Effect of age of habit on susceptibility to cycloheximide-induced amnesia in mice.

The amnesic effects of cycloheximide (CYC) on habits of different ages were investigated in a food-motivated, discrimination-reversal task. Groups of C57BL/6J mice were injected 30 min before training or immediately, 3 days, 6 days, or 9 days after training. Retention was tested 24 hr after CYC treatment. The usual amnesic effect of CYC occurred in animals injected before training. No amnesia was apparent in groups injected immediately, 3 days, or 9 days after training. However, a reliable and reproducible amnesia occurred in the group injected 6 days after training. This amnesia could be reversed by pretest treatment with a monoamine oxidase inhibitor, pheniprazine. Pheniprazine, given 7 days after training, also enhanced retrieval of memory in saline-injected mice.     

Role of muscarinic M1 receptors in inhibitory avoidance and contextual fear conditioning

The objective of the present study was to observe the effects of pre-training or post-training administration of dicyclomine, a M1 muscarinic antagonist, on inhibitory avoidance (IA) and contextual fear conditioning (CFC) and to investigate if the effects observed with the pre-training administration of dicyclomine are state-dependent. For each behavioral procedure (IA and CFC) groups of Wistar male rats were treated with saline or dicyclomine either 30 min before training (pre-training), immediately after training or 30 min before training/30 min before test (pre-training/pre-test). The animals were tested 24 h after training. The acquisition of IA and CFC was impaired by pre-training administration of dicyclomine. The consolidation of both tasks was not affected by dicyclomine given immediately after training. Pre-training/pre-test administration of dicyclomine impaired both tasks, an effect similar to that observed in the group which only received pre-training administration. Pre-test treatment induced dissociation between both tasks, impairing CFC retrieval, without interfering with the animals avoidance response. These results show that the dicyclomine did not affect IA and CFC consolidation, suggesting specific involvement of M1 muscarinic receptor only in acquisition these tasks, and these effects was not state-dependent. However, it is possible that the retrieval of these tasks may be mediated, at least in part, by different neurochemical mechanisms and may be dissociated by dicyclomine.     

Two critical periods of protein and glycoprotein synthesis in memory consolidation for visual categorization learning in chicks

A protein synthesis inhibitor, anisomycin (ANI), and an inhibitor of glycoprotein synthesis, 2-deoxygalactose (2-D-gal), were used to investigate memory consolidation following visual categorization training in 2-day-old chicks. ANI (0.6 micromole/chick) and 2-D-gal (40 micromoles/chick) were injected intracerebrally at different time intervals from 1 hr before to 23 hr after the training. Retention was tested 24 hr post-training. Both ANI and 2-D-gal injections revealed two periods of memory sensitivity to pharmacological intervention. ANI impaired retention when injected from 5 min before to 30 min after the training or from 4 hr to 5 hr post-training, thus demonstrating that consolidation of long-term memory in this task requires two periods of protein synthesis. 2-D-Gal first produced an amnesia when it was injected in the interval from 5 min before to 5 min after the training. Injections made between 5 min and 5 hr post-training were without effect on the retention. The second period of memory impairment by 2-D-gal started at 5 hr post-training and lasted until 21 hr after the training. Administration of 2-D-gal made 23 hr after the training did not influence retention in the test at either 24 hr or 26 hr. These results are consistent with the hypothesis that two waves of protein and glycoprotein synthesis are necessary for the formation of long-term memory. The prolonged duration of performance impairment by 2-D-gal in the present task might reflect an extended memory consolidation period for a categorization form of learning.     

Memory as a state dependent phenomenon: role of ACTH and epinephrine

Rats received an ip injection of saline, epinephrine HCl (5.0 micrograms/kg), or ACTH1--24(0.2 microgram/kg) immediately after training in a step-down inhibitory avoidance task, and an ip injection of saline, or epinephrine (0.5, 1.0, 2.0, 3.0, 5.0, or 10.0 mu/kg), or ACTH (0.02, 0.04 0.08, 0.12, 0.2, or 0.4 microgram/kg) 6 min prior to a test session of the same task 24 hr after training. Retention was excellent in the groups treated with saline after training, and poor in those treated with ACTH or epinephrine after training and tested under saline. The amnestic effect of the drugs was counteracted by their administration prior to testing, and there was a dose-response curve for this effect: partial recovery from amnesia was obtained with 20 or 40% of the amnestic dose and full recovery was obtained with 60 to 100% of that dose when the same drug was given after training and prior to testing; if the drugs given after training and prior to testing were not the same, full recovery was obtained only with twice the amnestic dose of the latter. These findings are consistent with the hypothesis that learning depends on the relation between the endogenous levels of these two hormones in the post-training period and during testing. The animals seem capable of discriminating between the two drugs only partially.     

Sequence-Specific Impairment of Memory Formation by NCAM Antisense Oligonucleotides

The functional role of NCAM gene expression in memory formation was studied in the one-trial passive avoidance task in day-old chicks by pretraining injections of one of three different 18-mer end-protected oligonucleotides corresponding to positions 190-, 207-, and 332- of the NCAM Ig1 domain. Twenty-four-hour-old chicks were trained by pecking at a bitter-tasting bead and tested for avoidance 30 min, 3, 8, or 24 hr later. Memory retention was significantly reduced only in the group of animals injected with the NCAM antisense corresponding to position 207- (AS-ODN-207), and only if given twice, both immediately after hatching and 12 hr before training. This antisense was without effect on the general behavior of the chicks, training or acquisition, and did not produce observable neurotoxic damage. Under such conditions amnesia was evident by 3 hr after training and lasted until at least 24 hr after training. The two other tested oligonucleotides were without behavioral effect. To control for nonsequence-specific effects of AS-ODN-207, brains from injected and trained animals were processed for Western blotting and probed using anti-NCAM, anti-L1, and anti-actin antibodies. NCAM antisense corresponding to position 207- significantly reduced the level of NCAM, whereas the level of L1 and actin remained unchanged. These results confirm our earlier conclusion that NCAM is necessary for longer term memory retention.     

Role of the biogenic amines in the reversal of cycloheximide-induced amnesia.

Amnesia was induced by pretraining injections of cycloheximide (CYC) in a food motivated discrimination reversal task. Magnitude of amnesia varied as a function of the amount of training on both the initial discrimination and the reversal and also as a function of the length of intertrial interval used on both the reversal and the test. Memory spontaneously recovered 48 hr. following reversal training. Recovery from amnesia was induced by pretesting injections of d-amphetamine and 2 monoamine oxidase inhibitors, pargyline and catron. This enhanced performance was a true recovery of the memory and not a result of enhanced learning or increased arousal. Depletion of catecholamines by alpha-methyl-para-tyrosine, a tyrosine hydroxylase inhibitor, and diethyldithiocarbamate, a dopamine beta hydroxylase inhbitor, resulted in an amnesia quantitatively and qualitatively similar to amnesia induced by CYC. These data support the hypothesis that CYC-induced amnesia is mediated via central catecholamines.     

Stability of long temporal gradients of retrograde amnesia in mice

Mice were given a single training trial and then received a series of four electroconvulsive shocks (ECS), 1 h apart, at one of several times after training (1-180 days). Retention was then tested at one of three times after ECS: 7, 14, or 28 days. Control animals that received sham treatment exhibited gradual forgetting with increasing training-retention intervals. Mice given ECS exhibited temporally graded retrograde amnesia, which affected memories acquired up to about 14 days before treatment. The retrograde amnesia was relatively stable, maintaining its temporally graded appearance for at least 28 days after ECS. Some recovery may have occurred in the case of memories acquired 7 days or longer before ECS, but memories acquired only 1 or 5 days before ECS did not recover. These findings extend the parallel between experimental amnesia in laboratory animals and human amnesia.     

Scopolamine amnesia of passive avoidance: a deficit of information acquisition

Despite its increasing use as an animal model of memory deficit in human dementia, relatively few studies have attempted to assess the memory processes involved in the anticholinergic-induced impairment of passive avoidance retention. In the present experiments, the influence of scopolamine administered prior to or immediately following training on 24-h retention of step-through passive avoidance was studied in NMRI mice. In low doses (0.3-3.0 mg/kg ip) pretraining administration (-5 min) of scopolamine induced a very strong amnesia. Post-training scopolamine induced a significant effect only at the highest dose tested (30 mg/kg). In a retention test of longer than normal duration (600 vs 180 s), which resulted in a more favorable comparison value in the control group, an intermediate post-training dose (10 mg/kg) induced a small effect which approached significance; a finding which may account for conflicting reports in the literature concerning the ability of scopolamine to induce a post-training deficit. The pretraining effect does not appear to have been solely the result of state-dependent learning; scopolamine (3 mg/kg) administered before both the training and test sessions induced a deficit of approximately the same magnitude as that found when administered before training or before testing only. The results indicate that scopolamine can induce a small post-trial effect, presumably through an influence on consolidation processes. The much larger effect of pretrial scopolamine, however, indicates a primary influence on processes related to information acquisition. Together with findings from the literature, the present experiments suggest that scopolamine-induced amnesia partially, but not completely, models the memory deficits of human dementia.     

Enhanced inhibitory avoidance learning prevents the long-term memory-impairing effects of cycloheximide,a protein synthesis inhibitor

Interference with activity of numerous cerebral structures produces memory deficiencies; in many instances, however, when animals are over-trained such interference becomes innocuous. Systemic administration of protein synthesis inhibitors impairs long-term retention; this effect has been interpreted to mean that protein synthesis is required for memory consolidation, though little is known about the effect of protein synthesis inhibitors on memory of enhanced learning in the rat. To further analyze the protective effect of enhanced learning against amnesic treatments, groups of Wistar rats were trained in a one-trial step-through inhibitory avoidance task, using different intensities of foot-shock during training. Cycloheximide (CXM; 2.8 mg/kg), an inhibitor of protein synthesis, was injected either 30 min before training or immediately after training. Twenty-four hours after training retention latencies were recorded. Our data showed that both pre- and post-training administration of CXM produced amnesia in those groups that had been trained with relatively low foot-shock intensities, but no impairment in retention was observed when relatively high intensities of foot-shock were administered. These and similar results lead us to conclude that protein synthesis inhibitors may interfere with memory consolidation, but their effect disappears when animals are submitted to an enhanced learning experience, calling into question the idea that protein synthesis is required for memory consolidation.     

Amnesia produced by anisomycin in an appetitive task is not due to conditioned aversion

Two experiments investigated the effects of lithium chloride (LiCl) and anisomycin (ANI) in a water reward Y-maze task. In Experiment 1, male CD-1 mice given weak or strong training were injected post-training with either saline or LiCl (150 mg/kg), which has been reported to produce conditioned aversion in mice. One day after training, both LiCl groups avoided the rewarded arm of the maze and drank less water than saline-injected controls. Two days after training, the strongly trained LiCl mice showed avoidance, while both LiCl groups drank less water. In Experiment 2, weakly trained mice given pre- and post-training ANI (30 mg/kg) were amnesic on the second test day compared to mice that received post-trial saline. However, water consumption was increased on the test day for both groups. LiCl produced a different pattern of results than ANI in this task. On the basis of these results, it is suggested that amnesia produced by ANI is due to impaired memory formation and not to conditioned aversion.     

AF-DX 116, a Presynaptic Muscarinic Receptor Antagonist, Potentiates the Effects of Glucose and Reverses the Effects of Insulin on Memory

Male Swiss mice were tested 24 h after training in a one-trial step-through inhibitory avoidance task. Low subeffective doses of -(+)-glucose (10 mg/kg, ip), but not its stereoisomer -(−)-glucose (30 mg/kg, ip), administered immediately after training, and AF-DX 116 (0.3 mg/kg, ip), a presynaptic muscarinic receptor antagonist, given 10 min after training, interact to improve retention. Insulin (8 IU/kg, ip) impaired retention when injected immediately after training, and the effects were reversed, in a dose-related manner, by AF-DX 116 (0.3, 1.0, or 3.0 mg/kg, ip) administered 10 min following insulin. Since AF-DX 116 possibly blocks autoreceptors mediating the inhibition of acetylcholine release from cholinergic nerve terminals, the present data support the view that changes in the central nervous system glucose availability, subsequent to modification of circulating glucose levels, influence the activity of central cholinergic mechanisms involved in memory storage of an inhibitory avoidance response in mice.     

The effects of calcium channel antagonists on short- and long-term retention in mice using spontaneous alternation behavior

The effect of calcium channel antagonists (CCA's) on working and reference memory in mice was studied using spontaneous alternation (SA) behavior in a T maze. Mice were given either one or four forced trials to either the right or the left arm on the training session (T1) followed by a free choice test (T2) at varying intervals after the initial trial. Untreated animals given one forced trial exhibited significantly greater levels of SA than chance at all delay intervals out to 20 min but not at 30, 60, or 180 min. Animals given four forced trials showed significant levels of SA 24 h after exposure but not at 72 h. Additional groups of mice were treated with amlodipine, nimodipine, diltiazem, and verapamil 1 h before T1. Mice given one forced trial were tested 30, 60, or 180 min after T1 while mice given four forced trials were tested 72 h after T1. Results showed that all of the CCA's except verapamil produced significant SA at the 30-min interval and nimodipine and diltiazem also significantly increased SA at the 60-min-delay interval. No significant effects were observed at the 180-min test. In the four trial groups, all of the CCA's with the exception of verapamil produced significant levels of SA at the 72-h interval. These results indicate that representative CCA's from both the dihydropyridine and the benzothiazapine classes can facilitate both short- and long-interval SA, thereby providing further confirmation that CCA's can enhance memory processing in young animals.     

Scopolamine effects on memory retention in mice: a model of dementia?

Scopolamine-treated normal young human subjects exhibit memory dysfunctions analogous to those observed in demented patients. The dysfunctions are reversible by physostigmine but not by d-amphetamine which suggests that the memory impairment is specifically related to reduced cholinergic transmission caused by scopolamine. Scopolamine-induced amnesia has been proposed as a model for dementia where reduced cholinergic function is the suspected cause. We report seven experiments in young adult mice which examine scopolamine's effects on memory retention and whether its amnestic effects are specifically blocked by cholinergic agonists or cholinomimetics. Young adult mice were trained to avoid footshock in a T maze and their retention tested 1 week after training. Pretraining subcutaneous injection of scopolamine improved retention scores of "undertrained" mice at a dose of 0.01 mg/kg but impaired at a dose of 0.1 mg/kg. Post-training injection showed no effect at 0.01 mg/kg, enhanced retention scores at 0.1 mg/kg, and impaired at 1.0 mg/kg. The impairment by 1.0 mg/kg was blocked by injection 45 min post-training of each of two cholinergic drugs but was also counteracted by six drugs which act upon five other neural systems (catecholamine, serotonin, glycine, GABA, and hormonal). When scopolamine was injected 40 min pretraining, and each of eight drugs was injected immediately after training, the amnestic effect of scopolamine was only partially counteracted. This suggests that scopolamine impaired acquisition, in addition to some impairment of memory processing. This was confirmed by a direct study of acquisition rates of the avoidance response; 0.1 mg/kg of scopolamine impaired acquisition. The overall results indicate that pretraining administration of scopolamine impairs learning and to some degree memory processing. Counteracting scopolamine-induced amnesia, by either pretraining or post-training drug administration, is not specific to the cholinergic system.     

Enhancement of the Post-training Cholinergic Tone Antagonizes the Impairment of Retention Induced by a Nitric Oxide Synthase Inhibitor in Mice

The present experiments examined the role of the central cholinergic system in the memory impairment induced by post-training administration of a nitric oxide synthase (NOS) inhibitor in mice. Male Swiss mice received a one-trial inhibitory avoidance training (0.8 mA, 50 Hz, 1-s footshock) followed immediately by an ip injection of the NOS inhibitor -N^G-nitroarginine methyl ester ( -NAME; 100 mg/kg). Retention (cut-off time, 300 s) was tested 48 h after training. The administration of -NAME results in memory impairment for the inhibitory avoidance task. The effects of -NAME (100 mg/kg, ip) on retention were reversed in a dose-related manner by the centrally acting anticholinesterase physostigmine (35, 70, or 150 μg/kg, sc) administered 30 min after the NOS inhibitor. Further, -NAME (100 mg/kg, ip)-induced memory impairment was completely antagonized by the centrally acting muscarinic cholinergic agonist oxotremorine (OTM; 25, 50, or 100 μg/kg, sc) when given 30 min after -NAME. The peripherally acting anticholinesterase neostigmine (150 μg/kg, sc) did not modify the memory-impairing effects of -NAME. These findings suggest that the memory impairment following post-training administration of a NOS inhibitor is mediated, at least in part, by a reduction of the activity of central muscarinic cholinergic mechanisms and are consistent with our previous view that nitric oxide may be involved in post-training neural processes underlying the storage of newly acquired information.     

Nicotine enhances contextual fear conditioning in C57BL/6J mice at 1 and 7 days post-training

Nicotine has been demonstrated to enhance learning processes. The present experiments extend these results to examine the effects of nicotine on acquisition and consolidation of contextual and cued fear conditioning, and the duration of nicotine's enhancement of conditioned fear. C57BL/6 mice were trained with two pairings of an auditory CS and a foot shock US. Multiple doses of nicotine were given before or immediately after training and on testing day (0.0, 0.050, 0.125, 0.250, and 0.375 mg/kg, i.p). Freezing to both the context and auditory CS was measured 24h after training and again 1 week after training. Mice did not receive nicotine for the 1-week retest. Nicotine (0.125 and 0.250 mg/kg) given on both training and testing days enhanced freezing to the context at 24h. In addition, elevated freezing to the context was seen 1 week post-training in mice previously treated with 0.125 and 0.250 mg/kg nicotine. Thus, nicotine-treated mice did show elevated levels of freezing when retested 1 week later, even though no nicotine was administered at the 1-week retest. Mice that received nicotine on training day or testing day only and mice that received nicotine with mecamylamine, a nicotinic receptor antagonist, were not different from saline-treated mice. In addition, post-training administration of nicotine did not enhance fear conditioning. The present results indicate that nicotine enhancement of contextual fear conditioning depends on administration of nicotine on training and test days but results in a long-lasting enhancement of memories of contextual fear conditioning that remains in the absence of nicotine.     

Amnesic effect of cycloheximide in the mouse mediated by adrenocortical hormones.

The involvement of adrenocortical hormones in the amnesic effect of cycloheximide was examined in mice. Subcutaneous injection of cycloheximide shortly before a training trial of a passive avoidance task resulted in an amnesia of the avoidance response. However, amnesia was absent in the adrenalectomized animals in which cerebral protein synthesis was suppressed by cycloheximide. Injection of corticosteroids antagonized the amnesic effect, most effecively if the steroids were given immediately after training. The influence of the hormonal treatments upon the amnesic effect was not ascribable to a change in general activity level. The amnesic effect of subcutaneously injected cycloheximide appears to be mediated by hormonal deficiency, and not related to suppression of the cerebral protein synthesis.     

The impairment of retention induced by pentylenetetrazol in mice may be mediated by a release of opioid peptides in the brain

Pentylenetetrazol (PTZ, 45 mg/kg, ip) impaired retention of a one-trial step-through inhibitory avoidance task when injected into male Swiss mice 10 min after training, as indicated by retention performance 48 h later. The amnestic effect of PTZ was prevented by naltrexone (0.01 or 0.10 mg/kg, ip) administered after training, but prior to PTZ-treatment. On the contrary, neither naltrexone methyl bromide (0.01, 0.10, or 10.0 mg/kg, ip), a quaternarium analog of naltrexone, nor MR2266 (0.01 or 0.10 mg/kg, ip), a putative kappa opiate receptor antagonist, modified the behavioral effects of PTZ. On the other hand, the body seizures produced by PTZ were unaffected by any of the three opiate receptor antagonists that were given before the convulsant. Taken together, these results suggest that the effects of PTZ on retention are mediated, at least in part, by opioid peptides of central origin, and rules out a possible participation of opioid peptides derived from prodynorphin-precursor molecule. Administration of beta-endorphin (0.01 or 0.10 microgram/kg, ip) 10 min prior to testing attenuate the retrograde amnesia caused by PTZ. The effect of beta-endorphin was prevented by the simultaneous administration of naltrexone (0.10 mg/kg, ip) prior to testing. Naltrexone has no effect of its own upon retrieval. These results suggest that the impairment of retention induced by PTZ is probably due, at least in part, to a release of opioid peptides in the brain during the post-training period. PTZ given after training do not affect consolidation or memory storage, as mice thus treated may retrieve the learned information when they are submitted to an appropriate neurohumoral and/or hormonal state in the test session, that is, beta-endorphin injection. Therefore, the action of PTZ would be primarily at the level of the mechanism that make stored information available for late retrieval.     

Chicks Injected with Antisera to either S-100α or S-100β Protein Develop Amnesia for a Passive Avoidance Task

The cellular expression of S-100β protein is upregulated in Alzheimer's disease and in Down's syndrome, and this protein has been implicated in memory-related processes in laboratory animals. However, the possibility that the α subunit of S-100 is also involved in memory has not yet been examined. In the present study, day-old black Australorp white Leghorn cockerel chicks (Gallus domesticus) received injections of monoclonal antisera to S-100α (1:50) or S-100β (1:500) into each hemisphere immediately after training on a one-trial passive avoidance task. The chicks displayed significantly lower retention levels than control birds that had been injected with antisera to carbonic anhydrase, or with saline (p< .01). S-100α antisera had an amnestic effect when injected between 0 and 20 min after training, with memory deficits occurring from 30 min postlearning, at the point of transition between the A and the B phases of the Gibbs-Ng intermediate memory stage. By contrast, the S-100β antisera needed to be injected either 5 min before or immediately after training and produced amnesia 10 min earlier, at the start of the A phase of the intermediate memory stage. We conclude that the two subunits of the S-100 protein are required at different points in the sequence of events leading to the consolidation of passive avoidance memory.     

Two time windows of anisomycin-induced amnesia for inhibitory avoidance training in rats: protection from amnesia by pretraining but not pre-exposure to the task apparatus

We have studied the effect of training conditions on hippocampal protein synthesis-dependent processes in consolidation of the inhibitory avoidance task. Adult male Wistar rats were trained and tested in a step-down inhibitory avoidance task (0.4 mA foot shock, 24 hr training–test interval). Fifteen minutes before or 0, 3, or 6 hr after training, animals received a 0.8-μl intrahippocampal infusion of the protein-synthesis inhibitor anisomycin (80 μg) or vehicle (PBS, pH 7.4). The infusion of anisomycin impaired retention test performance in animals injected 15 min before and 3 hr after the training session, but not at 0 or 6 h post-training. Pretraining with a low foot shock intensity (0.2 mA) 24 hr before training, prevented the amnestic effect of anisomycin injected at 15 min before or 3 hr after training. However, simple pre-exposure to the inhibitory avoidance apparatus did not alter the amestic effects of anisomycin. The results suggest that hippocampal protein synthesis is critical in two periods, around the time of, and 3 hr after training. A prior weak training session, however, which does not itself alter step-down latencies, is sufficient to prevent the amnestic effect of anisomycin, suggesting that even if not behaviorally detectable, weak training must be sufficient to produce some lasting cellular expression of the experience.