks of each session. (7 min, 17 sec) The duration of each individual block. (B) Illustration of the visuospatial working memory tasks. The match trials for 0-back, 1-back, and 2-back tasks are marked with vertical arrows. (100 msec) The duration of each stimulus; (3125 msec) the delay period between the stimuli. (C) Illustration of the audiospatial working memory tasks. (L) Left; (M) middle; (R) right.
The neural processes subserving working memory, and brain structures underlying this system, continue to develop during childhood. We investigated the effects of age and gender on audiospatial and visuospatial working memory in a nonclinical sample of school-aged children using n-back tasks. The results showed that auditory and visual working memory performance improves with age, suggesting functional maturation of underlying cognitive processes and brain areas. The gender differences found in the performance of working memory tasks suggest a larger degree of immaturity in boys than girls at the age period of 6-10 yr. The differences observed between the mastering of auditory and visual working memory tasks may indicate that visual working memory reaches functional maturity earlier than the corresponding auditory system.
 
Recent studies using lesion, infusion, and unit-recording techniques suggest that the infralimbic (IL) subregion of medial prefrontal cortex (mPFC) is necessary for the inhibition of conditioned fear following extinction. Brief microstimulation of IL paired with conditioned tones, designed to mimic neuronal tone responses, reduces the expression of conditioned fear to the tone. In the present study we used microstimulation to investigate the role of additional mPFC subregions: the prelimbic (PL), dorsal anterior cingulate (ACd), and medial precentral (PrCm) cortices in the expression and extinction of conditioned fear. These are tone-responsive areas that have been implicated in both acquisition and extinction of conditioned fear. In contrast to IL, microstimulation of PL increased the expression of conditioned fear and prevented extinction. Microstimulation of ACd and PrCm had no effect. Under low-footshock conditions (to avoid ceiling levels of freezing), microstimulation of PL and IL had opposite effects, respectively increasing and decreasing freezing to the conditioned tone. We suggest that PL excites amygdala output and IL inhibits amygdala output, providing a mechanism for bidirectional modulation of fear expression.
 
Mean swim distance (cm) to reach the hidden platform in the Morris water maze on each day of acquisition and reversal training for strains of mice with normal vision (A), albinism (B), retinal degeneration (C), and unknown visual abilities (D).
We calculated visual ability in 13 strains of mice (129SI/Sv1mJ, A/J, AKR/J, BALB/cByJ, C3H/HeJ, C57BL/6J, CAST/EiJ, DBA/2J, FVB/NJ, MOLF/EiJ, SJL/J, SM/J, and SPRET/EiJ) on visual detection, pattern discrimination, and visual acuity and tested these and other mice of the same strains in a behavioral test battery that evaluated visuo-spatial learning and memory, conditioned odor preference, and motor learning. Strain differences in visual acuity accounted for a significant proportion of the variance between strains in measures of learning and memory in the Morris water maze. Strain differences in motor learning performance were not influenced by visual ability. Conditioned odor preference was enhanced in mice with visual defects. These results indicate that visual ability must be accounted for when testing for strain differences in learning and memory in mice because differences in performance in many tasks may be due to visual deficits rather than differences in higher order cognitive functions. These results have significant implications for the search for the neural and genetic basis of learning and memory in mice.
 
The cAMP response element (CRE)-binding protein, CREB, is a transcription factor whose activity in the brain is critical for long-term memory formation. Phosphorylation of Ser133 in the kinase-inducible domain (KID), that in turn leads to the recruitment of the transcriptional coactivator CREB-binding protein (CBP), is thought to mediate the activation of CREB. However, the importance of phosphorylation for CREB binding to DNA and subsequent gene transcription in vivo is controversial. To definitively address the role of CREB phosphorylation in gene transcription and learning and memory, we derived mutant mice lacking the Ser133 phosphorylation site. These mice exhibit normal CREB-mediated gene transcription for a number of genes implicated in learning and memory processes. Furthermore these mice have no deficits in hippocampus- or striatum-dependent learning. Strikingly, our findings show that CREB phosphorylation at Ser133 is not necessary for CREB binding to CRE sites, CREB-mediated transcription, or CREB-mediated behavioral phenotypes associated with learning and memory. © 2015 Briand et al.; Published by Cold Spring Harbor Laboratory Press.
 
Numerous studies have suggested a role for ubiquitin-proteasome-mediated protein degradation in learning-dependent synaptic plasticity; however, very little is known about how protein degradation is regulated at the level of the proteasome during memory formation. The ubiquitin-specific protease 14 (USP14) is a proteasomal deubiquitinating enzyme that is thought to regulate protein degradation in neurons; however, it is unknown if USP14 is involved in learning-dependent synaptic plasticity. We found that infusion of a USP14 inhibitor into the amygdala impaired long-term memory for a fear conditioning task, suggesting that USP14 is a critical regulator of long-term memory formation in the amygdala.
 
Imaging CA3 and CA1 neuronal ensemble activity following exposure to two environmental contexts. In this study, rats were given two 6-min exposures to two environments (epoch 1 and epoch 2), separated by 20 min. Each of the environments was defined by a unique combination of local features (objects in a fixed configuration within the enclosure of the environment) and distal features (defined by the room, such as posters on the walls, lighting differences, etc). Environments A and B were designed to be as dissimilar as possible, with different enclosure geometries (Environment A was square, Environment B was circular), wall heights, different local objects within the environment (and in a different geometrical configuration), and different distal features, as the two environments were in different rooms. Environment A, for the purposes of this review, is the combination of three minor modifications to environment A: (Aobj) a change of the identity of the local objects; (Aconf) a change in the configuration of the same objects; (Ab) moving the exact local environment to a different room. The ensemble activity for both CA3 and CA1 in each of the groups comprising the A/A group was different from groups A/A and A/B, but not with each other, justifying their collapse into a single group. The sequence of the rats' exposure to environments in epochs 1 and 2 defined the behavioral group, with the A/A group being exposed to the same environment twice, the A/A group being exposed to two similar environments, and the A/B group being exposed to two completely different environments. (A) Confocal projection image of Arc/Homer catFISH in area CA3 from an A/B rat; (red) Arc RNA; (green) H1a RNA; (blue) nuclei. (Green arrow) Points to an H1a+ neuron, which was activated only during behavioral epoch 1; (red arrow) points to an Arc+ neuron, which was activated only during behavioral epoch 2; (yellow arrow) points to an Arc/H1a double+ neuron that was activated in both epoch 1 and epoch 2. For quantitative analyses of image stack data, only putative neuronal nuclei, characterized by their large volume and relatively diffuse staining with DAPI, were assessed for the presence or absence of intranuclear foci for Arc or H1a. (B) Analysis of similarity scores of rats from the different behavioral groups. Similarity scores are a single measure derived from the raw staining class data in which individual neuronal nuclei are classified as Arc+, H1a+, Arc/H1a double+, or negative. The derivation of the similarity score is described in detail elsewhere (Vazdarjanova and Guzowski 2004) and provides a normalized measure of overlap between neuronal ensembles active in the two behavioral sessions. A similarity score of 1 indicates a complete overlap of the neuronal ensembles activated by epoch 1 and epoch 2, whereas a similarity score of 0 indicates no overlap beyond that predicted by chance. Each symbol denotes data from an individual rat. In both CA3 and CA1, the similarity scores from rats of the A/A group were significantly different from those of the A/A and A/B groups (P < 0.0001). Notably , the similarity score of the A/A rats was greater in CA3 as compared to CA1 (P = 0.001). Conversely, the similarity score in the A/B condition in CA1 was significantly different from chance (P = 0.02) and greater than in CA3 (P = 0.015). Note the discontinuous pattern of overlap in the A/A, A/A, and A/B groups for region CA3 as compared to region CA1, which is more graded and linear in nature. (C) The within-rat ratio of CA3 to CA1 activity was strongly correlated across behavioral epochs and was not influenced by the nature of epoch 2. The fact that the CA3 and CA1 activity was not correlated across rats (data not shown), but that the CA3/CA1 within-rat ratio was correlated across sessions, suggests that the balance of CA3 to CA1 activity is highly specific to each individual rat. This individual difference in the intrinsic hippocampal CA3:CA1 network dynamic could play a role in determining the capacity of individuals for spatial navigation or mnemonic functions. This figure is based on data and figures from Vazdarjanova and Guzowski (2004) (Copyright 2004 by the Society for Neuroscience).  
Different functions have been suggested for the hippocampus and its subdivisions along both transversal and longitudinal axes. Expression of immediate-early genes (IEGs) has been used to map specific functions onto neuronal activity in different areas of the brain including the hippocampus (IEG imaging). Here we review IEG studies on hippocampal functional dissociations with a particular focus on the CA3 subregion. We first discuss the cellular functions of IEGs and the brain system interactions that govern their dynamic expression in hippocampal neurons to provide a more solid framework for interpreting the findings from IEG studies. We show the pitfalls and shortcomings of conventional IEG imaging studies and describe advanced methods using IEGs for imaging of neuronal activity or functional intervention. We review the current IEG evidence of hippocampal function, subregional-specific contribution to different stages of memory formation, systems consolidation, functional dissociation between memory and anxiety/behavioral inhibition along the septotemporal axis, and different neural network properties of hippocampal subregions. In total, IEG studies provide support for (1) the role of the hippocampus in spatial and contextual learning and memory, (2) its role in continuous encoding of ongoing experience, (3) septotemporal dissociations between memory and anxiety, and (4) a dynamic relationship between pattern separation and pattern completion in the CA3 subregion. In closing, we provide a framework for how cutting-edge IEG imaging and intervention techniques will likely contribute to better understanding of the specific functions of CA3 and other hippocampal subregions.
 
Persistent drug seeking/taking behavior involves the consolidation of memory. With each drug use, the memory may be reactivated and reconsolidated to maintain the original memory. During reactivation, the memory may become labile and susceptible to disruption; thus, molecules involved in plasticity should influence acquisition and/or reconsolidation. Recently, matrix metalloproteinases (MMPs) have been shown to influence neuronal plasticity, presumably by their regulation of extracellular matrix (ECM) molecules involved in synaptic reorganization during learning. We hypothesized that inhibition of MMP activity would impair the acquisition and/or reconsolidation of cocaine-conditioned place preference (CPP) in rats. Intracerebral ventricular (i.c.v.) microinjection of a broad spectrum MMP inhibitor, FN-439, prior to cocaine training suppressed acquisition of CPP and attenuated cocaine-primed reinstatement in extinguished animals. In a separate experiment, the cocaine memory was reactivated on two consecutive days with a cocaine priming injection. On these two days, artificial cerebral spinal fluid (aCSF) or FN-439 was administered either 30 min prior to or 1 min after cocaine-primed reinstatement sessions. Infusion of FN-439 partially impaired retrieval of the cocaine-associated context when given 30 min prior to cocaine. In both groups, however, FN-439 suppressed reinstatement compared with controls on the third consecutive test for cocaine-primed reinstatement, when no FN-439 was given. Control experiments demonstrated that two injections of FN-439 + cocaine given in the home cage, or of FN-439 + saline priming injections in the CPP chambers did not disrupt subsequent cocaine-primed reinstatement. These results show for the first time that (1) MMPs play a critical role in acquisition and reconsolidation of cocaine-induced CPP, and (2) rats demonstrate apparent disruption of reconsolidation by an MMP inhibitor after extinction and while they are under the influence of cocaine during reinstatement.
 
The formation of many forms of long-term memory requires several molecular mechanisms including regulation of gene expression. The mechanisms directing transcription require not only activation of individual transcription factors but also recruitment of transcriptional coactivators. CBP and p300 are transcriptional coactivators that interact with a large number of transcription factors and regulate transcription through multiple mechanisms, including an intrinsic histone acetyltransferase (HAT) activity. HAT activity mediates acetylation of lysine residues on the amino-terminal tails of histone proteins, thereby increasing DNA accessibility for transcription factors to activate gene expression. CBP has been shown to play an important role in long-term memory formation. We have investigated whether p300 is also required for certain forms of memory. p300 shares a high degree of homology with CBP and has been shown to interact with transcription factors known to be critical for long-term memory formation. Here we demonstrate that conditional transgenic mice expressing an inhibitory truncated form of p300 (p300Delta1), which lacks the carboxy-terminal HAT and activation domains, have impaired long-term recognition memory and contextual fear memory. Thus, our study demonstrates that p300 is required for certain forms of memory and that the HAT and carboxy-terminal domains play a critical role.
 
Drug addicts make poor decisions. These decision-making deficits have been modeled in addicts and laboratory animals using reversal-learning tasks. However, persistent reversal-learning impairments have been shown in rats and monkeys only after noncontingent cocaine injections. Current thinking holds that to represent the human condition effectively, animal models of addiction must utilize self-administration procedures in which drug is earned contingently; thus, it remains unclear whether reversal-learning deficits caused by noncontingent cocaine exposure are relevant to addiction. To test whether reversal learning deficits are caused by contingent cocaine exposure, we trained rats to self-administer cocaine, assessed cue-induced cocaine seeking in extinction tests after 1 and 30 d of withdrawal, and then tested for reversal learning more than a month later. We found robust time-dependent increases in cue-induced cocaine seeking in the two extinction tests (incubation of craving) and severe reversal-learning impairments.
 
Step-through latencies (mean + SEM) in seconds on a 48-h inhibitory avoidance retention test. ( A ) Immediate post-training infusions of OFQ/N (0.01, 0.1, 1, 10, or 100 pmol in 0.2 μL) into the BLA dose-dependently impaired memory consolidation. The ␤ 1 -adrenoceptor antagonist atenolol (2 nmol) administered concurrently into the BLA shifted the dose-response effects of OFQ/N to the left ( n = 9–14 rats per group). ( B ) Delayed infusions of OFQ/N (1 pmol in 0.2 μL) into the BLA administered 3 h after training impaired memory consolidation, but OFQ/N given 6 h after training 
Step-through latencies (mean + SEM) in seconds on a 48-h inhibitory avoidance retention test. (A) Immediate post-training infusions of the NOP receptor antagonist [Nphe 1 ]nociceptin(1-13)NH 2 (1, 10, or 100 pmol in 0.2 µL) dose-dependently enhanced memory consolidation. The 1-adrenoceptor antagonist atenolol (2 nmol) administered concurrently into the BLA blocked the enhancing effects of [Nphe 1 ]nociceptin(1-13)NH 2 (n = 8-12 rats per group). (B) Delayed infusions of [Nphe 1 ]nociceptin(1-13)NH 2 (10 pmol in 0.2 µL) administered into the BLA 3 h after training did not enhance memory consolidation (n = 8-10 rats per group). (✶✶) P < 0.01 compared with the corresponding saline group; () P < 0.01 compared with the corresponding [Nphe 1 ]nociceptin(1-13)NH 2 group.
Representative photomicrographs illustrating needle tracks terminating in the BLA (A) and CEA (B). The arrow points to the needle tip. The gray area in the diagram represents the different nuclei of the basolateral complex. (AB) Accessory basal nucleus; (B) basal nucleus; (L) lateral nucleus.
Extensive evidence indicates that the basolateral complex of the amygdala (BLA) mediates hormonal and neurotransmitter effects on the consolidation of emotionally influenced memory and that such modulatory influences involve noradrenergic activation of the BLA. As the BLA also expresses a high density of receptors for orphanin FQ/nociceptin (OFQ/N), an opioid-like peptide with anxiolytic and amnestic properties, the present experiments investigated whether the BLA is involved in mediating OFQ/N effects on memory consolidation and whether such effects require noradrenergic activity. OFQ/N (0.01-100 pmol in 0.2 microL) administered bilaterally into the BLA of male Sprague-Dawley rats immediately after aversively motivated inhibitory avoidance training induced dose-dependent impairment on a 48-h retention trial. The beta(1)-adrenoceptor antagonist atenolol (2.0 nmol) administered concurrently into the BLA potentiated the dose-response effects of OFQ/N. In contrast, immediate post-training infusions of the peptidergic OFQ/N receptor antagonist [Nphe(1)]nociceptin(1-13)NH(2) (1-100 pmol in 0.2 microL) into the BLA enhanced 48-h retention of inhibitory avoidance training, an effect that was blocked by coadministration of atenolol. Delayed infusions of OFQ/N or [Nphe(1)]nociceptin(1-13)NH(2) into the BLA administered either 6 or 3 h after training, respectively, or immediate post-training infusions of OFQ/N into the adjacent central amygdala did not significantly alter retention performance. These findings indicate that endogenously released OFQ/N interacts with noradrenergic activity within the BLA in modulating memory consolidation.
 
The impact of stress on recognition memory. Stress resulted in better recognition of the emotional story, but poorer recognition of the neutral story, compared with the control condition.
Positive correlation between salivary cortisol and false memory recall.
Stressful events frequently comprise both neutral and emotionally arousing information, yet the impact of stress on emotional and neutral events is still not fully understood. The hippocampus and frontal cortex have dense concentrations of receptors for stress hormones, such as cortisol, which at high levels can impair performance on hippocampally dependent memory tasks. Yet, the same stress hormones can facilitate memory for emotional information, which involves interactions between the hippocampus and amygdala. Here, we induced psychosocial stress prior to encoding and examined its long-term effects on memory for emotional and neutral episodes. The stress manipulation disrupted long-term memory for a neutral episode, but facilitated long-term memory for an equivalent emotional episode compared with a control condition. The stress manipulation also increased salivary cortisol, catecholamines as indicated by the presence of alpha-amylase, heart rate, and subjectively reported stress. Stressed subjects reported more false memories than nonstressed control subjects, and these false memories correlated positively with cortisol levels, providing evidence for a relationship between stress and false memory formation. Our results demonstrate that stress, when administered prior to encoding, produces different patterns of long-term remembering for neutral and emotional episodes. These differences likely emerge from differential actions of stress hormones on memory-relevant regions of the brain.
 
No-Reminder Condition. ( A ) Schematic representations of timelines for experimental procedures. The numbers in parentheses represent the number of rats in the various groups. ( B , C ) Contextual fear memory in control and hippocampal groups measured in terms of time spent freezing in the test session after conditioning; surgery intervals of 24 h ( B ) and 28 d ( C ) were used. In the 24-h test, controls displayed context specificity for the fear memory, while hippocampal rats exhibited no memory of the fear response. At 28 d, both control and hippocampal groups exhibited strong contextual fear memory in the training context (CXT-A) and in a different context (CXT-B) with no context specificity. Error bars represent 6 SEM. 
Reminder Condition. ( A ) Schematic representations of timelines for experimental procedures. The numbers in parentheses represent the number of rats in the various groups. ( B , C ) Contextual fear memory in control and hippocampal groups when either CXT-A ( B ) or CXT-B ( C ) was provided as a reminder 28 d after fear conditioning in CXT-A and 24 h before surgery. CXT-A, as a reminder, restored context specificity of the contextual fear memory and, consequently, controls froze only when tested in CXT-A. Hippocampal lesions eliminated the context-specific memory when it was reactivated by CXT-A. CXT-B was not an effective reminder for context specificity and resulted in generalized freezing in control and HPC groups. Error bars represent 6 SEM. 
Lesion representations. (Top) Photomicrographs of representative hippocampal lesion (left) in relation to a normal brain (right). (Bottom) Schematic drawings of minimal (gray) and maximal (black) extents of hippocampal lesions meeting the histological criterion included in our experiment.
After acquisition, memories associated with contextual fear conditioning pass through a labile phase, in which they are vulnerable to hippocampal lesions, to a more stable state, via consolidation, in which they engage extrahippocampal structures and are resistant to such disruption. The process is accompanied by changes in the form of the memory from being context-specific to context-general. However, when revived by a reminder, stable memories once again become labile and susceptible to hippocampal disruption, and memory reconsolidation is needed to stabilize them. This study addressed two questions with respect to this reconsolidation phenomenon: (1) How do reminders reinstate a hippocampally dependent memory trace? (2) As the memory changes from a stable to a labile state after a reminder, does its form remain invariant, or does it also change? Using contextual manipulations at retrieval in a test of contextual fear conditioning, we showed that when the fear-conditioning environment served as a reminder, the reinstated memory regained its context specificity and, as a result, became vulnerable again to the effects of hippocampal lesions. By comparison, exposure to a different environment during the reminder session reinstated a version of the original memory that was dependent primarily on general features of the original context and, consequently, was less affected by hippocampal lesions. These findings, which relate loss of reactivated memories after hippocampal destruction (or inactivation) to changes in memory representation, are interpreted as consistent with the transformation hypothesis of memory processing.
 
Dorsal versus ventral regions of rodent medial prefrontal cortex differentially control fear and drug seeking. The four major subdivisions of rodent medial prefrontal cortex are depicted along the Paxinos and Watson anatomical boundaries (3.0 mm anterior to bregma) (Paxinos and Watson 2005). Activity in the prelimbic (PL) region promotes the expression of conditioned fear and cocaine-seeking behavior. Dorsal to PL is the dorsal anterior cingulate cortex (ACd), which may also promote fear and drug seeking. The infralimbic (IL) cortex, which lies ventral to PL, promotes the extinction of conditioned fear and cocaineseeking behavior. The ventral-most dorsopeduncular cortex (DP) may resemble IL in the ability to inhibit fear and drug seeking. Hence, dorsal regions of medial prefrontal cortex increase fear and drug seeking (arrows up), while ventral regions exert the opposite effect on behavior, decreasing both fear and drug seeking (arrows down). 
Circuit diagram depicting prefrontal regulation of conditioned fear and cocaine-seeking behaviors. The dorsal and ventral subdivisions of medial prefrontal cortex (PFC) are shown at the center, with their respective outputs to the amygdala controlling fear shown at right, and those to the nucleus accumbens controlling cocaine seeking shown at left. The prelimbic (PL) cortex projects to the basal (BA) nucleus of the amygdala, which excites the central (CE) nucleus of the amygdala, thereby promoting the expression of conditioned fear. The BA also receives excitatory input from lateral (LA) amygdala, which also drives the expression of conditioned fear. The infralimbic (IL) cortex, in contrast, excites a class of GABAergic inhibitory neurons known as the intercalated (ITC) cell masses. These neurons inhibit the CE, thereby inhibiting conditioned fear and promoting extinction. By comparison, PL and IL control cocaine seeking via their differential projections to the core and shell subdivisions of the nucleus accumbens. The PL projects to the core, which promotes the expression of cocaine-seeking behavior. For cueinduced cocaine seeking, this may involve an intermediate projection through the BA to access the core (thin green line). The IL projects to the shell, which promotes the expression of extinction. It remains to be determined how output from these two divisions of the accumbens differentially affects cocaine-seeking behavior (see text for details). Green depicts pathways that activate fear and cocaine seeking. Red depicts pathways that inhibit fear and cocaine seeking.
Extinction is a form of inhibitory learning that suppresses a previously conditioned response. Both fear and drug seeking are conditioned responses that can lead to maladaptive behavior when expressed inappropriately, manifesting as anxiety disorders and addiction, respectively. Recent evidence indicates that the medial prefrontal cortex (mPFC) is critical for the extinction of both fear and drug-seeking behaviors. Moreover, a dorsal-ventral distinction is apparent within the mPFC, such that the prelimbic (PL-mPFC) cortex drives the expression of fear and drug seeking, whereas the infralimbic (IL-mPFC) cortex suppresses these behaviors after extinction. For conditioned fear, the dorsal-ventral dichotomy is accomplished via divergent projections to different subregions of the amygdala, whereas for drug seeking, it is accomplished via divergent projections to the subregions of the nucleus accumbens. Given that the mPFC represents a common node in the extinction circuit for these behaviors, treatments that target this region may help alleviate symptoms of both anxiety and addictive disorders by enhancing extinction memory.
 
In essentially every domain of neuroscience, the generally implicit assumption that few, if any, meaningful differences exist between male and female brain function is being challenged. Here we address how this development is influencing studies of the neurobiology of learning and memory. While it has been commonly held that males show an advantage on spatial tasks, and females on verbal tasks, there is increasing evidence that sex differences are more widespread than previously supposed. Differing performance between the sexes have been observed on a number of common learning tasks in both the human and animal literature, many neither purely spatial nor verbal. We review sex differences reported in various areas to date, while attempting to identify common features of sexually dimorphic tasks, and to place these differences in a neurobiological context. This discussion focuses on studies of four classes of memory tasks for which sex differences have been frequently reported: spatial, verbal, autobiographical, and emotional memory. We conclude that the female verbal advantage extends into numerous tasks, including tests of spatial and autobiographical abilities, but that a small but significant advantage may exist for general episodic memory. We further suggest that for some tasks, stress evokes sex differences, which are not normally observed, and that these differences are mediated largely by interactions between stress and sex hormones.
 
Transitive inference training and testing protocol. (A) Description of the stages to train animals on each of the odor pairs. (B) Schematic of the behavioral apparatus used to train and test the animals. Each letter refers to a specific odor used in the task. The " þ " indicates the rewarded odor in each pairing.  
Histological verification of the extent of medial prefrontal cortex damage. (A) Representative sections from three levels along the anterior – posterior axis of the mouse prefrontal cortex in a sham-operated animal. (B) Corresponding sections from an animal given ibotenic acid infusions into the medial prefrontal cortex. (C) A diagram shows the extent of the largest (light gray) and smallest (black) lesion across the 10 animals.  
Performance on the odor pairs. (A) Performance (+SEM) across all four stages of training. Graph shows total number of days to reach criterion. (B) Total number of days to reach criterion across each of four training stages. (C ) Performance (+SEM) on odor pairs presented during probe testing. (SHAM) Sham-operated group; (MPFCX) medial prefrontal lesion group; ( Ã ) P , 0.05.  
Probe pair performance. (A) Preference index for the transitive probe B vs. D. (B) Preference index for the nontransitive probe A vs. E. (SHAM) Sham-operated group; (MPFCX) medial prefrontal lesion group; ( Ã ) P , 0.05.  
"Transitive inference" refers to the ability to judge from memory the relationships between indirectly related items that compose a hierarchically organized series, and this capacity is considered a fundamental feature of relational memory. Here we explored the role of the prefrontal cortex in transitive inference by examining the performance of mice with selective damage to the medial prefrontal cortex. Damage to the infralimbic and prelimbic regions resulted in significant impairment in the acquisition of a series of overlapping odor discrimination problems, such that animals with prefrontal lesions required twice as many trials to learn compared to sham-operated controls. Following eventually successful acquisition, animals with medial prefrontal lesions were severely impaired on a transitive inference probe test, whereas they performed as well as controls on a test that involved a nontransitive judgment from a novel odor pairing. These results suggest that the prefrontal cortex is part of an integral hippocampal-cortical network essential for relational memory organization.
 
Histone acetylation plays a critical role during long-term memory formation. Several studies have demonstrated that the histone acetyltransferase (HAT) CBP is required during long-term memory formation, but the involvement of other HAT proteins has not been extensively investigated. The HATs CBP and p300 have at least 400 described interacting proteins including transcription factors known to play a role in long-term memory formation. Thus, CBP and p300 constitute likely candidates for transcriptional coactivators in memory formation. In this study, we took a loss-of-function approach to evaluate the role of p300 in long-term memory formation. We used conditional knock-out mice in which the deletion of p300 is restricted to the postnatal phase and to subregions of the forebrain. We found that p300 is required for the formation of long-term recognition memory and long-term contextual fear memory in the CA1 area of the hippocampus and cortical areas.
 
Evidence suggests that plasticity of the amygdalar and hippocampal GABAergic system is critical for fear memory formation. In this study we investigated in wild-type and genetically manipulated mice the role of the activity-dependent 65-kDa isozyme of glutamic acid decarboxylase (GAD65) in the consolidation and generalization of conditioned fear. First, we demonstrate a transient reduction of GAD65 gene expression in the dorsal hippocampus (6 h post training) and in the basolateral complex of the amygdala (24 h post training) during distinct phases of fear memory consolidation. Second, we show that targeted ablation of the GAD65 gene in Gad65(-/-) mice results in a pronounced context-independent, intramodal generalization of auditory fear memory during long-term (24 h or 14 d) but not short-term (30 min) memory retrieval. The temporal specificity of both gene regulation and memory deficits in Gad65 mutant mice suggests that GAD65-mediated GABA synthesis is critical for the consolidation of stimulus-specific fear memory. This function appears to involve a modulation of neural activity patterns in the amygdalo-hippocampal pathway as indicated by a reduction in theta frequency synchronization between the amygdala and hippocampus of Gad65(-/-) mice during the expression of generalized fear memory.
 
Schematic drawing of target brain sites. (A) Representative ventral periaqueductal gray lesion. (B) Sites of stimulation electrodes in the dorsal periaqueductal gray. Sham-lesion animals submitted to electrical () or chemical stimulation (). vPAG-lesion animals submitted to electrical (·) or chemical stimulation (), respectively.
(A) Aversive thresholds determined in rats bearing (hatched columns) or not bearing (sham-open columns) electrolytic lesion of the vPAG. (B) Percentage of time spent freezing by rats with sham (hatched columns) or vPAG (open columns) lesions during 6 min. Pre refers to the baseline period; Post, to the testing session 24 h after conditioning. N = 8 for all groups, except for the freezing threshold (lesion = 8, control = 7).  
(A) Freezing and escape behaviors induced by microinjections of semicarbazide (8.0 µg/0.2µL) into the dPAG in rats bearing (hatched columns) or not bearing (open columns) electrolytic lesion of the vPAG. N = 9 for both groups.  
Previously-reported evidence showed that freezing to a context previously associated with footshock is impaired by lesion of the ventral periaqueductal gray (vPAG). It has also been shown that stepwise increase in the intensity of the electrical stimulation of the dorsal periaqueductal gray (dPAG) produces alertness, then freezing, and finally escape. These aversive responses are mimicked by microinjections of GABA receptor antagonists, such as bicuculline, or blockers of the glutamic acid decarboxylase (GAD), such as semicarbazide, into the dPAG. In this work, we examined whether the expression of these defensive responses could be the result of activation of ventral portion of the periaqueductal gray. Sham- or vPAG electrolytic-lesioned rats were implanted with an electrode in the dPAG for the determination of the thresholds of freezing and escape responses. The vPAG electrolytic lesions were behaviorally verified through a context-conditioned fear paradigm. Results indicated that lesion of the vPAG disrupted conditioned freezing response to contextual cues associated with footshocks but did not change the dPAG electrical stimulation for freezing and escape responses. In a second experiment, lesion of the vPAG also did not change the amount of freezing and escape behavior produced by microinjections of semicarbazide into the dPAG. These findings indicate that freezing and escape defensive responses induced by dPAG stimulation do not depend on the integrity of the vPAG. A discussion on different neural circuitries that might underlie different inhibitory and active defensive behavioral patterns that animals display during threatening situations is presented.
 
Many behavioral and electrophysiological studies in animals and humans have suggested that sleep and circadian rhythms influence memory consolidation. In rodents, hippocampus-dependent memory may be particularly sensitive to sleep deprivation after training, as spatial memory in the Morris water maze is impaired by rapid eye movement sleep deprivation following training. Spatial learning in the Morris water maze, however, requires multiple training trials and performance, as measured by time to reach the hidden platform is influenced by not only spatial learning but also procedural learning. To determine if sleep is important for the consolidation of a single-trial, hippocampus-dependent task, we sleep deprived animals for 0-5 and 5-10 h after training for contextual and cued fear conditioning. We found that sleep deprivation from 0-5 h after training for this task impaired memory consolidation for contextual fear conditioning whereas sleep deprivation from 5-10 h after training had no effect. Sleep deprivation at either time point had no effect on cued fear conditioning, a hippocampus-independent task. Previous studies have determined that memory consolidation for fear conditioning is impaired when protein kinase A and protein synthesis inhibitors are administered at the same time as when sleep deprivation is effective, suggesting that sleep deprivation may act by modifying these molecular mechanisms of memory storage.
 
In six experiments we studied the effects of a single re-exposure to a conditioned stimulus (CS; "retrieval trial") prior to extinction training (extinction-reconsolidation boundary) on the development of and recovery from fear extinction. A single retrieval trial prior to extinction training significantly augmented the renewal and reinstatement of extinguished responding. Augmentation of recovery was not observed if the retrieval and extinction training occurred in different contexts. These results contrast with those reported in earlier papers by Monfils and coworkers in rats and by Schiller and coworkers in humans. We suggest that these contrasting results could depend on the contrasting influences of either: (1) occasion-setting contextual associations vs. direct context-CS associations formed as a consequence of the retrieval trial or (2) discrimination vs. generalization between the circumstances of conditioning and extinction.
 
Propranolol administered systemically impairs IA retrieval but has no effect on reconsolidation. Experimental timelines are shown above each experiment. Values of latencies are expressed in seconds (s) and shown as means + SEM. (A) Animals were trained in IA with 0.9 mA footshock intensity, 48 h later they were tested (Test 1), and immediately after, they were injected i.p. with propranolol or saline. Rats were tested again 48 h later (Test 2). No significant effect of treatment was found among groups. (B) Animals underwent the same experimental protocol as in A, except that 0.6 mA footshock intensity was used during training. No significant effect of treatment was found among groups. (C ) Animals underwent the same experimental protocol as in A, except that 0.25 mA footshock intensity was used during training. No significant effect of treatment was found among groups. (D) Animals underwent the same experimental protocol as in B, except that they received an i.p. injection of propranolol 30 min before reactivation (Test 1). Compared with saline, propranolol significantly disrupted IA retention at Test 1 ( * * , P , 0.01), but not at Test 2. 
Propranolol administered systemically after reactivation disrupts both cue-and contextual-FC reconsolidation. Cue reactivation influences contextual memory. Experimental timelines are shown above each experiment. Freezing scores are expressed as percent of the total time of testing and shown as means + SEM. (A) Rats were trained in FC. Forty-eight hours later, they were presented with the tone in a different context and immediately after, received an i.p. injection of either propranolol or saline. Forty-eight hours and 96 h later, the animals were tested for cue and contextual freezing, respectively. Propranolol significantly disrupted FC in both cue and context tests ( * * * , P , 0.001). (B) Rats were trained in FC and exposed to the training context 48 h later. Immediately after context exposure, rats received an i.p. injection of either propranolol or saline. Forty-eight hours and 96 h later, the animals were tested for contextual and cue freezing, respectively. Propranolol significantly disrupted contextual fear memory but did not affect cued fear memory ( * * * , P , 0.001). (C) Rats were trained and received propranolol or saline injections as in A in the absence of reactivation. Forty-eight hours and 96 h after the injection, the animals were tested for cue and contextual freezing, respectively. No significant differences were found among groups. (D) Rats were trained in FC and 48 h later were exposed to a novel context without any cue presentation and immediately after were injected with propranolol or saline i.p. Forty-eight hours and 96 h later, the animals were tested for cue and contextual freezing, respectively. No significant differences were found among groups. 
Propranolol disrupts freezing but not IA memory. Experimental timelines are shown above each experiment. Values of latencies are expressed in seconds and shown as means + SEM. Freezing scores are expressed as percent of the total time of testing and shown as means + SEM. (A) Rats were trained in IA with 0.6 mA footshock intensity and, during training, presented with a tone. Forty-eight hours later, they were tested in IA, and immediately after, received an i.p. injection of propranolol or saline. Forty-eight hours and 96 h later, the animals were tested in IA and cue fear memory, respectively. Compared with saline, propranolol had no effect on IA retention, but significantly disrupted freezing (Student's t-test, [ * * ] P , 0.01). (B) Rats underwent the same protocol as in A, except that a footshock of 0.9 mA was used during training. Compared with saline, propranolol had no effect on IA retention, but significantly disrupted freezing (Student's t-test, [ * ] P , 0.05). 
Previous studies suggested that the beta-adrenergic receptor antagonist propranolol might be a novel, potential treatment for post-traumatic stress disorder (PTSD). This hypothesis stemmed mainly from rodent studies showing that propranolol interferes with the reconsolidation of Pavlovian fear conditioning (FC). However, subsequent investigations in humans have produced controversial evidence about the effect of propranolol on fear memories and an effect on PTSD symptomatology has yet to be reported. Thus, it remains to be established whether propranolol interferes with the reconsolidation of fear memories at large. To address this question, we tested the effect of systemic injections of propranolol administered before or after the retrieval of an inhibitory avoidance (IA) memory elicited with different footshock intensities. In parallel, the same treatment was tested on the reconsolidation of Pavlovian FC. Propranolol showed no effect on the reconsolidation of IA, although the pre-retrieval administration resulted in a significant retrieval impairment. This impairment was transient, and memory returned to control levels at later times. In agreement with previous studies, we found that systemic administration of propranolol disrupts the reconsolidation of Pavlovian FC and that its injection following a retrieval elicited by cue exposure also interferes with the reconsolidation of contextual FC. Hence, propranolol disrupts the reconsolidation of Pavlovian FC, but has no effect on the reconsolidation of IA. The results indicate that the efficacy of systemic administration of propranolol in disrupting the reconsolidation of fear memories is limited.
 
A distributed limbic-corticostriatal circuitry is implicated in cue-induced drug craving and relapse. Exposure to drug-paired cues not only precipitates relapse, but also triggers the reactivation and reconsolidation of the cue-drug memory. However, the limbic cortical-striatal circuitry underlying drug memory reconsolidation is unclear. The aim of this study was to investigate the involvement of the nucleus accumbens core and the basolateral amygdala in the reconsolidation of a cocaine-conditioned stimulus-evoked memory. Antisense oligodeoxynucleotides (ASO) were infused into each structure to knock down the expression of the immediate-early gene zif268, which is known to be required for memory reconsolidation. Control infusions used missense oligodeoxynucleotides (MSO). The effects of zif268 knockdown were measured in two complementary paradigms widely used to assess the impact of drug-paired CSs upon drug seeking: the acquisition of a new instrumental response with conditioned reinforcement and conditioned place preference. The results show that both intranucleus accumbens core and intrabasolateral amygdala zif268 ASO infusions at memory reactivation impaired the reconsolidation of the memory underlying a cocaine-conditioned place preference. However, knockdown of zif268 in the nucleus accumbens at memory reactivation had no effect on the memory underlying the conditioned reinforcing properties of the cocaine-paired CS measured subsequently, and this is in contrast to the marked impairment observed previously following intrabasolateral amygdala zif268 ASO infusions. These results suggest that both the basolateral amygdala and nucleus accumbens core are key structures within limbic cortical-striatal circuitry where reconsolidation of a cue-drug memory occurs. However reconsolidation of memory representations formed during Pavlovian conditioning are differentially localized in each site.
 
Human and preclinical models of addiction demonstrate that gonadal hormones modulate acquisition of drug seeking. Little is known, however, about the effects of these hormones on extinction of drug-seeking behavior. Here, we investigated how 17β-estradiol (E2) affects expression and extinction of cocaine seeking in female rats. Using a conditioned place preference (CPP) paradigm, ovariectomized rats were maintained throughout conditioning with 2 d of E2 treatment followed by 2 d of vehicle treatment, or were injected with E2 daily. Hormone injections were paired or explicitly unpaired with place conditioning sessions. Expression of a cocaine CPP was of equal magnitude regardless of conditioning protocol, suggesting that E2 levels during conditioning did not affect subsequent CPP expression. During extinction, daily E2 administration initially enhanced expression of the cocaine CPP, but resulted in significantly faster extinction compared to controls. Whereas E2-treated rats were extinguished within 8 d, vehicle-treated rats maintained CPP expression for more than a month, indicative of perseveration. To determine whether E2 could rescue extinction in these rats, half were given daily E2 treatment and half were given vehicle. E2-treated rats showed rapid extinction, whereas vehicle-treated rats continued to perseverate. These data demonstrate for the first time that E2 is necessary for extinction of cocaine seeking in female rats, and that it promotes rapid extinction when administered daily. Clinically, these findings suggest that monitoring and maintaining optimal E2 levels during exposure therapy would facilitate therapeutic interventions for female cocaine addicts.
 
Grouping procedure. Each box represents one stage of the behavioral protocol: 5 min of contextual pre-exposure, an immediate footshock on training day, or 5 min of context exposure during test.
Mean percent freezing after contextual fear conditioning. Rats underwent three phases of contextual fear conditioning on three different days: pre-exposure, training, and testing. The 17/18/19 group and-/24/25 group significantly differed from the 17/24/25 group, 23/ 25/25 group, and the 23/30/31 group. Groups * significantly differed from groups ^ (P , 0.05). Groups with the same symbol did not significantly differ.
Mean percent freezing after contextual fear conditioning. Rats were pre-exposed to either the training context (pre-exposed A) or a control context (pre-exposed B) on PD 17, received a footshock immediately upon placement into the training context on either PD 18 or PD 24, and were tested on PD 25. The ( * ) symbol indicates that this group significantly differed from all other groups, P , 0.05.
Histology. ( A ) Coronal brain sections (30 m m) stained with cresyl violet from a bilateral dorsal hippocampus lesion on PD 15 / 16 ( top ) and PD 21 / 22 ( bottom ). ( B ) Diagrammatic reconstruc- tion of coronal sections adapted from Sherwood and Timiras (1970) and reprinted here with express 
Mean percent freezing after contextual fear conditioning in hippocampal lesion and sham lesion rats. Rats underwent bilateral hippocampus or sham lesions prior to the pre-exposure phase. Mean percent freezing exhibited by the sham-lesioned rats significantly differed from the hippocampus-lesioned rats in both the 17 / 24 / 25 and 23 / 30 / 31 groups ( ∗ P , 0.05). 
Long-term memory for fear of an environment (contextual fear conditioning) emerges later in development (postnatal day; PD 23) than long-term memory for fear of discrete stimuli (PD 17). As contextual, but not explicit cue, fear conditioning relies on the hippocampus; this has been interpreted as evidence that the hippocampus is not fully developed until PD 23. Alternatively, the hippocampus may be functional prior to PD 23, but unable to cooperate with the amygdala for fearful learning. The current experiments investigate this by separating the phases of conditioning across developmental stages. Rats were allowed to learn about the context on one day and to form the fearful association on another. Rats exposed to the context on PD 17 exhibited significant fear only when trained and tested a week later (PD 23, 24), but not on consecutive days (PD 18, 19), demonstrating that rats can learn about a context as early as PD 17. Further experiments clarify that it is associative mechanisms that are developing between PD 18 and 23. Finally, the hippocampus was lesioned prior to training to ensure the task is being solved in a hippocampus-dependent manner. These data provide compelling evidence that the hippocampus is functional for contextual learning as early as PD 17, however, its connection to the amygdala or other relevant brain structures may not yet be fully developed.
 
It has been demonstrated previously on the radial maze that the emergence of an age-related mnemonic impairment is critically dependent on the form which the discrimination problems took. Hence, when the arms were presented one by one (i.e., successive go-no-go discrimination), both adult and aged mice learned to distinguish between positive (baited) and negative (unbaited) arms readily, as evidenced by their increased readiness to enter positive relative to negative arms (i.e., by a differential in arm-entry latencies). A selective impairment in the aged mice was seen when these arms were presented subsequently as pairs, such that the mice were confronted with an explicit choice (i.e., simultaneous 2-choice discrimination). When discriminative performance was measured by the differential run speed between positive and negative arms, aged mice were also impaired. This was particularly pronounced in the 2-choice discrimination condition. We examined the effects of tacrine (3mg/kg, subcutaneously) or S 17092 (10mg/kg, orally) in aged mice on the three behavioral indices of this 2-stage spatial discrimination paradigm. The results indicated that: (1) Tacrine, but not S 17092, enhanced the acquisition of go-no-go discrimination as reflected in arm-entry latencies; (2) both drugs improved choice accuracy in simultaneous discrimination, although the effect of tacrine was less striking and, in particular, far from statistical significance in the very first 2-choice responses; and (3) neither drugs significantly affected run-speed performance. We conclude further that the specific patterns of drug effects on the three indices of discriminative performance might suggest that each index is associated with a distinct form of mnemonic expression relying on separate neural systems.
 
Flupenthixol blocks an increase in goal approach induced by a motivational state shift, but does not affect reward palatability. (A) The effects of acute administration of flupenthixol (0.5 mg/kg i.p) on magazine entries during noncontingent re-exposure to the sucrose reward in either the control 3-h or novel 23-h elevated food deprivation state. (B) The effects of acute administration of flupenthixol during the same test on a reward " liking " -related lick frequency measure (the y-axis is truncated at 3.5 licks/sec based on our observation of this frequency as the floor licking rate). * P , 0.05, * * P , 0.01, * * * P , 0.001. N ¼ 16, per drug group.  
Flupenthixol does not affect instrumental incentive learning. The test of the effects of instrumental incentive learning on reward seeking was conducted off drug and lever press performance was unrewarded . (A) The effects of flupenthixol (0.5 mg/kg i.p) during the previous day's noncontingent re-exposure session on reward-seeking actions (normalized to baseline pretest response rates) in both the control 3-h and novel elevated 23-h food-deprived state. (B) For the vehicle-treated rats only, reward-seeking response rate plotted as a percentage of baseline (BL) for the test of incentive learning separated into five 1-min time bins. (C) For the flupenthixol-treated rats only, rewardseeking response rate, plotted as a percentage of BL, for the test of incentive learning separated into five 1-min time bins. * P , 0.05, * * P , 0.01. N ¼ 16, per drug group.  
Here we attempted to clarify the role of dopamine signaling in reward seeking. In Experiment 1, we assessed the effects of the dopamine D(1)/D(2) receptor antagonist flupenthixol (0.5 mg/kg i.p.) on Pavlovian incentive motivation and found that flupenthixol blocked the ability of a conditioned stimulus to enhance both goal approach and instrumental performance (Pavlovian-to-instrumental transfer). In Experiment 2 we assessed the effects of flupenthixol on reward palatability during post-training noncontingent re-exposure to the sucrose reward in either a control 3-h or novel 23-h food-deprived state. Flupenthixol, although effective in blocking the Pavlovian goal approach, was without effect on palatability or the increase in reward palatability induced by the upshift in motivational state. This noncontingent re-exposure provided an opportunity for instrumental incentive learning, the process by which rats encode the value of a reward for use in updating reward-seeking actions. Flupenthixol administered prior to the instrumental incentive learning opportunity did not affect the increase in subsequent off-drug reward-seeking actions induced by that experience. These data suggest that although dopamine signaling is necessary for Pavlovian incentive motivation, it is not necessary for changes in reward experience, or for the instrumental incentive learning process that translates this experience into the incentive value used to drive reward-seeking actions, and provide further evidence that Pavlovian and instrumental incentive learning processes are dissociable.
 
(See also Supplemental Fig. S1.) Effect of reconsolidation-extinction procedure on the expression of remote fear memory. (A) Experimental schedule. Fear-conditioned mice were randomly assigned to three different experimental groups 29 d after training: Ret-Ext (n ¼ 8) mice underwent the retrieval trial (Ret) in conditioning chamber A for 3 min and then underwent 30 min of extinction training (Extinction) 1 h after retrieval in the same chamber; Ext (n ¼ 8) mice were submitted to 33 min of extinction training in chamber A; No-Treat (n ¼ 8) mice were not exposed to either memory retrieval or to an extinction session. All mice were submitted to a contextual memory test (CTX) in chamber A and to a fear sensitization test (TONE) in chamber B (T1). Thirty days after T1 all mice underwent CTX in chamber A and TONE in chamber C (T2) in order to evaluate spontaneous recovery and fear incubation, respectively. (B) Percentage of freezing responses recorded during retrieval and extinction. (C,D) Percentage of freezing response recorded during (C ) contextual (CTX) and (D) fear sensitization (TONE) tests. All data are shown as mean + SEM. ( * ) P , 0.05 vs. all other groups. 
(See also Supplemental Fig. S2.) Fear-conditioned mice (No-treat, Ret-Ext, and Ext) showed higher anxiety levels, social withdrawal, and spatial memory deficits compared with no-shocked control mice (NS, n ¼ 8). (A) In the Plus Maze, anxiety levels were evaluated as percentage of time spent in open arms (left) and the number of crossings to open arms (right). (B) Social avoidance/approach index was calculated as the ratio of the time spent in social and nonsocial chambers of the apparatus (T social /(T social + T nonsocial ). (C ) Escape latency is expressed in seconds required to reach the hidden platform in a spatial water maze task (six trials per day; trial/block 1-18). ( * ) P , 0.05 vs. all other groups. 
(See also Supplemental Fig. S3.) Effect of extinction after a longer retrieval trial on expression of remote fear memory. (A) Experimental schedule. Fear-conditioned mice were randomly assigned to two different experimental groups 29 d after training: L-Ret-Ext (n ¼ 8) mice underwent a longer retrieval trial (Long-Ret) in conditioning chamber A (15 min) and then submitted to 30 min of extinction training (Extinction) 1 h after retrieval in the same chamber; No-Treat (n ¼ 8) mice were not exposed to either memory retrieval or extinction session. All mice were submitted to a contextual memory test (CTX) in chamber A and to a fear sensitization test (TONE) in chamber B (T1). Thirty days after T1, all mice underwent CTX in chamber A and TONE in chamber C (T2) in order to evaluate spontaneous recovery and fear incubation, respectively. (B) Percentage of freezing response recorded during long retrieval and extinction. (C,D) Percentage of freezing response recorded during (C) contextual (CTX) and (D) fear sensitization (TONE) tests. All data are shown as mean + SEM. ( * ) P , 0.05 vs. L-Ret-Ext mice. 
(See also Supplemental Fig. S4.) An immediate footshock procedure induces fear sensitization but not contextual memory. (A) Experimental schedule. Mice were randomly assigned to three different groups: C/S-Hi (n ¼ 9) received a high electric footshock (0.7 mA); C/S-PT (n ¼ 9) received a pain threshold electric footshock; S-Hi (n ¼ 9) received an immediate high electric footshock (0.7 mA) and was immediately removed from the apparatus. Thirty days after training (T1), all mice were submitted to a contextual memory test (CTX) in conditioning chamber A and to a fear sensitization test (TONE) in chamber B. Thirty days after T1, all mice underwent CTX in chamber A and TONE in chamber C (T2) in order to evaluate spontaneous recovery and fear incubation, respectively. (B,C ) Percentage of freezing response recorded during (B) contextual (CTX) and (C ) fear sensitization (TONE) tests. All data are shown as mean + SEM. ( * ) P , 0.05.
(See also Supplemental Fig. S5.) An immediate footshock procedure induces anxiety behavior, social withdrawal, and spatial memory deficits. Performances of shocked mice (C/S-Hi; C/S-PT; S-Hi) were compared with those of no-shocked control mice (NS, n ¼ 8). (A) In the Plus Maze, anxiety levels were evaluated as percentage of time spent in open arms (left) and number of crossings to open arms (right). (B) Social avoidance/approach index was calculated as the ratio of the time spent in social and nonsocial chambers of the apparatus (T social / (T social + T nonsocial ). (C ) Escape latency is expressed in seconds required to reach the hidden platform in a spatial water maze task (six trials per day; trial/block 1-18). ( * ) P , 0.05. 
Long-lasting memories of adverse experiences are essential for individuals' survival but are also involved, in the form of recurrent recollections of the traumatic experience, in the aetiology of anxiety diseases (e.g., post-traumatic stress disorder [PTSD]). Extinction-based erasure of fear memories has long been pursued as a behavioral way to treat anxiety disorders; yet, such a procedure turns out to be transient, context-dependent, and ineffective unless it is applied immediately after trauma. Recent evidence indicates that, in both rats and humans, extinction training can prevent the return of fear if administered within the reconsolidation window, when memories become temporarily labile and susceptible of being updated. Here, we show that the reconsolidation-extinction procedure fails to prevent the spontaneous recovery of a remote contextual fear memory in a mouse model of PTSD, as well as the long-lasting behavioral abnormalities induced by traumatic experience on anxiety and in both social and cognitive domains (i.e., social withdrawal and spatial learning deficits). Such a failure appears to be related to the ineffectiveness of the reconsolidation-extinction procedure in targeting the pathogenic process of fear sensitization, a nonassociative component of traumatic memory that causes animals to react aberrantly to harmless stimuli. This indicates fear sensitization as a major target for treatments aimed at mitigating anxiety and the behavioral outcomes of traumatic experiences.
 
Silencing of a single gene, FMR1, is linked to a highly prevalent form of mental retardation, characterized by social and cognitive impairments, known as fragile X syndrome (FXS). The FMR1 gene encodes fragile X mental retardation protein (FMRP), which negatively regulates translation. Knockout of Fmr1 in mice results in enhanced long-term depression (LTD) induced by metabotropic glutamate receptor (mGluR) activation. Despite the evidence implicating FMRP in LTD, the role of FMRP in long-term potentiation (LTP) is less clear. Synaptic strength can be augmented heterosynaptically through the generation and sequestration of plasticity-related proteins, in a cell-wide manner. If heterosynaptic plasticity is altered in Fmr1 knockout (KO) mice, this may explain the cognitive deficits associated with FXS. We induced homosynaptic plasticity using the β-adrenergic receptor (β-AR) agonist, isoproterenol (ISO), which facilitated heterosynaptic LTP that was enhanced in Fmr1 KO mice relative to wild-type (WT) controls. To determine if enhanced heterosynaptic LTP in Fmr1 KO mouse hippocampus requires protein synthesis, we applied a translation inhibitor, emetine (EME). EME blocked homo- and heterosynaptic LTP in both genotypes. We also probed the roles of mTOR and ERK in boosting heterosynaptic LTP in Fmr1 KO mice. Although heterosynaptic LTP was blocked in both WT and KOs by inhibitors of mTOR and ERK, homosynaptic LTP was still enhanced following mTOR inhibition in slices from Fmr1 KO mice. Because mTOR will normally stimulate translation initiation, our results suggest that β-AR stimulation paired with derepression of translation results in enhanced heterosynaptic plasticity.
 
Major brain functions depend on neuronal processes that favor the plasticity of neuronal circuits while at the same time maintaining their stability. The mechanisms that regulate brain plasticity are complex and engage multiple cascades of molecular components that modulate synaptic efficacy. Protein kinases (PKs) and phosphatases (PPs) are among the most important of these components that act as positive and negative regulators of neuronal signaling and plasticity, respectively. In these cascades, the PP protein phosphatase 2B or calcineurin (CaN) is of particular interest because it is the only Ca(2+)-activated PP in the brain and a major regulator of key proteins essential for synaptic transmission and neuronal excitability. This review describes the primary properties of CaN and illustrates its functions and modes of action by focusing on several representative targets, in particular glutamate receptors, striatal enriched protein phosphatase (STEP), and neuromodulin (GAP43), and their functional significance for synaptic plasticity and memory.
 
A shows that there was no significant effect of dose on attentional accuracy [F(2, 24) = 1.53, P < 0.05]. However, these animals made significantly more omissions at the high dose [F(2, 24) = 9.42, P < 0.01; see Fig. 2B] and were also slower to respond to the target [F(2, 24) = 7.55, P < 0.01; means (SEM) in csec, veh-67.2 (2.93), 3 µg dose-67.2 (2.89), 10 µg dose-77.5 (2.85)]. ANOVA showed a dose-dependent effect of scopolamine on the number of premature responses [F(2, 24) = 5.93, P < 0.01] that was characterized by an increase in the number of responses made at the low dose and a reduction in responses made at the high dose [means (SEM), veh-113 (21.2), 3 µg dose-137 (21.5), 10 µg dose-94 (20.8)]. The number of perseverative nose pokes made during the delay also significantly reduced at the high dose [F(2, 24) = 4.12, P < 0.01; means (SEM), veh-98 (18.9), 3 µg dose-93 (18.0), 10 µg dose-67 (16.9)]. During the memory phase of the task, overall, choice accuracy systematically declined as a function of delay [F(3, 36) = 11.71, P < 0.001]. The high dose of scopolamine produced a delay-independent reduction in choice accuracy across all delays [F(2, 24) = 13.31, P < 0.001; see Fig. 2C]. There was no dose x delay interaction [F(6, 72) = 0.45, P > 0.05]. In addition, the high dose of scopolamine made all animals slower to respond correctly to the choice stimulus [F(2, 24) = 5.60, P < 0.01; means (SEM) in csec, veh-116 (4.70), 3 µg dose-111 (3.22), 10 µg dose-123 (3.79)] and lengthened magazine latencies [F(2, 24) = 6.71, P < 0.01; means (SEM) in csec, veh-143 (8.86), 3 µg dose-143 (8.20), 10 µg dose-220 (36.25)]. Thus, although a high dose of scopolamine infused into the mPFC did not impair the animals' ability to correctly detect the target in the attentional phase, these animals were unable to accurately respond to the matching target stimulus during the choice (memory) phase of the task, even at the short delay. Speed of responding was also significantly impaired.
Mean (SEM) performance of animals with intra-mPFC infusions of vehicle, 3 and 10 µg scopolamine on attentional (target) accuracy (A), target omissions (B), and memory for the target during the choice phase (C).
Representative photomicrographs of sections of the basal forebrain showing ChAT-IR neurons in the Ch4 region of sham ( a , left ) and 192 IgG-saporin lesioned ( b , right ) rats. It can be seen that the magnocellular ChAT-IR neurons of the Ch4 cell group (nucleus basalis magnocellularis; nBM) are greatly reduced in number after intrabasalis infusions of 192 IgG-saporin. (v) Ventral; (m) medial; (gp) globus pallidus. 
Mean (SEM) performance of Sham controls and nBM lesioned animals on postoperative baseline testing for 8 d on attentional (target) accuracy (A), target omissions (B), and memory for the target during the choice phase (C).
Two experiments examined the effects of reductions in cortical cholinergic function on performance of a novel task that allowed for the simultaneous assessment of attention to a visual stimulus and memory for that stimulus over a variable delay within the same test session. In the first experiment, infusions of the muscarinic receptor antagonist scopolamine into the medial prefrontal cortex (mPFC) produced many omissions but did not impair rats' ability to correctly detect a brief visual stimulus. However, these animals were highly impaired in remembering the location of that stimulus following a delay period, although in a delay-independent manner. In the second experiment, another group of animals with selective 192 IgG-saporin lesions of the nucleus basalis magnocellularis (nBM) were not impaired under conditions of low-attentional demand. However, when the stimulus duration was reduced, a significant memory impairment was observed, but similar to the results of the first experiment, the nBM-lesioned animals were not impaired in attentional accuracy, although aspects of attention were compromised (e.g., omissions). These findings demonstrate that (1) cortical cholinergic depletion produces dissociable deficits in attention and memory, depending on the task demands, (2) delay-independent mnemonic deficits produced by scopolamine are probably due to impairments other than simple inattention, and (3) working memory deficits are not simply dependent on attentional difficulties per se. Together, these findings implicate the nBM cortical cholinergic system in both attentional and mnemonic processing.
 
Top-cited authors
Larry Squire
  • University of California, San Diego
Stephen Maren
  • Texas A&M University
Robert Clark
  • University of California, San Diego
Reginald Frederick Westbrook
  • UNSW, Sydney, Australia
Lynn Nadel
  • The University of Arizona