Effects of reversible inactivations of the medial septal area on reference and working memory versions of the Morris water maze

Involvement of the medial septal area (MSA) in reference memory and working memory versions of the Morris water maze (MWM) task was investigated in rats with reversible inactivation of this area by drugs injected through a single cannula aimed at the MSA. In Experiment 1, rats were trained in a reference memory version of the MWM with two blocks of four trials per day for 3 consecutive days. Acquisition was impaired by pretrial MSA injection of 10 ng tetrodotoxin (TTX) in 1 μl saline but not of saline alone into MSA. In Experiment 2, intraseptal injection of TTX (10 ng, 1 μl) immediately after two blocks of four trials had no effect on the consolidation of spatial reference memory. In Experiment 3, intraseptal injection of TTX (10 ng, 1 μl) impaired retrieval of well established spatial reference memory in rats which had received 8 trials per day for 3 consecutive days. In Experiments 4 and 5, rats were trained in a working memory version of MWM task to find a new target position in trial 1 and retrieval of this information was tested 75 min later in trial 2. Intraseptal injection of lidocaine (4%, 1 μl) prior to training impaired working memory performance while immediately posttraining injection of lidocaine had no effect. It is concluded that normal activity of the MSA is necessary for the memory formation at the time of training but its involvement in posttraining consolidation is unlikely. The MSA function is required for retrieval of well established spatial reference memory.     

Reversible lesion of the rat's orbitofrontal cortex interferes with hippocampus-dependent spatial memory

In this study, tetrodotoxin (TTX) inactivation was employed to evaluate the involvement of the rat's orbitofrontal cortex (OFC) in hippocampus-dependent spatial memory using Morris water maze (MWM) and place avoidance learning (PAL) tasks. In Experiment 1, rats trained in MWM task with two blocks of four trials per day for 3 consecutive days received bilateral injections of either TTX or saline into the OFC 60 min before each daily training session. The acquisition of spatial memory was evaluated 24h after the last training day and it was shown an impairment by the TTX. In Experiment 2, bilateral intra-OFC injections of TTX or saline were made immediately after two blocks of four trials. Testing 24h later, it was revealed that TTX also impairs spatial memory consolidation. In Experiments 3 and 4, rats were trained in a single 30-min session to avoid a 60 degrees segment of the stable circular (80-cm diameter) arena, entering which was punished by a mild shock (PAL task) and retention was tested 24h later in a 30-min extinction session. Bilateral injections of TTX or saline were made into the OFC 60 min before training or immediately after training. Again, TTX impaired the place avoidance retention when it was injected into the OFC either before (acquisition phase) or after (consolidation phase) training. These findings indicate that functional integrity of the OFC is necessary for both the acquisition and the consolidation of hippocampus-dependent spatial memory in rats.     

Different effects of scopolamine on the retrieval of spatial memory and fear memory

Retrieval of memory is fundamental for our life as individuals. The participation of cholinergic system in memory consolidation process has been extensively studied, but there are few data concerning the function of this system in memory retrieval process. In the current study, we inject non-selective muscarinic antagonist scopolamine peripherally 20 min before training or testing to see whether cholinergic modulation has effects on the acquisition or retrieval of spatial memory by water maze task and fear memory by inhibitory avoidance task. We find that the cholinergic system is essential for the acquisition of both spatial memory and fear memory. As for the memory retrieval, the cholinergic system has a positive role in the retrieval of spatial memory, because mice injected with scopolamine 20 min before the testing in the water maze show impaired spatial memory retrieval. Whereas injection of scopolamine 20 min before the testing in the inhibitory avoidance task does not cause memory retrieval deficits. That indicates the cholinergic system is not essential for the retrieval of fear memory.     

Effects of mammillary body lesions on spatial reference and working memory tasks.

This work examines the effects of electrolytic mammillary body (MB) lesions on the performance of rats in different spatial memory tasks in the Morris water maze. The first experiment assessed the effect of MB lesion on performance in a spatial reference memory task (place learning with multiple trials). The second experiment examined the effect of a lesion in this nucleus on performance in a spatial working memory task (single-trial place learning). The results show that lesion of the MB impairs the animals performance in spatial working memory tasks but does not impair acquisition in spatial reference memory tasks (place learning, transfer task, reversal task) or in a visual-cued task. However, the deficit in the spatial working memory task does not appear to vary with the delay between acquisition and retention trials (30 s and 5 min). Our results demonstrate a clear role of the mammillary bodies in the processing of spatial information in a working memory task. Lesion of the MB impairs performance in a working memory task but does not affect reference memory processes.     

Effects of excitotoxic median raphe lesions on scopolamine-induced working memory deficits in inhibitory avoidance

The aim of the present study was to investigate the effects of excitotoxic damage of the serotonergic cell bodies in the median raphe nucleus (MRN) on the scopolamine-induced working memory deficits in a single-trial light/dark inhibitory avoidance task. Rats were given 1 mg/kg of scopolamine hydrobromide (intraperitonal, i.p.) or saline before the inhibitory avoidance training, in which initial preference to the dark compartment (escape latency) was used to measure nonmnemonic behaviors, and response latency to enter the dark compartment immediately after the shock was used to measure working memory. It was found that scopolamine significantly reduced escape latencies in sham-lesioned rats, whereas it had no effect in the rats with MRN lesions. Although MRN lesion per se did not alter response latency, it prevented scopolamine-induced decrease in this parameter. These results suggest that the antagonistic interactive processes between serotonergic projections of the MRN and the muscarinic cholinergic system modulate nonmnemonic attentional component of working memory formation in the inhibitory avoidance.     

Effect of reversible inactivation of reuniens nucleus on memory processing in passive avoidance task

The reuniens nucleus (RE) is the largest nucleus of the midline thalamic nuclei (MLN). RE has strongly connections with the amygdala and hippocampus, the structures that are involved in the learning and memory processes. In our previous report we have shown the role of RE in the spatial learning and memory using Morris water maze (MWM) task. Since RE is connected to multiple limbic structures, its involvement in the emotional learning and memory is a possibility. The present study was designed to elucidate the role of RE in acquisition, consolidation, and retrieval on the passive avoidance (PA) task which depends on a distributed network including the thalamus, amygdala, medial prefrontal cortex (mPFC) and hippocampus. For this purpose, rats were chronically implanted with a cannula aimed at the RE through which 0.5 μl tetracaine (2%) or saline were injected. Rats were trained in a PA task and their retention test was performed 24 h later. The injection of saline or tetracaine was applied 5 min before or 5, 90, and 360 min after the acquisition trial and 5 min before the retention tests. Our findings showed that inactivation of RE before training did not affect acquisition, but affected memory retention 24 h later in PA task. Moreover, inactivation of RE only 5 min after training impaired consolidation but not after 90 or 360 min. Also, inactivation of the RE, 5 min before the retrieval test impaired memory retrieval in PA task. In conclusion, it seems that RE is involved in memory processes in rats.     

Intra-hippocampal microinjections of anisomycin did not block glucocorticoid-induced impairment of memory retrieval in rats: an evidence for non-genomic effects of glucocorticoids

Previous findings indicated that intra-cerebral as well as systemic injections of glucocorticoid hormones shortly before retention testing induce impairment of long-term memory retrieval, but the underlying mechanisms (genomic or non-genomic) are not known. To clarify this issue, we investigated the involvement of protein synthesis in the hippocampus in glucocorticoid-induced impairment of memory retrieval. Young rats carrying bilateral cannulae aimed at the hippocampus were trained in a water maze (MWM) task with six trials per day for six consecutive days. Retention of the spatial training was assessed 24h after the last training session with a 60-s probe trial. Corticosterone (1mg/kg) was injected 30 min before retention testing with or without prior bilateral intra-hippocampal injections of anisomycin (80 or 160 microg/mul per site) as an inhibitor of protein synthesis. The control animals received vehicle or saline solutions. The results show that corticosterone-induced impairment of memory retrieval was not blocked by intra-hippocampal infusions of anisomycin. Our data suggest that corticosterone influences memory retrieval via a protein independent mechanism and provide evidence for rapid actions (non-genomic) of glucocorticoids on memory retrieval.     

EFFECTS OF EXCITOTOXIC MEDIAN RAPHE LESIONS ON SCOPOLAMINE-INDUCED WORKING MEMORY DEFICITS IN INHIBITORY AVOIDANCE

The aim of the present study was to investigate the effects of excitotoxic damage of the serotonergic cell bodies in the median raphe nucleus (MRN) on the scopolamine-induced working memory deficits in a single-trial light/dark inhibitory avoidance task. Rats were given 1 mg/kg of scopolamine hydrobromide (intraperitonal, IP) or saline before the inhibitory avoidance training, in which initial preference to the dark compartment (escape latency) was used to measure nonmnemonic behaviors, and response latency to enter the dark compartment immediately after the shock was used to measure working memory. It was found that scopolamine significantly reduced escape latencies in sham-lesioned rats, whereas it had no effect in the rats with MRN lesions. Although MRN lesion per se did not alter response latency, it prevented scopolamine-induced decrease in this parameter. These results suggest that the antagonistic interactive processes between serotonergic projections of the MRN and the muscarinic cholinergic system modulate nonmnemonic attentional component of working memory formation in the inhibitory avoidance.     

Functional inactivation of orexin 1 receptors in CA1 region impairs acquisition, consolidation and retrieval in Morris water maze task

Orexin containing neurons in the lateral hypothalamic area (LHA) produce orexin-A (hypocretin-1) and orexin-B (hypocretin-2) and send their axons to the hippocampus, which predominantly expresses orexin 1 receptors (OX1Rs) showing a higher affinity to orexin-A. Recent studies have shown that central administration of orexin-A has an effect on learning and memory but literature concerning the role of orexinergic system in cognition remains controversial. Therefore, we examined the effect of pre-training, post-training and pre-probe trial intrahippocampal CA1 administration of a selective OX1R the orexin 1 receptor antagonist SB-334867-A (1.5, 3, 6 microg/0.5 microl) on acquisition, consolidation and retrieval in a single-day testing version of Morris water maze (MWM) task. Our results show that, SB-334867-A impaired acquisition, consolidation and retrieval of MWM task as compared with the control group. This drug had no effect on escape latency of a non-spatial visual discrimination task. Therefore, it seems that endogenous orexins, especially orexin-A, play an important role in spatial learning and memory in the rat.     

The effects of NMDA-induced retrohippocampal lesions on performance of four spatial memory tasks known to be sensitive to hippocampal damage in the rat

Four separate cohorts of rats were employed to examine the effects of cytotoxic retrohippocampal lesions in four spatial memory tasks which are known to be sensitive to direct hippocampal damage and/or fornix-fimbria lesions in the rat. Selective retrohippocampal lesions were made by means of multiple intracerebral infusions of NMDA centred on the entorhinal cortex bilaterally. Cell damage typically extended from the lateral entorhinal area to the distal ventral subiculum. Experiment 1 demonstrated that retrohippocampal lesions spared the acquisition of a reference memory task in the Morris water maze, in which the animals learned to escape from the water by swimming to a submerged platform in a fixed location. In the subsequent transfer test, when the escape platform was removed, rats with retrohippocampal lesions tended to spend less time searching in the appropriate quadrant compared to controls. Experiment 2 demonstrated that the lesions also spared the acquisition of a working memory version of the water maze task in which the location of the escape platform was varied between days. In experiment 3, both reference and working memory were assessed using an eight-arm radial maze in which the same four arms were constantly baited between trials. In the initial acquisition, reference memory but not working memory was affected by the lesions. During subsequent reversal learning in which previously baited arms were now no longer baited and vice versa, lesioned animals made significantly more reference memory errors as well as working memory errors. In experiment 4, spatial working memory was assessed in a delayed matching-to-position task conducted in a two-lever operant chamber. There was no evidence for any impairment in rats with retrohippocampal lesions in this task. The present study demonstrated that unlike direct hippocampal damage, retrohippocampal cell loss did not lead to a general impairment in spatial learning, implying that the integrity of the retrohippocampus and/or its interconnection with the hippocampal formation is not critical for normal hippocampal-dependent spatial learning and memory. This outcome is surprising for a number of current hippocampal theories, and suggests that other cortical as well as subcortical inputs to the hippocampus might be of more importance, and further raises the question regarding the functional significance of the retrohippocampal region. Introduction     

Effects of excitotoxic median raphe lesion on working memory deficits produced by the dorsal hippocampal muscarinic receptor blockade in the inhibitory avoidance in rats

The experiments investigated the interactions between median raphe nucleus (MRN) serotonergic and septo-hippocampal muscarinic cholinergic systems in the modulation of forming and storing performances of working memory. Rats with ibotenic acid-induced MRN-lesion bilaterally received scopolamine (2-4 microg/each side) infusion into the dentate gyrus of the dorsal hippocampus and were tested in a single trial step-through inhibitory avoidance. Initial preference to the dark compartment (escape latency) was taken as the measure of non-mnemonic behaviours and response latency to enter the dark compartment immediately after the foot-shock was used to measure working memory. The high-dose scopolamine infusion 10 min before the training decreased escape latencies in the sham-lesioned rats, whereas had no effect in the MRN-lesioned rats. Although MRN lesion per se did not alter response latency, it alleviated pre-training scopolamine-induced decrease, but aggravated post-training scopolamine-induced reduction in this parameter. These results suggest that the antagonistic interactive processes between MRN-serotonergic and hippocampal cholinergic systems modulate non-mnemonic component of working memory formation, whereas the storing performance of working memory is modulated by the synergistic interactions between these systems in the hippocampus, mainly in the dentate gyrus.     

The locus coeruleus involves in consolidation and memory retrieval, but not in acquisition of inhibitory avoidance learning task

The locus coeruleus (LC) located at the level of the pons, is involved in cognitive functions such as learning and memory. The bilateral lidocaine-induced reversible inactivation of this nucleus has been considered in order to study its role in the phases of memory processing (acquisition, consolidation and retention) without any interference with the function of the same structure either during earlier and/or later phases of the same process. In this study, inhibitory avoidance (IA) learning task used to find the LC function in acquisition, consolidation and retrieval. Saline or lidocaine 4% (0.5 microl/side) microinjected into the LC, for assessing the acquisition (5 min before training), consolidation (5, 90 and 360 min after training) and memory retrieval, 5 min before testing. The retention test was done 24h after learning. Our results indicated that: (1) The bilateral functional inactivation of LC before training did not affect acquisition, but affected subsequent memory retention 24h later in IA task. (2) The lidocaine-induced inactivation of LC only 5 min after training impaired consolidation but did not affect it after 90 or 360 min. (3) Inactivation of the LC, 5 min before pre-retrieval test, impaired memory retrieval in IA task. In conclusion, it seems that the nucleus locus coeruleus does not affect acquisition while it involves in the memory consolidation and retrieval of inhibitory avoidance learning task.     

Raphe cells grafted into the hippocampus can ameliorate spatial memory deficits in rats with combined serotonergic/cholinergic deficiencies

The ability of embryonic raphe cells grafted into the hippocampus to restore spatial learning ability was tested in rats with combined serotonergic/cholinergic deficits. Embryonic raphe cells (E14) were transplanted into the hippocampus of serotonin-depleted rats. Two to 3 months after transplantation, control, lesioned and grafted rats were tested in a spatial memory task (a water maze) with and without the addition of atropine. All 3 groups could negotiate the water maze equally well, in non-drug conditions. The injection of atropine caused a severe disruption of performance only in the serotonin depleted rats. The presence of an active serotonergic graft was examined in the intact rat hippocampus using the serotonin releasing drug fenfluramine (FFA). A pronounced depression of hippocampal EEG was observed in control and grafted but not in lesioned rats 15 min after the injection of FFA. These results suggest the involvement of serotonin in cognitive functions in the rat. Furthermore, it is suggested that an interaction between serotonergic and cholinergic neurotransmission occurs in the hippocampus.     

High levels of estradiol disrupt conditioned place preference learning, stimulus response learning and reference memory but have limited effects on working memory.

The present study investigated the effects of high levels of estradiol in female rats on four different radial arm maze tasks: the hippocampus-dependent spatial working-reference memory task; the prefrontal cortex-hippocampus dependent delayed win-shift task; the striatum-dependent cued win-stay task; and the amygdala-dependent conditioned place preference task. Ovariectomized female rats were injected daily with either 10 microg of estradiol benzoate or sesame oil vehicle approximately 4 h prior to testing. In Experiment 1, treatment with estradiol disrupted learning on the spatial working-reference memory task by increasing the number of reference memory errors to reach criterion. In Experiment 2, treatment with estradiol had no significant effect on the delayed win-shift task. In Experiment 3, treatment with estradiol resulted in impaired performance on a striatum-dependent cued win-stay task. In Experiment 4, treatment with estradiol impaired the acquisition of a conditioned place-preference task. Taken together these findings suggest that high levels of estradiol inhibit reference memory, stimulus response learning, and amygdala-dependent appetitive conditioning while having little effect on working memory.     

Exposure to 3,4-methylenedioxymethamphetamine (MDMA) on postnatal days 11-20 induces reference but not working memory deficits in the Morris water maze in rats: implications of prior learning.

3,4-methylenedioxymethamphetamine (MDMA) in previous experiments has been shown to induce long-term spatial and sequential learning and memory deficits in adult offspring after exposure to the drug on postnatal (P) days 11-20, but not after exposure on P1-10. Herein we further tested for the effects of MDMA (0, 5, 10 or 20 mg/kg x 2/day) after exposure on P11-20 on reference and working memory in the Morris water maze (MWM), on reference memory in the Barnes maze, and on cued learning in the visible platform version of the MWM. The MWM and Barnes mazes were counterbalanced such that half the litters received the MWM-first and the other half received the Barnes maze first. Effects on MWM performance as a function of test order were observed. For animals that received the Barnes maze first, spatial MWM learning and memory trends were seen but they were not significantly different between MDMA groups and saline controls. For those receiving the MWM-first, there are consistent impairments on all measures in the MDMA groups compared to controls on MWM performance (latency, path length, and cumulative distance from the goal). On probe trials, MDMA animals receiving the MWM-first showed increased distance from the target site compared to controls. There were no MDMA effects seen on cued trials in the MWM or on straight channel swimming trials regardless of test order, indicating that MDMA had no effects on swimming ability or on the skills needed to learn the MWM. Similarly, there were no effects of MDMA on MWM working memory regardless of test order. No MDMA effects on the Barnes maze were found regardless of test order, however, the interpretation of this finding was compromised by the poor performance of the animals on this task.     

Effect of reversible inactivation of the supramammillary nucleus on spatial learning and memory in rats

Memory includes processes such as acquisition, consolidation and retrieval. Reference memory (RM) and working memory (WM) are two kinds of memory that can be assessed in rodents using spatial tasks, especially using the Morris water maze. The Morris water maze is particularly sensitive to hippocampal lesions. The supramammillary nucleus (SuM) has strong links with the hippocampus and septum. The role of the SuM on spatial learning is controversial. In the present study, involvement of SuM in the different steps of spatial RM and WM was investigated in the Morris water maze using reversible inactivation of SuM with lidocaine. Lidocaine (0.5 microl, 4%) was injected into the SuM through a guide cannula implanted above the SuM. The rats were trained on RM and WM versions of the Morris water maze. SuM was inactivated before training or immediately after training or before the probe trial of retrieval tests. Reversible inactivation of the SuM impaired consolidation of RM, and of consolidation and retrieval of WM. Therefore, it seems that activity of SuM neurons plays a role in spatial RM and WM learning and memory in the rat.     

Differential participation of the NBM in the acquisition and retrieval of conditioned taste aversion and Morris water maze

Deficits in both learning and memory after lesions of the cholinergic basal forebrain, in particular the nucleus basalis magnocellularis (NBM), have been widely reported. However, the participation of the cholinergic system in either acquisition or retrieval of memory process is still unclear. In this study, we tested the possibility that excitotoxic lesions of the NBM affect either acquisition or retrieval of two tasks. In the first experiment, animals were trained for two conditioned taste aversion tasks using different flavors, saccharine and saline. The acquisition of the first task was before NBM lesions (to test retrieval) and the acquisition of the second task was after the lesions (to test acquisition). Accordingly, in the first part of the second experiment, animals were trained in the Morris water maze (MWM), lesioned and finally tested. In the final part of this experiment, another set of animals was lesioned, then trained in the MWM and finally tested. All animals were able to retrieve conditioned taste aversion (CTA) and MWM when learned before NBM lesions; however, lesions disrupted the acquisition of CTA and MWM. The results suggest that the NBM and cholinergic system may play an important role in acquisition but not during retrieval of aversive memories.     

Hippocampal CA3-region is crucial for acquisition and memory consolidation in Morris water maze task in mice

This experiment investigated the involvement of the dorsal hippocampal CA3-region in the different phases of learning and memory in spatial and non-spatial tasks. To do so, we temporarily inactivated the CA3-subfield by a focal injection of diethyldithiocarbamate (DDC) which chelates most of the heavy metals present in this region. The effects of temporary inactivation of the CA3-region were examined in an associative task, the Morris water maze (MWM). To study the different phase of memory we used a new behavioural massed-procedure founded on four massed training sessions in the spatial and the non-spatial (cue) version of this task. In the spatial version, we showed that a bilateral injection of DDC into the CA3-region impairs the acquisition but not the recall of spatial information. The main result of this study is that the same injection performed immediately after the training session also perturbed memory consolidation. In the cue version of the MWM, we found no difference between the DDC-injected mice and their controls in acquisition or memory consolidation of non-spatial information. These results suggest that the hippocampal CA3-region is essential for spatial memory processes and specifically in memory consolidation of spatial information.     

Effects of pentylenetetrazol-induced brief convulsive seizures on spatial memory and fear memory

Previous studies have demonstrated that in the pentylenetetrazol (PTZ) kindling model, recurrent seizures either impair or have no effect on learning and memory. However, the effects of brief seizures on learning and memory remain unknown. Here, we found that a single injection of a convulsive dose of PTZ (50 mg/kg, ip) induced brief seizures in Sprague–Dawley rats. Administration of PTZ before training impaired the acquisition of spatial memory in the Morris water maze (MWM) and fear memory in contextual fear conditioning. However, the administration of PTZ immediately after training did not affect memory consolidation in either task. These findings suggest that brief seizures have different effects on acquisition and consolidation of spatial and fear memory.     

The effect of intrahippocampal injection of testosterone enanthate (an androgen receptor agonist) and anisomycin (protein synthesis inhibitor) on spatial learning and memory in adult, male rats

In most mammals, the hippocampus has a well-documented role in spatial memory acquisition. High concentration of androgen receptors in fundamental centers of learning and memory in brain such as hippocampus shows that there may be some relationships between androgen receptors and cognitive aspects of brain. Previous studies, which have shown sex-dependent differences in hippocampal morphology and physiology, suggest a modulatory role for sex steroids in hippocampal function. Androgens have been shown to modulate some hippocampal-mediated behaviors including learning and memory. To study the mechanism of action of androgens in processes underlying learning and memory, anisomycin, a protein synthesis inhibitor was used to prevent the genomic effects of testosterone. Therefore, the effects of anisomycin and testosterone together were assessed on rat's performance in MWM. Rats received anisomycin (2.5 microg/0.5 microl), testosterone (80 microg/0.5 microl) or both anisomycin (2.5 microg/0.5 microl) and testosterone (80 microg/0.5 microl) through the connulas in the CA1 region. Anisomycin was injected 20 min and testosterone was injected 35 min before training each day. The results showed that anisomycin (2.5 microg/0.5 microl) and testosterone (80 microg/0.5 microl) increased latencies to find the invisible platform. But the group that received testosterone and anisomycin together was decrease in latency and traveled distance to find the invisible platform.