Effects of posttraining treatments in the posterior cingulate cortex on short- and long-term memory for inhibitory avoidance in rats.

Adult male Wistar rats were bilaterally implanted with indwelling cannulae in the caudal region of the posterior cingulate cortex. After recovery, animals were trained in a step-down inhibitory avoidance task (3.0-s, 0.4-mA foot shock) and received, right after training, a 0.5-microl infusion of vehicle (phosphate-buffered saline, pH 7.4), of the GABA(A) receptor agonist muscimol (0.1 or 0.5 microg), of the cAMP-dependent protein kinase (PKA) stimulant Sp-cAMPS (0.1 or 0.5 microg), or of the PKA inhibitor Rp-cAMPS (0.1 or 0.5 microg). Animals were tested twice, 1.5 h and, again, 24 h after training, in order to examine the effects of these agents on short- and long-term memory, respectively. Muscimol (0.5 but not 0.1 microg) hindered retention for both short- and long-term memory (p <.05). Rp-cAMPS (0.1 or 0.5 microg) hindered retention for short-term memory (p <.05). In addition, these animals showed lower, but not significantly lower, latencies than controls in the test session for long-term memory (p >.10). A trend toward an amnesic effect on long-term memory was also observed after Sp-cAMPS infusion at 0.1 microg (p <.10). These results show that strong stimulation of GABAergic synapses in the caudal region of the rat posterior cingulate cortex right after training impairs short- and long-term memory (the latter less dramatically). The same occurs by inhibiting PKA activity with regard to STM and possibly to LTM.

Involvement of the serotonergic type 1A (5-HT1A) receptor in the agranular insular cortex in the consolidation of memory for inhibitory avoidance in rats.

Adult male Wistar rats were bilaterally implanted with indwelling cannulae in the agranular insular cortex of the prefrontal cortex. After recovery, animals were trained in a step-down inhibitory avoidance task (3.0-s, 0.4-mA footshock) and received, immediately after training, a 0.5-microl infusion of the serotonergic type 1A (5-HT1A) receptor agonist dipropylamino-8-hydroxy-1,2,3,4-tetrahydronaphthalene hydrobromide (8-OH-DPAT) or of the 5- HT1A receptor antagonist 1-(2-methoxyphenyl)-4-[4-(2-phthalimido)butyl] piperazine hydrobromide (NAN-190), or of vehicle alone (20% DMSO). Retention testing was carried out 24 h after training. 8-OH-DPAT (1.25 and 6.25 microg but not 0.0125 or 0.125 microg) was amnesic. NAN-190 was not effective at 0.125 or 1.25 microg any dose but reversed amnesia when given at 1.250 microg simultaneously with both effective doses of 8-OH-DPAT. These results show that an overactivation of 5-HT1A receptors in the agranular insular cortex impairs memory consolidation of inhibitory avoidance, in rats, immediately after training. This suggests that these receptors of the insular cortex may modulate memory consolidation.

Training in the step-down inhibitory avoidance task time-dependently increases cAMP-dependent protein kinase activity in the entorhinal cortex.

The cAMP/cAMP-dependent protein kinase (PKA) signaling pathway has been implicated in synaptic plasticity changes and memory consolidation. Several cortical structures are involved in the consolidation of memory for inhibitory avoidance. The aim of the present work was to observe the effects of training in the inhibitory avoidance task on the levels of PKA activity in the entorhinal, parietal and posterior cingulate cortex (EC, PARIET and PC), and the medial precentral area (Fr2) of the rat, at different post-training times (0, 1.5, 3 and 6h). PKA activity, assayed using [gamma-32P]ATP and kemptide, a selective substrate, increased in the EC 3 h after training, but no changes were observed in PARIET, PC and Fr2. These results suggest that the late phase of memory consolidation of inhibitory avoidance requires a functional PKA signaling pathway in the EC in a way that a 'peak' of PKA activity is observed.

Simultaneous modulation of retrieval by dopaminergic D(1), beta-noradrenergic, serotonergic-1A and cholinergic muscarinic receptors in cortical structures of the rat.

Retrieval of inhibitory avoidance has been recently shown to require intact glutamate receptors, protein kinases A and C and mitogen-activated protein kinase in the CA1 region of the rat hippocampus and in the entorhinal, posterior parietal and anterior cingulate cortex. These enzymatic activities are known to be modulated by dopamine D(1), beta-noradrenergic, 5HT1A and cholinergic muscarinic receptors. Here we study the effect on retrieval of this task of well-known agonists and antagonists of these receptors infused in the same brain cortical regions and into the basolateral amygdala, in rats. The drugs used were SKF38393 (D(1) agonist), noradrenaline, 8-HO-DPAT (5HT1A agonist), oxotremorine (muscarinic agonist), SCH23390 (D(1) antagonist), timolol (beta antagonist), NAN-190 (5HT1A antagonist) and scopolamine (muscarinic antagonist). All were studied at two different dose levels. The localised infusion of SKF38393, noradrenaline, NAN-190 and oxotremorine into any of the cortical structures mentioned 10 min prior to a 24-h retention test session of one-trial step-down inhibitory avoidance enhanced retention test performance. SCH2330, timolol, 8-HO-DPAT and scopolamine hindered retention test performance. In the basolateral amygdala only an enhancing effect of noradrenaline and an inhibitory effect of timolol were seen. Three hours after the infusions, retention test performance returned to normal in all cases. None of the treatments affected locomotion or rearing in an open field or behaviour in the elevated plus maze. Therefore, their effects on retention testing can be attributed to an influence on retrieval. In conclusion, memory retrieval of this apparently simple task requires the participation of CA1, entorhinal, posterior parietal and anterior cingulate cortex, and is strongly modulated by, dopaminergic D(1), beta-noradrenergic, muscarinic cholinergic and 5HT1A receptors in the four areas. The first three types of receptor enhance, and the latter inhibits, retrieval. Only beta-adrenoceptors appears to be involved in the modulation of retrieval of this task by the amygdala. The results bear on the well-known influence of emotion and mood on retrieval, and indicate that this involves many areas of the brain simultaneously. In addition, the results point to similarities and differences between the modulatory mechanisms that affect retrieval and those involved in the consolidation of the same task.

The ubiquitin-proteasome cascade is required for mammalian long-term memory formation.

It has been recently demonstrated that ubiquitin-proteasome-mediated proteolysis is required for long-term synaptic facilitation in Aplysia. Here we show that the hippocampal blockade of this proteolytic pathway is also required for the formation of long-term memory in the rat. Bilateral infusion of lactacystin, a specific proteasome inhibitor, to the CA1 region caused full retrograde amnesia for a one-trial inhibitory avoidance learning when given 1, 4 or 7h, but not 10 h, after training. Proteasome inhibitor I produced similar effects. In addition, inhibitory avoidance training resulted in an increased ubiquitination and 26S proteasome proteolytic activity and a decrease in the levels of IkappaB, a substrate of the ubiquitin-proteasome cascade, in hippocampus 4 h after training. Together, these findings indicate that the ubiquitin-proteasome cascade is crucial for the establishment of LTM in the behaving animal.

LY294002, an inhibitor of phosphoinositide 3-kinase given into rat hippocampus impairs acquisition, consolidation and retrieval of memory for one-trial step-down inhibitory avoidance.

Adult male Wistar rats were bilaterally implanted with indwelling cannulae in the CA1 region of the dorsal hippocampus. Once recovered from surgery, animals were submitted to one session of step-down inhibitory avoidance training (3.0 s, 0.4 mA footshock). Animals received a 0.5-microl infusion of saline, or of LY294002 (5, 50 or 500 microM), an inhibitor of the phosphoinositide 3-kinase (PI 3-K) family. Infusions were given 10 min before training, immediately post-training or 10 min prior to a 24-h retention test. In the pre- and post-training groups, the animals were tested twice: at 1.5 and 24 h after training, for short- (STM) and long-term memory (LTM), respectively. Pre- and post-training infusion of the drug inhibited both STM and LTM. Pre-test infusions impaired LTM retrieval. The effects can not be attributed to influences on locomotor, exploratory, pro- or anti-conflict behaviour, since LY294002 had no influence on elevated plus-maze behaviour. The results suggest that hippocampal PI 3-K is necessary for memory acquisition, consolidation and retrieval of the consolidation of step-down inhibitory avoidance in rats. This could be due to an interaction with the N-methyl-d-aspartate (NMDA) receptor complex or with activity of the extracellularly regulated protein kinase (ERK)-Ras signalling pathway.

Molecular signalling pathways in the cerebral cortex are required for retrieval of one-trial avoidance learning in rats.

Rats were implanted bilaterally with cannulae in the CA1 region of the dorsal hippocampus, the entorhinal cortex, anterior cingulate cortex, posterior parietal cortex, or the basolateral complex of the amygdala. The animals were trained in one-trial step-down inhibitory avoidance and tested 24 h later. Prior (10 min) to the retention test, through the cannulae, they received 0.5 microl infusions of a vehicle (2% dimethylsulfoxide in saline), or of the following drugs dissolved in the vehicle: the glutamate NMDA receptor blocker, aminophosphonopentanoic acid (AP5, 2.0 or 5.0 microg), the AMPA receptor blocker, 6,7-dinitroquinoxaline-2,3 (1H,4H)dione (DNQX, 0.4 or 1.0 microg), the metabotropic receptor antagonist, methylcarboxyphenylglycine (MCPG, 0.5 or 2.5 microg), the inhibitor of cAMP-dependent protein kinase (PKA), Rp-cAMPs (0.1 or 0.5 microg), the PKA stimulant, Sp-cAMPs (0.5 microg), or the inhibitor of the mitogen-activated protein kinase (MAPK), PD098059 (10 or 50 microM). All these drugs, at the same doses, had been previously found to alter long-term memory formation of this task. Here, retrieval test performance was blocked by DNQX, MCPG, Rp-cAMPs and PD098059 and enhanced by Sp-cAMPs infused into CA1 or the entorhinal cortex. The drugs had similar effects when infused into the parietal or anterior cingulate cortex, except that in these two areas AP5 also blocked retrieval, and in the cingulate cortex DNQX had no effect. Infusions into the basolateral amygdala were ineffective except for DNQX, which hindered retrieval. None of the treatments that affected retrieval had any influence on performance in an open field or in a plus maze; therefore, their effect on retention testing can not be attributed to an influence on locomotion, exploration or anxiety. The results indicate that the four cortical regions studied participate actively in, and are necessary for, retrieval of the one-trial avoidance task. They require metabotropic and/or NMDA glutamate receptors and PKA and MAPK activity. In contrast, the basolateral amygdala appears to participate only through a maintenance of its regular excitatory transmission mediated by glutamate AMPA receptors.

Involvement of the medial precentral prefrontal cortex in memory consolidation for inhibitory avoidance learning in rats.

Adult male Wistar rats were trained in a step-down inhibitory avoidance learning task (3.0-s, 0.4-mA foot shock), received a 0.5-microl infusion of muscimol (0.02, 0.1, or 0.5 microg), AP5 (0.16, 0.34, 0. 5, 1.6, or 5.0 microg), SCH 23390 (0.05, 0.34, 0.5, or 1.75 microg), saline, or vehicle (DMSO 20%) into the anterior medial precentral area (Fr2) (CI) immediately after training, and were tested 24 h later. Muscimol (0.02, 0.1, or 0.5 microg), AP5 (0.34 or 0.5 microg), or SCH (0.5 or 1.75 microg) were amnesic. Then, animals were infused with muscimol (0.1 or 0.5 microg), AP5 (0.34, 0.5, or 5.0 microg), or SCH (0.5 microg) at other posttraining times and/or into the junction of Fr1-Fr2 (CII). Muscimol (0.1 and 0.5 microg) or SCH into CI were amnesic when given 90 or 180 min after training, but not when given 270 min after training. Muscimol (0.5 microg, but not 0.1 microg) or SCH into CII were amnesic when given 90 min after training, but not when given 0 or 180 min after training. AP5 (0.5, but not 5.0 microg) was amnesic when given into CI, but not into CII, at 0 or 180 min posttraining, and a trend toward an amnesic effect was seen at 90 min posttraining. The results suggest that 1) the glutamatergic, GABAergic, and dopaminergic systems in Fr2 are involved in the consolidation of memory for inhibitory avoidance learning, either directly or as parts of modulatory systems; and 2) timing of involvement of anterior Fr2 (CI) is different from that of posterior Fr2 (CII).

Short- and long-term memory are differentially affected by metabolic inhibitors given into hippocampus and entorhinal cortex.

Rats were implanted with cannulae in the CA1 area of the dorsal hippocampus or in the entorhinal cortex and trained in one-trial step-down inhibitory avoidance. Two retention tests were carried out in each animal, one at 1.5 h to measure short-term memory (STM) and another at 24 h to measure long-term memory (LTM). The purpose of the present study was to screen the effect on STM of various drugs previously shown to affect LTM of this task when given posttraining at the same doses that were used here. The drugs and doses were the guanylyl cyclase inhibitor LY83583 (LY, 2.5 microMg), the inhibitor of Tyr-protein kinase at low concentrations and of protein kinase G (PKG) at higher concentrations lavendustin A (LAV, 0.1 and 0.5 microMg), the PKG inhibitor KT5823 (2.0 microMg), the protein kinase C (PKC) inhibitor staurosporin (STAU, 2.5 microMg), the inhibitor of calcium/ calmodulin protein kinase II (CaMKII) KN62 (3.6 microMg), the protein kinase A (PKA) inhibitor KT5720 (0.5 microMg), and the mitogen-activated protein kinase kinase (MAPKK) inhibitor PD098059 (PD, 0.05 microMg). PD was dissolved in saline; all the other drugs were dissolved in 20% dimethyl sulfoxide. In all cases the drugs affected LTM as had been described in previous papers. The drugs affected STM and LTM differentially depending on the brain structure into which they were infused. STM was inhibited by KT5720, LY, and PD given into CA1 and by STAU and KT5720 given into the entorhinal cortex. PD given into the entorhinal cortex enhanced STM. LTM was inhibited by STAU, KN62, KT5720, KT5823, and LAV (0.5 microMg) given into CA1 and by STAU, KT5720, and PD given into the entorhinal cortex. The results suggest that STM and LTM involve different physiological mechanisms but are to an extent linked. STM appears to require PKA, guanylyl cyclase, and MAPKK activity in CA1 and PKA and PKC activity in the entorhinal cortex; MAPKK seems to play an inhibitory role in STM in the entorhinal cortex. In contrast, LTM appears to require PKA and PKC activity in both structures, guanylyl cyclase, PKG, and CaMKII activity in CA1, and MAPKK activity in the entorhinal cortex.

S100B infusion into the rat hippocampus facilitates memory for the inhibitory avoidance task but not for the open-field habituation.

Adult male Wistar rats were bilaterally implanted with indwelling cannulae in the hippocampus. Forty-eight hours after surgery, animals were habituated to an open-field box during 2 min, being tested 24 h later; next they were trained in a step-down inhibitory avoidance task (3.0 s, 0.4 mA foot-shock), being tested again 24 h later. Immediately after the training session of each task, animals received a 0.5-microl infusion of calcium-phosphate-buffered saline (PBS) and S100B (20, 200, 2000, or 20,000 nM). In the inhibitory avoidance task, animals infused with the two highest concentrations of S100B, 2 and 20 microM, obtained higher scores of retention relative to controls in the test session (p<0.05), and a trend toward an increase was observed in animals infused with 200 nM (p<0. 10). In both sessions of the habituation task, groups were not different regarding crossings, rearings, and time for leaving the first square (p>0.10). These results indicate that, in rats, post-training increased hippocampal levels of S100B right after training facilitate, in a dose-dependent way, long-term memory for an inhibitory avoidance task, but not for an open-field habituation.

Differential effects of post-training muscimol and AP5 infusions into different regions of the cingulate cortex on retention for inhibitory avoidance in rats.

Adult male Wistar rats were bilaterally implanted with indwelling cannulae in four different coordinates of the cingulate cortex: (1) the anterior cingulate (AC), (2) the rostral region of the posterior cingulate (RC), (3) the upper portion of the caudal region of the posterior cingulate (UC), and (4) the lower portion of the caudal region of the posterior cingulate (LC). After recovery, animals were trained in a step-down inhibitory avoidance task (3.0-s, 0.4-mA foot shock). Either immediately, or 90 or 180 min after training, animals received a 0.5-microl infusion of vehicle (phosphate buffer, pH 7.4), of muscimol (0.5 microg), or of AP5 (5.0 microg). Retention testing was carried out 24 h after training. Muscimol was amnestic when given into any of the three coordinates of the posterior cingulate cortex 90 min after training, and when given into LC immediately post-training. In addition, AP5 was amnestic when given into UC 90 min post-training, but not when given into any other region and/or at any other time. None of the treatments had any effect when given into AC. The results suggest that memory processing of the inhibitory avoidance task is regulated by the posterior but not by the anterior cingulate cortex, through muscimol-sensitive synapses, relatively late after training. AP5-sensitive synapses appear to play a very limited role in these processes, restricted to UC.

The amygdala is involved in the modulation of long-term memory, but not in working or short-term memory.

Rats with cannulae implanted in the junction between the central and the basolateral nuclei of the amygdala were trained in one-trial step-down inhibitory avoidance and tested at 3 s for working memory (WM) or 1.5 or 24 h later for short-term memory (STM) and long-term memory (LTM), respectively. Several drugs were infused 6 min prior to training in the animals in which WM was measured or 0 min posttraining in those in which STM and LTM were measured: the glutamate receptor antagonists CNQX (0.5 microg) and AP5 (5.0 microg), the indirect GABA A receptor antagonist picrotoxin (0.08 microg), the cholinergic muscarinic receptor blocker scopolamine (2. 0 microg), norepinephrine (0.3 microg), the protein kinase C inhibitor staurosporin (1.0 microg), or the calcium/calmodulin dependent protein kinase II inhibitor Kn-62 (3.5 ng). None of the drugs had any effect on either WM or STM. All had, as previously shown, strong effects on LTM: picrotoxin and norepinephrine enhanced it, and CNQX, AP5, scopolamine, Kn-62, and staurosporin inhibited it. The results do not support the idea that memory of this task is formed in the amygdala; they indicate that the amygdala is not involved in WM or STM processing and support the idea that the amygdala modulates LTM storage processes carried out elsewhere.

Differential involvement of cortical receptor mechanisms in working, short-term and long-term memory.

Rats received, through bilaterally implanted indwelling cannulae, 0.5 microliter infusions of 6-cyano-7-nitroquinoxaline2,3-dione (CNQX) (0.5 microgram), D-2-amino-5-phophono pentanoic acid (AP5) (5.0 micrograms), muscimol (0.5 microgram), scopolamine (2.0 micrograms), SCH23390 (2.5 micrograms), saline or a vehicle into the CA1 region of the hippocampus, or into the antero-lateral prefrontal (PRE), posterior parietal (PP) and entorhinal cortex (EC). The infusions were given 6 min prior to one-trial step-down inhibitory avoidance training in order to measure their effect on working memory (WM), or immediately post-training in order to measure their effect on short-term (STM) and long-term memory (LTM), 1.5 and 24 h later, respectively. WM was inhibited by CNQX or muscimol given into any of the cortical areas, by SCH23390 given into CA1, PRE or PP, and by scopolamine given into PRE or EC. STM was unaffected by any of the treatments given into PRE, and was inhibited by CNQX or muscimol given into CA1, PP and EC and by scopolamine given into PP, and enhanced by SCH given into CA1. LTM was inhibited by CNQX, muscimol, scopolamine or SCH23390 given into PRE, by scopolamine given into PP, by SCH23390 given into the entorhinal cortex, and by AP5, CNQX, muscimol or scopolamine given into CA1. The results indicate a differential involvement of the various neurotransmitter systems in the three types of memory in the various brain areas, and a separation of the mechanisms and of the regions involved in each. In addition, some of the findings suggested links between WM and LTM processing in PRE, between WM and STM processing in EC and PP, and between all three types of memory in CA1.

Short- and long-term memory are differentially regulated by monoaminergic systems in the rat brain.

Rats with cannulae implanted in the dorsal CA1 region of the hippocampus or in the entorhinal cortex (EC) were trained in one-trial step-down inhibitory avoidance and tested 1.5 or 24 h later, in order to measure short-term memory (STM) and long-term memory (LTM) respectively. Several drugs infused immediately post-training inhibited STM without altering LTM: the D1 receptor agonist SKF38393 (7.5 microgram) given into either CA1 or EC, the beta blocker timolol (0.3 microgram) given into EC, the 5HT1A receptor agonist 8-HO-DPAT (2.5 microgram) given into CA1, and the 5HT1A antagonist NAN-190 (2.5 microgram) given into EC. These findings indicate that STM is not a necessary step toward LTM. Intraentorhinal 8-HO-DPAT enhanced STM and depressed LTM. The D1 antagonist SCH23390 (0.5 microgram) enhanced STM without affecting LTM when given into CA1, and blocked LTM without affecting STM when given into EC. Intraentorhinal norepinephrine (0.3 microgram) enhanced both STM and LTM, and the same drug when given into CA1 enhanced LTM selectively. None of the drugs had any effect on retrieval of either STM or LTM when given prior to testing. The data indicate that STM and LTM are differentially modulated by D1, beta, and 5HT1A receptors in CA1 and EC.