Induction and activity-dependent reversal of persistent LTP and LTD in lateral perforant path synapses in vivo.

The reversibility of long-term potentiation (LTP) and heterosynaptic long-term depression (LTD) lasting weeks was examined in the lateral perforant path of freely moving adult Sprague-Dawley rats. LTP lasting weeks was rapidly reversed within minutes by high-frequency heterosynaptic stimulation of the medial perforant path, in an N-methyl-D-aspartate receptor-dependent manner. LTP reversal also occurred, albeit more slowly and to a lesser extent, when animals were given 1-3 weeks of overnight exposure to an enriched environment (EE). LTD likewise was reversed upon repeated EE exposure. A covert similarity between the degrees of LTP and LTD reversal was revealed when the small potentiation effect of EE treatment by itself on lateral path responses was taken into account. Despite its ability to reverse previously acquired synaptic plasticity, two weeks of EE treatment had no effect on animals' retention of the platform location in a spatial watermaze task, although it did facilitate new learning. These data are in agreement with the hypothesis that hippocampal synapses retain the capacity for rapid synaptic change even when otherwise relatively stable plasticity has previously been induced. Slow reversal of such plasticity did not correlate with a loss of memory retention, possibly because either slow changes permit reorganization of representations such that both old and new information can be accommodated, or else the new information is synaptically represented in orthogonal fashion to the old information.

Long-term regulation of N-methyl-D-aspartate receptor subunits and associated synaptic proteins following hippocampal synaptic plasticity.

Synaptic plasticity in the dentate gyrus is dependent on activation of the N-methyl-D-aspartate (NMDA)-subtype of glutamate receptors. In this study, we show that synaptic plasticity in turn regulates NMDA receptors, since subunits of the NMDA receptor complex are bidirectionally and independently regulated in the dentate gyrus following activation of perforant synapses in awake animals. Low-frequency stimulation that produced a mild synaptic depression resulted in a decrease in the NMDA receptor subunits NR1 and NR2B 48 h following stimulation. High-frequency stimulation that produced long-term potentiation resulted in an increase in NR1 and NR2B at the same time point. Further investigations revealed that in contrast to NR2B, NR1 levels increased gradually after long-term potentiation induction, reaching a peak level at 48 h, and were insensitive to the competitive NMDA receptor antagonist 3-3(2-carboxypiperazin-4-yl) propyl-1-phosphate. The increased levels of NR1 and NR2B at 48 h were found associated with synaptic membranes and with increased NMDA receptor-associated proteins, postsynaptic density protein 95, neuronal nitric oxide synthase and Ca(2+)/calmodulin-dependent protein kinase II, alpha subunit. These data suggest that the persistence of long-term potentiation is associated with an increase in the number of NMDA receptor complexes, which may be indicative of an increase in synaptic contact area.

Heterosynaptic metaplasticity in the hippocampus in vivo: a BCM-like modifiable threshold for LTP.

The homeostatic maintenance of the "modification threshold" for inducing long-term potentiation (LTP) is a fundamental feature of the Bienenstock, Cooper, and Munro (BCM) model of synaptic plasticity. In the present study, two key features of the modification threshold, its heterosynaptic expression and its regulation by postsynaptic neural activity, were tested experimentally in the dentate gyrus of awake, freely moving rats. Conditioning stimulation ranging from 10 to 1,440 brief 400-Hz trains, when applied to medial perforant path afferents, raised the threshold for LTP induction heterosynaptically in the neighboring lateral perforant path synapses. This effect recovered slowly over a 7- to 35-day period. The same conditioning paradigms, however, did not affect the reversal of long-term depression. The inhibition of LTP by medial-path conditioning stimulation was N-methyl-D-aspartate (NMDA) receptor-dependent, but antidromic stimulation of the granule cells could also inhibit lateral path LTP induction, independently of NMDA receptor activation. Increased calcium buffering is a potential mechanism underlying the altered LTP threshold, but the levels of two important calcium-binding proteins did not increase after conditioning stimulation, nor was de novo protein synthesis required for generating the threshold shift. These data confirm, in an in vivo model, two key postulates of the BCM model regarding the LTP threshold. They also provide further evidence for the broad sensitivity of synaptic plasticity mechanisms to the history of prior activity, i.e., metaplasticity.

Sequential increase in Egr-1 and AP-1 DNA binding activity in the dentate gyrus following the induction of long-term potentiation.

Establishment of long-term potentiation (LTP) at perforant path synapses is highly correlated with increased expression of Egr and AP-1 transcription factors in rat dentate gyrus granule cells. We have investigated whether increased transcription factor levels are reflected in increased transcription factor activity by assessing Egr and AP-1 DNA binding activity using gel shift assays. LTP produced an increase in binding to the Egr element, which was NMDA receptor-dependent and correlated closely with our previously reported increase in Egr-1 (zif/268) protein levels. Supershift analysis confirmed involvement of Egr-1, but not Egr-2 in the DNA binding activity. AP-1 DNA binding was also rapidly elevated in parallel with protein levels, however, the peak increase in activity was delayed until 4 h, a time point when we have previously shown that only jun-D protein was elevated. These data indicate that binding of Egr-1 and AP-1 to their response elements is increased in two phases. This may result in activation of distinct banks of target genes which contribute to the establishment of persistent LTP.

Immediate early gene transcription and synaptic modulation.

Long-term changes in gene expression appear to be critical to the formation of memory, but little is known about its stimulus- transcription coupling. Numerous studies in the last decade, by focusing on unraveling this signal transduction pathway, have investigated the potential role of the immediate-early genes in this process. The krox family of immediate-early gene proteins are of particular interest because they may be involved in stabilizing the synaptic modifications that underlie hippocampal long-term potentiation (LTP). A potential upstream mediator of krox induction is cyclic AMP-responsive element binding protein (CREB), a posttranslationally activated transcription factor that has been implicated in numerous memory paradigms. In this study we investigated whether the activation of CREB by phosphorylation may have a role in the development of rat perforant- path-stimulated LTP and associated dentate granule cell krox-24 mRNA expression. Contrary to what was expected, we failed to show any difference in the levels of phosphorylated CREB after LTP or following endogenous synaptic facilitation stimulated by novelty. Using these same model systems we also investigated the protein levels of brain- derived neurotrophic factor (BDNF), another immediate-early gene that is induced following a durable form of LTP. However, BDNF protein was not induced within the hippocampus after LTP and was transiently decreased following novel environmental stimulation.

A double dissociation within the hippocampus of dopamine D1/D5 receptor and beta-adrenergic receptor contributions to the persistence of long-term potentiation.

We compared the effects of the D1/D5 receptor antagonist SCH-23390 with the beta-adrenergic receptor antagonist propranolol on the persistence of long-term potentiation in the CA1 and dentate gyrus subregions of the hippocampus. In slices, SCH-23390 but not propranolol reduced the persistence of long-term potentiation in area CA1 without affecting its induction. The drugs exerted reverse effects in the dentate gyrus, although in this case the induction of long-term potentiation was also affected by propranolol. The lack of effect of SCH-23390 on the induction and maintenance of long-term potentiation in the dentate gyrus was confirmed in awake animals. The drug also had little or no effect on the expression of inducible transcription factors. In area CA1 of awake animals, SCH-23390 blocked persistence of long-term potentiation beyond 3 h, confirming the results in slices. To rule out a differential release of catecholamines induced by our stimulation protocols between brain areas, we compared the effects of the D1/D5 agonist SKF-38393 with the beta-adrenergic agonist isoproterenol on the persistence of a weakly induced, decremental long-term potentiation in CA1 slices. SKF-38393 but not isoproterenol promoted greater persistence of long-term potentiation over a 2-h period. In contrast, isoproterenol but not SKF-38392 facilitated the induction of long-term potentiation. These data demonstrate that there is a double dissociation of the catecholamine modulation of long-term potentiation between CA1 and the dentate gyrus, suggesting that long-term potentiation in these brain areas may be differentially consolidated according to the animal's behavioural state.

Biphasic changes in the levels of N-methyl-D-aspartate receptor-2 subunits correlate with the induction and persistence of long-term potentiation.

N-Methyl-D-aspartate glutamate receptors (NMDAR) form ion channels made up of polypeptides from two classes of subunits; NR1 is obligatory for function whereas members of the NR2 class regulate the properties of the channel. Long-term potentiation (LTP) of synaptic transmission is an event largely dependent on NMDAR activation, and is studied as the primary cellular model of memory in the mammalian brain. While there has been a focus on non-NMDARs in mediating the expression of LTP, we report here biochemical evidence for plasticity of the NMDAR that is associated with LTP persistence in awake animals. Following the establishment of LTP in perforant path synapses of the dentate gyrus, we observed a rise in NR2B protein levels 48 h post-tetanus which was dependent upon activation of NMDARs during the tetanization, and which strongly correlated with the degree of LTP measured at this time-point. We also observed a transient increase in both NR2B and NR2A protein levels 20 min post-tetanus that returned to control levels by 4 h. These early increases were not observed in anaesthetized animals which do not sustain persistent LTP. Our data demonstrate a marked plasticity of NMDAR subunit expression, which may affect LTP persistence, as well as the subsequent ability to induce LTP at previously activated synapses.

Synaptic activity-dependent modulation of mitochondrial gene expression in the rat hippocampus.

In order to identify genes that may underlie the maintenance of long-term potentiation (LTP) at perforant path synapses, complementary DNA libraries were synthesised from dentate gyrus total RNA extracts prepared 48 h after the induction of LTP and from control dentate gyrus extracts. Through differential screening of the LTP library we have identified the mitochondrial 12S rRNA (mt12SrRNA) as a transcript that was elevated at this late time. Northern blot analyses showed that the elevation in mt12SrRNA expression began around 8 h and persisted for at least 2 weeks post-tetanus. We then examined the expression patterns of other mitochondrially-encoded genes and demonstrated a similar elevation in their expression. mt12SrRNA levels were also elevated in other hippocampal regions, including areas CA3 and CA1 and were elevated following low-frequency stimulation or in the presence of an N-methyl-D-aspartate receptor antagonist where induction of LTP was precluded. Taken together, these observations suggest that a long-lasting up-regulation of energy production may be triggered by synaptic activity and this activity need not be of sufficient strength to induce LTP, but may be related to the induction of a metaplastic state.

TrkB expression in dentate granule cells is associated with a late phase of long-term potentiation.

Recent studies have demonstrated that the neurotrophins nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) are induced in hippocampal neurons following the induction of long-term potentiation (LTP), a model of memory, and that BDNF and NT-3 (but not NGF) can induce LTP-like increases in synaptic efficacy. Receptors for these neurotrophins have been cloned and characterized and we investigated whether LTP alters the expression of two neurotrophin receptors, trkB (BDNF receptor) and trkC (NT-3 receptor) in dentate granule neurons of the hippocampus using in situ hybridization analysis. Results show that trkB is strongly induced in these neurons in an N-methyl-D-aspartate (NMDA) receptor-dependent manner. Moreover, the induction of trkB and trkC mRNAs was attenuated by sodium pentobarbital, which interferes with the durability of LTP. Low-frequency stimulation of the perforant path had no effect on trkB mRNA levels but significantly reduced trkC mRNA in dentate granule cells. Thus, both BDNF and its receptor trkB are induced in granule cells by stimulation that produces durable LTP, suggesting that this neurotrophin and its receptor play an important role in memory formation and may be suitable targets for the development of cognitive-enhancing drugs in the treatment of diseases, such as Alzheimer's.

Low-frequency stimulation does not readily cause long-term depression or depotentiation in the dentate gyrus of awake rats.

The ability of low-frequency stimulation (LFS) to induce either long-term depression (LTD) or depotentiation was assessed for perforant path synapses in the dentate gyrus of awake, adult rats. Neither LFS at 1 Hz (100 or 900 pulses) nor LFS at 3 Hz (900 pulses) was sufficient to produce either LTD or depotentiation. LFS at 3 Hz did produce a transient response depression of previously potentiated synapses, but this lasted less than 24 h and was secondary to seizure-like afterdischarges. We conclude that the LFS protocols so effective at eliciting LTD and depotentiation in area CA1 are ineffective for perforant path synapses in the dentate gyrus.

Analysis of the decremental nature of LTP in the dentate gyrus.

The persistence of long-term potentiation (LTP) in the dentate gyrus was compared for two tetanization protocols: 50 trains on one day, or 50 trains on 5 consecutive days. LTP induction was significantly greater in the 250 train condition, but the LTP decay rate over weeks was similar between conditions. The decay of LTP could not be accounted for by deterioration of the preparation. Successive days of stimulation caused repetitive induction of immediate early genes, but did not prolong LTP, suggesting that either the effects of gene expression on LTP stabilization had saturated, or that these genes play other roles in synaptic plasticity.