4.1 LTP underlies memory function
Synaptic plasticity is the ability of a synapse to continuously adapt thorough time according to the flow of neuronal activity by the strengthening or weakening of synapses.
Santiago Ramon y Cajal proposed in 1894, the idea that learning is preceded by strengthening of synapses by enhancement of communication efficiency (Cajal, 1894).
Following Cajal proposal, in 1949 Donald Hebb formalized this idea and formally postulated: “When an axon of cell A is near enough to excite a cell B and repeatedly or persistently takes part in firing it, some growth process or metabolic change takes place in one or both cells such that A’s efficiency, as one of the cells firing B, is increased” (Hebb, 1949). Learning and memory-induced changes in synaptic strength could thus follow a coincidence in activity between the pre- and post-synaptic cells.
Two main forms of long-term synaptic plasticity are described in the CNS: long-term potentiation (LTP) and long-term depression (LTD) (Figure 5).
Figure 5. NMDAr-dependent long-term potentiation (LTP) and depression (LTD) in the hippocampus, from Lüscher and Malenka (2012).
A. Representative drawing from Ramon y Cajal (1909) of the trisynaptic pathway in the hippocampus.
dependent LTP and LTD are induced at CA3-CA1 pyramidal cells synapses (blue on red). NMDAr-independent LTP is induced at DG-CA3 synapses (green on blue). B. Representative electron microscopy image of two asymmetric CA3-CA1 synapses (scale bar 200 nm). C. Example traces of bidirectional change
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in CA3-CA1 synaptic efficacy by LTD and LTP induction in the same synapses monitored by extracellular field recordings in acute slice preparation of the hippocampus. D. Proposed mechanisms for LTP and LTD expression. Modest activation of NMDAr via calcium influx mediated by weak presynaptic activity preferentially activates phosphatases and dephosphorylate AMPAr and thus promote their endocytosis and LTD is observed. Conversely, strong presynaptic activity paired with strong postsynaptic depolarization triggers LTP mainly via CAMKII, receptor phosphorylation and exocytosis.
In 1966, Tim Bliss and Terje Lømo reported for the first time that long-term synaptic efficacy changes in hippocampus of the rabbit could be induced by high-frequency electrical stimulations (known also as tetanic stimulations) (Bliss and Lomo, 1973; Lynch, 2004; Luscher and Malenka, 2012; Nicoll, 2017). The basic mechanisms of LTP involve a simultaneous pre-synaptic excitatory input and intracellular post-synaptic depolarization, which allow calcium influx through N-methyl-D-aspartate receptors (NMDAr) that activate intracellular signaling cascades able to adapt synaptic efficacy (Figure 5 D). This is the case of LTP at EC-DG and CA3-CA1 hippocampal synapses where LTP induction is dependent of NMDAr activation and trigger post-synaptic modifications (see chapter below). Activation of Calcium/calmodulin-dependent kinase II (CamKII) lead to phosphorylation of AMPAr and their insertion at post-synaptic membranes (Ehlers, 2000).
AMPAr insertion mediates increased postsynaptic-evoked currents (Figure 5 A-D).
Studies have reported synapses containing only NMDAr and no AMPAr, called silent synapses that are activated by LTP protocol and trigger AMPAr insertion (Isaac et al., 1995; Liao et al., 1995). In contrast to LTP in CA1, LTP at DG-CA3 synapses is known to be mediated by a pre-synaptic mechanism. DG-CA3 LTP is mediated by an increase in the release of neurotransmitter (Zalutsky and Nicoll, 1990; Staubli et al., 1990; Xiang et al., 1994). These studies revealed an increase in the release probability of neurotransmitters at CA3 synapses after LTP induction, mainly mediated by increase in pre-synaptic cAMP and required normal PKA function (Huang et al., 1994; Lopez-Garcia et al., 1996). Another key element found to mediate this form of pre-synaptic LTP is the Ca2+/ calmodulin-activated adenylyl cyclase 1 (AC1) (Villacres et al., 1998). LTP at mossy fibers can be observed in conditions where NMDAr are dispensable (Harris and Cotman, 1986) but studies have shown that at mossy fibers synapses NMDAr are present and do participate to AMPAr responses (Weisskopf and Nicoll, 1995).
The demonstration that synaptic potentiation is linked to memory encoding has proved to be of great complexity and continues to be tested nowadays. Morris carried out one of the first attempts with this aim in 1986 (Morris et al., 1986). In this study, Morris and colleagues reported that chronic intraventricular infusion of NMDAr blockers (D, L-AP5) impaired specifically spatial learning in rats performing Morris water maze task (MWM) but had no major effects on other behavioral tasks (visual discrimination or sensorimotor
activity). They confirmed that in vivo application of AP5 suppressed LTP induction, providing the first correlative evidence of the link between LTP and learning processes.
Short after, Kentros et al. (1998) showed that NMDAr blockade during spatial exploration of a novel environment prevented the long-term stabilization of CA1 place fields, while short-term stability was not affected, suggesting the involvement of a form of LTP in the formation and maintenance of brain spatial representations (Kentros et al., 1998).
Moser et al. in 1998 exploited the fact that electrically induced LTP saturates synaptic efficacy to test the role of synaptic plasticity in memory (Moser et al., 1998). By electrically potentiating connections in the rats’ hippocampus, they found that consequent spatial learning was impaired. In this same line, Gruart et al. (2006), recorded neuronal activity at the CA3-CA1 synapses during an associative learning task (eyeblink conditioning) (Gruart et al., 2006). They observed that acquisition and/or extinction of the conditioned eyeblink was parallel to changes in CA3-CA1 synaptic strength (potentiation or decrease). Evoked-LTP impaired the acquisition of the CRs as well as learning induced increase in fEPSPs, suggesting a saturation of CA3-CA1 connectivity.
Whitlock et al. in 2006, showed that rats trained in the inhibitory avoidance tasks (IA), displayed in vivo CA1 synaptic changes in glutamate receptors and increases in fEPSP as it happens by evoked-LTP induction (Whitlock et al., 2006). A confirmation came with the observation that induction of LTP was occluded after IA learning.
In 2014, Nabavi et al. showed that it is possible to artificially create an episodic-like memory in mice by induction of LTP and LTD (Nabavi et al., 2014). In their study, they combined optogenetic induction of LTP and/or LTD to interfere with auditory fear memory formation. They revealed that LTP seems to underlie neuronal ensembles (see chapter 6) coding for the associated memory and suggest that LTD induction can be used to inactivate the formed memory by disassembling these neuronal populations (see chapter 6).
4.2 Synaptic plasticity modifies gene expression
While the early phase of LTP depends on rapidly regulated kinase activity and AMPAR trafficking (Huang, 1998; Malenka and Bear, 2004; Luscher and Malenka, 2012), the late phase of LTP (between 1 and 8 h after induction, L-LTP) require de novo protein synthesis. Trafficking of proteins and mRNA from the soma to distal dendritic sites and to the activated synapses provides a pool of new synaptic proteins necessary for stabilization of L-LTP (Kelleher et al., 2004). Immediate Early Genes (IEGs), rapidly and transiently transcribed in response to neuronal activity (Guzowski et al., 2005; Bramham et al., 2008)
play a major role in L-LTP. Their activation does not require protein synthesis as it is the case for most of the genes (Herrera and Robertson, 1996), allowing very rapid transcription. Ca2+ influx through NMDAr and L-type VDCC (Berridge, 1998; Flavell and Greenberg, 2008) trigger diverse kinase signaling cascades that converge on transcription factors within the nucleus and ultimately control the expression of neuronal activity-regulated genes (Figure 6).
Figure 6. Network of activity-dependent induction of gene expression, adapted from Flavell and Greenberg (2008).
Cellular calcium influx either from L-type voltage-gated calcium channel (L-VGCC) or from ionotropic glutamate NMDA receptor trigger a wide range of intracellular signaling cascades. These cascades can lead to activation of pre-existing activity-regulated transcription factors in the nucleus. These activity-regulated transcription factors, when are dephosphorylated can trigger induction of activity-regulated transcription factors and synaptic proteins regulators. Genetic mutations in the genes that encode several of these signaling molecules give rise to neurological disorders in humans (yellow boxes). Only a subset of the signaling pathways that mediate activity- dependent transcription are shown here.
IEGs are divided in two main categories: regulatory transcription factors (RTFs) and effector factors (EFs). The former regulates the expression of other genes, while the latter directly control specific cell’s functions. Among the first class, special attention has been given to the RTF IEG c-fos as it is involved in neuronal plasticity mechanisms required for long-term memory consolidation processes (Grimm et al., 1997; Lamprecht and Dudai, 1996; Morrow et al., 1999; Tischmeyer and Grimm, 1999). Since its early discovery, c-fos has been used as a marker for neuronal activity (Sagar et al., 1988; Garner et al., 2012).
c-fos is tightly regulated by increases in neuronal activity in many brain regions (Kovacs,
Figure 3.
Signal transduction networks mediating neuronal activity-dependent gene expression. Calcium influx through either neurotransmitter receptors or voltage-gated calcium channels leads to the activation of many calcium-regulated signaling enzymes, which sets in motion several signal transduction cascades. These pathways converge on preexisting transcription factors in the nucleus and lead to their activation through direct posttranslational protein modifications.
Several of the activity-regulated genes encode transcriptional regulators, which in turn promote the transcription of additional activity-regulated genes. Many other activity-regulated genes encode proteins that function in dendrites or at synapses and thereby coordinate activity-dependent dendritic and synaptic remodeling within the neuron. Genetic mutations in the genes that encode several of these signaling molecules give rise to neurological disorders in humans ( yellow boxes). Only a subset of the signaling pathways that mediate activity-dependent transcription are shown here.
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2008). c-fos gene regulation is very rapid and has a transient temporal dynamic. For example, following a stimulating event (i.e. memory formation), c-fos mRNA expression levels have a peak of around 30 minutes, while its protein expression is known to peak around 90-120 minutes (Guzowski, 2002; Zangenehpour and Chaudhuri, 2002). Different studies reported that c-fos expression highly correlated to natural (Dragunow and Faull, 1989; Rakhade et al., 2007) or optogenetic induced neuronal firing (Schoenenberger et al., 2009). Also, synaptic NDMA activation trigger c-fos mRNA expression in hippocampal neurons (Cole et al., 1989). More attention has been paid on the role of c-fos in the hippocampus showing its involvement in the formation of long-term memories (Countryman et al., 2005; He et al., 2002; Katche et al., 2010).
Effector IEGs (non-RTF IEGs) have diverse cellular functions including functional and structural synapse (Vazdarjanova et al., 2002; Shepherd et al., 2006). Arc/Arg3.1 mRNA and protein levels are tightly regulated by different signals (BDNF, mGluR1 receptors, muscarinic acetylcholine receptors and N-methyl-D-aspartate receptors (NMDAr) (Ying et al., 2002; Waung et al., 2008; Teber et al., 2004; Steward and Worley, 2001).
4.3 Structural plasticity during leaning
Neuronal network continuously undergoes structural rearrangements, also known as structural plasticity, through different mechanisms including the formation, stabilization and elimination of synapses (Holtmaat and Svoboda, 2009). How structural plasticity relate to memory? There is correlational evidence showing emergence of new synapses formation and spine growth following learning-induced synaptic potentiation (Zito et al., 2009) (Nagerl et al., 2007), but there is no clear consensus and evidence of how structural plasticity underlie memory retrieval and consolidation. However, a recent study from Hayashi-Takagi et al. has been able to causally link synapse formation and memory (Hayashi-Takagi et al., 2015). They developed a novel approach to label and manipulate motor learning formation of synapses in the cortex of mice. By in vivo imaging, they first reveal that motor learning induces synapse remodeling of a small subset of cells and that these cells presented potentiation. Second, they showed that selective elimination (shrinkage) of learning-induced synapses disrupted motor learning without affecting new learning. These results first demonstrated that newly formed synapses induced by motor learning are necessary for memory formation and retention (Figure 7).
Figure 7. Structural plasticity reflects learning, adapted from Caroni et al. (2012).
Schematics represent excitatory dendrites (grey lines) and their synapses (dots). New formed synapses are labelled in green, lost synapses in red and stable synapses in blue. Arrows indicate the gain and loss of synapses. Learning induces structural synapses rearrangement on dendrites of principal excitatory cells by increasing their turnover. Consolidation result in the stabilization of learning-induced synapses.
4.4 System consolidation: a model for the long-term storage of memories
Memories are not immediately formed. They are initially labile to be then consolidated and permanently stored in the brain (Frankland and Bontempi, 2005). The first theoretical model for system consolidation was proposed by David Marr in 1970 (Marr, 1970) and subsequently refined by Squire and Alvarez (Squire and Alvarez, 1995). Information is firstly and temporarily retained by the hippocampus before being transferred to cortical areas where is then reorganized and stored for long periods of time. This idea posed the basis for the development of more contemporary models. Since the hippocampus plays a temporary function for storing information, then it should have strong interactions with the cortex where memories can be permanently reposed. Blockade of communication between the HPC and the cortex should impair memory consolidation. This is exactly what Remondes and Shuman have demonstrated in their study (Remondes and Schuman, 2004). Lesions of the temporoammonic input from the layer III of the EC to the CA1 allowed normal spatial memory formation (MWM task, 24 hours’ test) but disrupted its consolidation as confirmed by impairment of memory recall 4 weeks later. Many other studies have confirmed that prefrontal cortex alterations interfere mostly with remote episodic memories but not with recent ones (Maviel et al., 2004; Zola-Morgan and Squire, 1990; Frankland et al., 2004). Imaging studies revealed that spatial memories activate cortical regions in mice (Maviel et al., 2004; Bontempi et al., 1999). Taken together these findings suggest that remote memories are distributed in cortical networks with particular focus on the prefrontal cortex (PFC).
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