**4.2 LTD in Hippocampus**

Unlike LTP, LTD in hippocampus occurs when the postsynaptic cells are weakly depolarized, whereas LTP induction involves strong postsynaptic depolarization. The hippocampal LTD is governed by BCM (Bienenstock Cooper Munro) theory (Bienenstock et al., 1982). It says that the synapses that are active when the postsynaptic cells are weakly polarized undergo LTD. If APs preceed EPSPs, LTD results; i.e., unpaired stimulation causes lower calcium signals and therefore LTD.

In the CA1 region LTD is homosynaptic, and depends on NMDARs and on protein synthesis but in the DG, it is independent of NMDARs and protein synthesis and is found both as heterosynaptic and homosynaptic forms (Kemp and Vaughan, 2007).

The best understood type of LTD is induced in hippocampal area CA1 by LFS via an NMDAR dependent rise in postsynaptic intracellular calcium and the activation of a protein phosphatase cascade which will be discussed hereforth. A brief application of NMDA can also lead to depression, i.e., a form of chem-LTD. LTD is triggered by postsynaptic calcium entry, like LTP, after activation by presynaptic stimulus. The main receptors involved are AMPAR and NMDAR. If the postsynaptic depolarization by AMPAR is weak, it cannot activate NMDARs completely. The partial removal of Mg2+ block results in reduced Ca2+ entry.

LTD is an activity dependent reduction in the efficacy of neuronal synapses. It can generally last for hours or longer. It brings about a long lasting decrease in synaptic strength. LTD can be defined as a long lasting decrease in the synaptic response of neurons to stimulation of their afferents following a long patterned stimulus (Collingrigde et al., 2010). LTD is generally considered as a reversal of LTP as it is understood that if synapses continue to increase in strength, eventually they would reach some level of maximum efficacy which might cause saturation and then they may be unable to encode new information. This would result in neurons coming to a stage of complete inactivity or over activity. LTD is also considered to be the initial step in synaptic elimination (Bastrikova et al., 2008; Beckner et

al., 2008) as it is known that those synapses which lose their efficacy are eliminated.

maintenance mechanism is not fully understood (Bliss and Cooke, 2011).

LTD can be either homosynaptic or heterosynaptic. Homosynaptic LTD is induced by a conditioning input. It is input specific. It is restricted to the individual synapse which is activated by a low frequency stimulus (LFS) i.e., it happens in the same synapse that receives the induction. It is associative and it correlates with postsynaptic activation of the neuron by an active presynaptic neuron. Homosynaptic LTD is in turn of two types. LTD which follows an LTP is often known as depotentiation. If LTD is observed from base line conditions, with low frequency stimulus, then it is *de novo* LTD. Heterosynaptic LTD refers to depression at synapses neighboring the activated ones but are not directly activated themselves (Abraham et al., 2007). Heterosynaptic LTD occurs at synapses that are not potentiated. It occurs consequent to a non-conditioning input in association with either LTP or LTD. LTD relies on both pre and postsynaptic expression mechanisms although the

Unlike LTP, LTD in hippocampus occurs when the postsynaptic cells are weakly depolarized, whereas LTP induction involves strong postsynaptic depolarization. The hippocampal LTD is governed by BCM (Bienenstock Cooper Munro) theory (Bienenstock et al., 1982). It says that the synapses that are active when the postsynaptic cells are weakly polarized undergo LTD. If APs preceed EPSPs, LTD results; i.e., unpaired stimulation causes

In the CA1 region LTD is homosynaptic, and depends on NMDARs and on protein synthesis but in the DG, it is independent of NMDARs and protein synthesis and is found

The best understood type of LTD is induced in hippocampal area CA1 by LFS via an NMDAR dependent rise in postsynaptic intracellular calcium and the activation of a protein phosphatase cascade which will be discussed hereforth. A brief application of NMDA can also lead to depression, i.e., a form of chem-LTD. LTD is triggered by postsynaptic calcium entry, like LTP, after activation by presynaptic stimulus. The main receptors involved are AMPAR and NMDAR. If the postsynaptic depolarization by AMPAR is weak, it cannot activate NMDARs completely. The partial removal of Mg2+ block results in reduced Ca2+ entry.

both as heterosynaptic and homosynaptic forms (Kemp and Vaughan, 2007).

**4. Long term depression** 

**4.1 Types of LTD** 

**4.2 LTD in Hippocampus** 

lower calcium signals and therefore LTD.

Therefore instead of kinases, phosphatases get activated as they require comparatively lower Ca2+ concentrations for activation. The protein phosphatase activated is PP2B or calcineurin. PP2B can in turn activate PP1. PP2B dephosphorylates and inactivates Inhibitor-1. This relieves the inhibition of PP1 by Inhibitor-1 thereby activating it (Mulkey et al., 1993).

In hippocampus, plasticity is mediated by conductance changes of AMPARs which are in turn regulated by phosphorylation. The majority of AMPARs at hippocampal synapses are GluR1/GluR2 and GluR2/GluR3 heteromers. The trafficking of GluR1 plays a dominant role in plasticity. Activated PP1 brings about dephosphorylation of GluR1 at Ser845 (Fig. 6) and promotes AMPAR internalization (Lee et al., 2000). Inhibition of PP2B blocks GluR1 internalization and thereby LTD, suggesting the importance of phosphatase activity in LTD (Beattie et al., 2000). Targeting PP1 precisely to synapses upon NMDAR activation is crucial for LTD expression and is facilitated by PP1 binding proteins like spinophilin, neurabin, etc. (Morshita et al., 2001). In hippocampus, AMPARs are stabilized on the membrane by NSF and clathrin adaptor protein AP2, which bind to the NSF binding site on GluR2. During NMDAR-LTD, AP2 replaces NSF and this initiates AMPAR endocytosis. Clathrin mediated endocytosis of AMPARs is triggered by a neuronal calcium sensor known as hippocalcin. Upon activation, hippocalcin translocates to the plasma membrane, where it forms a complex with AP2 and GluR2 and initiates clathrin mediated AMPAR endocytosis. Protein interacting with C-kinase 1 (PICK1) is another protein that binds directly to GluR2 and it can also bind to PKC. PICK1 competes with AMPAR binding protein (ABP) and glutamate receptor interacting protein (GRIP) for binding to C-terminal of GluR2 and promotes internalization. PICK1 also helps in modifying neuronal architecture by interacting with F-actin. PP2B interacts with A-kinase anchor protein-150 (AKAP-150) which in turn interacts with PSD-95. PSD-95 further interacts with NMDAR thereby positioning PP2B near NMDAR (Bhattacharya et al., 2009). This helps in the activation of PP2B by Ca2+ influx through NMDARs. Activated PP2B can mediate the NMDAR-induced endocytosis of AMPARs that underlies one major form of LTD. Disruption of the interaction between PSD-95 and AKAP-150 strongly inhibited NMDAR-dependent endocytosis of AMPARs (Bhattacharya et al., 2009). Phosphorylation of Ser295 of PSD-95 occurs *in vivo*, and it enhances the ability of PSD-95 to accumulate in the PSD, to recruit surface AMPA receptors, and to strengthen synaptic transmission. During LTD, PSD-95 is dephosphorylated at Ser295 facilitating its removal from PSD. This mechanism also plays a role in the NMDAR-dependent endocytosis of AMPAR (Kim et al., 2007).

Although a major form of LTD is mediated by NMDARs, the ultimate direction of change in synaptic efficacy is brought about by changes in AMPAR function (Collingridge et al., 2010). Calcium influx through the NMDAR is central to the induction of both LTP and LTD because intracellular application of calcium chelators, such as BAPTA or EGTA, prevents induction of plasticity. Since induction of LTP and LTD are controlled by the postsynaptic NMDAR, any presynaptic component of expression requires a retrograde messenger that can signal to the presynaptic terminal that coincidence has occurred. Two candidates are nitric oxide (NO) and endocannabinoids (eCB) (Bliss & Cook, 2011). *N*arachidonylethanolamine (AEA) and 2-arachidonoylglycerol (2-AG) are two major eCBs that activate type I cannabinoid receptors (CB1) receptors on the presynaptic neuron in the brain (Di Marzo et al., 1998). Upon stimulation, eCBs are released from postsynaptic neurons and travel across the synaptic cleft to activate CB1 on presynaptic terminals, resulting in depression of synaptic transmission.

Molecular Mechanisms in Synaptic Plasticity 313

The events described above depict the importance of GluN2B subunit in LTD. It is also known that CaMKII can phosphorylate GluN2B at Ser1303, both *in vitro* and *in vivo* (Omkumar et al., 1996). This phosphorylation prevents the binding of CaMKII to GluN2B *in vitro* (Strack et al., 2000; O'Leary et al., 2011). Studies from our lab have shown that the phosphorylation status at Ser1303 enables GluN2B to distinguish between the Ca2+/CaM activated form and autonomously active Thr286-autophosphorylated form of CaMKII. This highlights the need for a dephosphorylation mechanism at GluN2B-Ser1303. It has been shown that phosphatases in PSD can dephosphorylate GluN2B-Ser1303 (Rajeevkumar et al., 2009). Although the physiological role of GluN2B-Ser1303 is not known to date, it is likely to be involved in LTD mechanism as phosphatases get activated during induction of LTD.

Another major form of LTD worth mentioning is the one which is dependent on group 1 metabotropic glutamate receptors (mGluR). Chemical LTD is typically induced by activation of mGlu receptors. The most commonly induced chemical LTD is by (*S*)-3, 5 dihydroxyphenylglycine (DHPG), an agonist of mGluR, one which is effective even in the absence of Ca2+. Normally mGluR-LTD is induced in CA1 synapses by a train of LFS consisting of single pulses. The mGluR antagonist α-methyl-4-caboxyphenylglycine (MCPG) blocked depotentiation and *de novo* LTD in CA1 showing the involvement of mGluR in LTD

Glutamate binding to mGluR initiates a signaling cascade, involving the breakdown of the membrane lipid PIP2 (Phosphoinositol 4, 5 - bisphosphate) by phospholipase C (PLC) to the important signaling molecules IP3 (Inositol 1, 4, 5 - triphosphate). This also causes release of diacylglycerol (DAG) and calcium mobilization. This leads to the activation of the calcium sensitive kinase, PKC. This enzyme then phosphorylates AMPAR but in such a manner that the conductance is reduced (Bliss et al., 2011). An offshoot is the production of NO, the retrograde messenger. Group I mGluRs (mGluR1/5) activate PLC, leading to Ca2+ mobilization, and activation of the ERK–MAPK pathway through which they modulate signals of synapse-to-nucleus communication and triggers protein synthesis. mGluR1 and mGluR2 receptor subtypes mediate *de novo* LTD at cerebellar PF-PC synapses and

mGluRs are the critical regulators of activity-dependent protein synthesis in dendrites. Signaling by mGluR1/5 is critical to synaptic circuitry formation during development and is implicated in LTD (Zukin et al., 2009). mGluR1/5 elicit synapse specific modifications in synaptic strength and spine morphology by stimulating rapid local translation of dendritic mRNAs including *Fmr1*, that encodes fragile X mental retardation protein (FMRP) (Greenough et al., 2001). Expression of mGluR-LTD at Schaffer collateral to CA1 pyramidal cell synapses is mediated by persistent internalization of AMPARs and in adolescent mice requires *de novo*  protein synthesis (Huber et al., 2000; Snyder et al., 2001). But mGluR-LTD at cortical synapses (Desai et al., 2006) requires neither local protein synthesis nor FMRP function. mGluR-LTD is also seen in CA1 in neonates and is protein synthesis dependent. (Nosyreva & Huber, 2005). PICK1 is also required for mGluR-LTD at different synapses. mGluRs are associated with protein tyrosine phosphatases rather than Ser/Thr ones. DHPG, a potent agonist of mGluR, induced LTD that involves tyrosine dephosphorylation of GluA2 and is associated with AMPARs endocytosis (Gladding et al., 2009). DHPG-induced LTD appears not to require

(Bolshakow et al., 1994).

hippocampal mossy fibre synapses respectively.

Fig. 6. Mechanisms of LTD induction in hippocampus and cerebellum. Key signaling pathways that lead to LTD in hippocampus and in cerebellum involve AMPAR regulation. Hippocampal LTD involves activation of phosphatases like PP2B and PP1 which dephosphorylate GluR1 resulting in reduced AMPAR conductance, whereas in cerebellum, kinases like CaMKII and PKC are activated resulting in GluR2 phosphorylation and thereby causing AMPAR endocytosis and reduced current.

Of the NMDARs, GluN2B containing NMDARs are supposed to be important for LTD especially in hippocampus as a study using conditional knockout mice showed that the selective ablation of GluN2B subunits in pyramidal neurons in CA1 specifically impairs CA1 NMDAR-LTD. This also results in deficits in several hippocampal dependent learning and memory tasks, providing strong evidence for a key role of this particular from of LTD in memory formation. (Brigman et al., 2010; Collingridge et al., 2010). BDNF is released from glutamatergic neurons in response to high frequency stimulus and is found to have a role in LTP. While BDNF affects synaptic potentiation at hippocampal synapses, proBDNF is involved in LTD. proBDNF, by activating its receptor known as the p75 neurotrophin receptor (p75NTR ), facilitates hippocampal (LTD). Deletion of p75NTR-/- in mice selectively impaired the NMDAR dependent LTD, without affecting other forms of synaptic plasticity. p75NTR-/- mice also showed a decrease in the expression of GluN2B, an NMDA receptor subunit uniquely involved in LTD. p75NTR-/- mice showed a decrease in the expression of GluN2B in the hippocampus and also a marked reduction in GluN2B-mediated currents at the CA1 synapse. Activation of p75NTR by proBDNF enhanced GluN2B dependent LTD and GluN2B mediated synaptic currents. (Woo et al., 2005).

Fig. 6. Mechanisms of LTD induction in hippocampus and cerebellum. Key signaling pathways that lead to LTD in hippocampus and in cerebellum involve AMPAR regulation.

dephosphorylate GluR1 resulting in reduced AMPAR conductance, whereas in cerebellum, kinases like CaMKII and PKC are activated resulting in GluR2 phosphorylation and thereby

Of the NMDARs, GluN2B containing NMDARs are supposed to be important for LTD especially in hippocampus as a study using conditional knockout mice showed that the selective ablation of GluN2B subunits in pyramidal neurons in CA1 specifically impairs CA1 NMDAR-LTD. This also results in deficits in several hippocampal dependent learning and memory tasks, providing strong evidence for a key role of this particular from of LTD in memory formation. (Brigman et al., 2010; Collingridge et al., 2010). BDNF is released from glutamatergic neurons in response to high frequency stimulus and is found to have a role in LTP. While BDNF affects synaptic potentiation at hippocampal synapses, proBDNF is involved in LTD. proBDNF, by activating its receptor known as the p75 neurotrophin receptor (p75NTR ), facilitates hippocampal (LTD). Deletion of p75NTR-/- in mice selectively impaired the NMDAR dependent LTD, without affecting other forms of synaptic plasticity. p75NTR-/- mice also showed a decrease in the expression of GluN2B, an NMDA receptor subunit uniquely involved in LTD. p75NTR-/- mice showed a decrease in the expression of GluN2B in the hippocampus and also a marked reduction in GluN2B-mediated currents at the CA1 synapse. Activation of p75NTR by proBDNF enhanced GluN2B dependent LTD and

Hippocampal LTD involves activation of phosphatases like PP2B and PP1 which

causing AMPAR endocytosis and reduced current.

GluN2B mediated synaptic currents. (Woo et al., 2005).

The events described above depict the importance of GluN2B subunit in LTD. It is also known that CaMKII can phosphorylate GluN2B at Ser1303, both *in vitro* and *in vivo* (Omkumar et al., 1996). This phosphorylation prevents the binding of CaMKII to GluN2B *in vitro* (Strack et al., 2000; O'Leary et al., 2011). Studies from our lab have shown that the phosphorylation status at Ser1303 enables GluN2B to distinguish between the Ca2+/CaM activated form and autonomously active Thr286-autophosphorylated form of CaMKII. This highlights the need for a dephosphorylation mechanism at GluN2B-Ser1303. It has been shown that phosphatases in PSD can dephosphorylate GluN2B-Ser1303 (Rajeevkumar et al., 2009). Although the physiological role of GluN2B-Ser1303 is not known to date, it is likely to be involved in LTD mechanism as phosphatases get activated during induction of LTD.

Another major form of LTD worth mentioning is the one which is dependent on group 1 metabotropic glutamate receptors (mGluR). Chemical LTD is typically induced by activation of mGlu receptors. The most commonly induced chemical LTD is by (*S*)-3, 5 dihydroxyphenylglycine (DHPG), an agonist of mGluR, one which is effective even in the absence of Ca2+. Normally mGluR-LTD is induced in CA1 synapses by a train of LFS consisting of single pulses. The mGluR antagonist α-methyl-4-caboxyphenylglycine (MCPG) blocked depotentiation and *de novo* LTD in CA1 showing the involvement of mGluR in LTD (Bolshakow et al., 1994).

Glutamate binding to mGluR initiates a signaling cascade, involving the breakdown of the membrane lipid PIP2 (Phosphoinositol 4, 5 - bisphosphate) by phospholipase C (PLC) to the important signaling molecules IP3 (Inositol 1, 4, 5 - triphosphate). This also causes release of diacylglycerol (DAG) and calcium mobilization. This leads to the activation of the calcium sensitive kinase, PKC. This enzyme then phosphorylates AMPAR but in such a manner that the conductance is reduced (Bliss et al., 2011). An offshoot is the production of NO, the retrograde messenger. Group I mGluRs (mGluR1/5) activate PLC, leading to Ca2+ mobilization, and activation of the ERK–MAPK pathway through which they modulate signals of synapse-to-nucleus communication and triggers protein synthesis. mGluR1 and mGluR2 receptor subtypes mediate *de novo* LTD at cerebellar PF-PC synapses and hippocampal mossy fibre synapses respectively.

mGluRs are the critical regulators of activity-dependent protein synthesis in dendrites. Signaling by mGluR1/5 is critical to synaptic circuitry formation during development and is implicated in LTD (Zukin et al., 2009). mGluR1/5 elicit synapse specific modifications in synaptic strength and spine morphology by stimulating rapid local translation of dendritic mRNAs including *Fmr1*, that encodes fragile X mental retardation protein (FMRP) (Greenough et al., 2001). Expression of mGluR-LTD at Schaffer collateral to CA1 pyramidal cell synapses is mediated by persistent internalization of AMPARs and in adolescent mice requires *de novo*  protein synthesis (Huber et al., 2000; Snyder et al., 2001). But mGluR-LTD at cortical synapses (Desai et al., 2006) requires neither local protein synthesis nor FMRP function. mGluR-LTD is also seen in CA1 in neonates and is protein synthesis dependent. (Nosyreva & Huber, 2005).

PICK1 is also required for mGluR-LTD at different synapses. mGluRs are associated with protein tyrosine phosphatases rather than Ser/Thr ones. DHPG, a potent agonist of mGluR, induced LTD that involves tyrosine dephosphorylation of GluA2 and is associated with AMPARs endocytosis (Gladding et al., 2009). DHPG-induced LTD appears not to require

Molecular Mechanisms in Synaptic Plasticity 315

LTD can be induced by prolonged periods of LFS by pairing baseline synaptic stimulation with depolarization i.e. normally at a frequency of 10 Hz. LFS of 1 Hz for 15 minutes brings about LTD in CA1 area of hippocampus of anaesthetized rabbit (Bliss & Lomo, 1973). This stimulation is preceded by baseline stimulation at frequencies such as 0.1–0.05 Hz to establish a reference level for basal synaptic transmission. Electrically induced LTD is typically generated by low frequency stimulation (in the range of 1–3 Hz) given for prolonged periods of time (5–15 min) (Braunewell et al., 2001). Coincident EPSPs with action potentials (APs) evokes large calcium signals and leads to LTP whereas unpaired stimulation brings about LTD (Markram et al., 1997). LTD, like LTP possess the characteristics of longevity, input-specificity and associativity. The relative contributions of pre and postsynaptic mechanisms may vary at different times after induction and also across different classes of synapses. In cerebellum, PF-LTD can be induced by paired PF and

Elucidating the functional role of LTD *in vivo* has been challenging due to difficulty of inducing it *in vivo* and due to the lack of selective inhibitors for the same (Collingridge et al.,

LTD has been implicated in cerebellar motor learning. For example, studies using PKC transgenic mice showed that chronic PKC inhibition restricted to cerebellar PF-PC synapses exhibited compromised LTD and defective adaptation of VOR. But still a causal relationship remains to be demonstatred between PF-PC LTD and motor learning (De Zeeuw et al., 1998). To the contrary, recent studies using knockout mice which target the expression of PF-PC LTD by blocking internalization of AMPARs show that LTD is not necessary for motor learning. The mutant mice lacked PF-PC LTD but had no difficulty in performing motor learning skills like VOR adaptation, eyeblink conditioning, and locomotion learning on the Erasmus Ladder which covers a wide range of cerebellar learning behaviors

LTD is implicated in hippocampus dependent learning because of its property of depotentiation. Depotentiation in CA1 and DG has been shown when rats explore a novel environment or a familiar environment containing novel objects. Impairment of reversal learning in water maze was found to be associated with severely impaired hippocampal LTD in dopamine transporter knockout mice. LTD is also involved in learning regarding novelty detection. In freely moving rats, LTD is facilitated during exploration of complex

The various molecular mechanisms involved in LTD in brain have been discussed. LTD has been implicated in several physiological processes including learning and memory and also in development of visual system. Future studies will be focused on the aspects of protein synthesis and turnover involved in LTD in detail. Also the mechanisms by which LTD could be induced by neurotransmitters other than glutamate remains to be elucidated.

CF stimulation (Ito et al., 1982), which is typically applied at 1–4 Hz for 5 min.

2010). But still some of the physiological functions of LTD are known.

environments containing novel objects (Collingridge et al., 2010).

**4.4 LTD induction** 

**4.5 Physiological functions of LTD** 

(Schonewille et al., 2011).

(Collingridge et al., 2010).

extracellular Ca2+ (Fitzjohn et al., 2001). It also doesn't require CaMKII. Certain other proteins which are activated by mGluRs are arg 3.1(Arc), striatal-enriched protein tyrosine phosphatase (STEP) and microtubule associated protein 1B which are involved in AMPAR internalization (Collingridge et al., 2010).

#### **4.3 LTD in cerebellum**

Cerebellum is involved in motor learning and non-declarative memory. It is required in the adaptation of vestibulo ocular reflex (VOR) movements of the eyes needed to keep the retinal image stable. In the 1980s, Ito and colleagues provided the first experimental evidence for plasticity in the cerebellar cortex.

Of the three layers of cerebellum; the molecular layer, granular layer and the Purkinje layer, the Purkinje cells synapse on deep cerebellar nuclei which are the major output from the cerebellum (Bear et al., 2001). Purkinje cells modify the output. Purkinje cells receive excitatory input from two sources, viz, climbing fibre (CF) and parallel fibre (PF). Each Purkinje cell receives input from one inferior olive cell (which arises from medulla) via CF and this input is very powerful. This generates a very large EPSP that always strongly activates the postsynaptic Purkinje cells. The other input to the Purkinje cells, viz the PF arises from cerebellar granule (CG) cells. The CG cells in turn receive mossy fibres which arise from precerebellar nuclei. Purkinje cells receive synapses from more than one PF. The plasticity at PF-PC synapse is governed by Marr Albus (Marr, 1969; Albus, 1971) theory which explains the mechanism behind motor learning. It says that the plasticity of the PF synapse is effective if it is active at the same time as the CF input to the Purkinje cell. Activating CF results in massive calcium influx. Despite the large calcium influx, paired CF and PF stimulation results in LTD in Purkinje cells (Ito et al., 1982), whereas PF stimulation alone causes LTP (Lev Ram et al., 2002).

CF stimulation results in large input of EPSP. As a result voltage gated sodium channels open causing massive depolarization. This activates VGCCS, facilitating calcium entry. At the same time, the activation of PF results in glutamate release which binds to AMPA receptor and allows sodium ion entry. Altering AMPAR affects synaptic efficacy by changing the channel density. The other receptor activated is mGluR. It activates the release of downstream second messengers such as DAG and results in activation of PKC. αCaMKII is also activated along with PKC (Hansel, 2006). These kinases can phosphorylate the AMPAR. PKC can phosphorylate GluR2, the subunit of AMPAR at Ser-880 and this brings about receptor endocytosis (Chung et al., 2003). It is a critical event in the induction of cerebellar LTD (Fig. 6). This phosphorylation disrupts the binding of GluR2 to GRIP1 facilitating binding to PICK1 (Xia et al., 2000). Purkinje cells are enriched with GluR2/GluR3 receptors whereas it is poor in expressing GluR1 subunits. Purkinje cells lack NMDAR and thus LTD is mainly AMPAR mediated. The cerebellar LTD is the type of plasticity where information is stored as a decrease in the effectiveness of synaptic connection.

It is interesting to see that the LTP and LTD induction cascades in hippocampus and cerebellar Purkinje synapses are different and exhibit a mirror image like relationship (Jorntell & Hansel, 2006). As already discussed, LTD in cerebellum, is being brought about by kinases whereas in hippocampus it is brought about by phosphatases.

#### **4.4 LTD induction**

314 Neuroscience – Dealing with Frontiers

extracellular Ca2+ (Fitzjohn et al., 2001). It also doesn't require CaMKII. Certain other proteins which are activated by mGluRs are arg 3.1(Arc), striatal-enriched protein tyrosine phosphatase (STEP) and microtubule associated protein 1B which are involved in AMPAR

Cerebellum is involved in motor learning and non-declarative memory. It is required in the adaptation of vestibulo ocular reflex (VOR) movements of the eyes needed to keep the retinal image stable. In the 1980s, Ito and colleagues provided the first experimental

Of the three layers of cerebellum; the molecular layer, granular layer and the Purkinje layer, the Purkinje cells synapse on deep cerebellar nuclei which are the major output from the cerebellum (Bear et al., 2001). Purkinje cells modify the output. Purkinje cells receive excitatory input from two sources, viz, climbing fibre (CF) and parallel fibre (PF). Each Purkinje cell receives input from one inferior olive cell (which arises from medulla) via CF and this input is very powerful. This generates a very large EPSP that always strongly activates the postsynaptic Purkinje cells. The other input to the Purkinje cells, viz the PF arises from cerebellar granule (CG) cells. The CG cells in turn receive mossy fibres which arise from precerebellar nuclei. Purkinje cells receive synapses from more than one PF. The plasticity at PF-PC synapse is governed by Marr Albus (Marr, 1969; Albus, 1971) theory which explains the mechanism behind motor learning. It says that the plasticity of the PF synapse is effective if it is active at the same time as the CF input to the Purkinje cell. Activating CF results in massive calcium influx. Despite the large calcium influx, paired CF and PF stimulation results in LTD in Purkinje cells (Ito et al., 1982), whereas PF stimulation

CF stimulation results in large input of EPSP. As a result voltage gated sodium channels open causing massive depolarization. This activates VGCCS, facilitating calcium entry. At the same time, the activation of PF results in glutamate release which binds to AMPA receptor and allows sodium ion entry. Altering AMPAR affects synaptic efficacy by changing the channel density. The other receptor activated is mGluR. It activates the release of downstream second messengers such as DAG and results in activation of PKC. αCaMKII is also activated along with PKC (Hansel, 2006). These kinases can phosphorylate the AMPAR. PKC can phosphorylate GluR2, the subunit of AMPAR at Ser-880 and this brings about receptor endocytosis (Chung et al., 2003). It is a critical event in the induction of cerebellar LTD (Fig. 6). This phosphorylation disrupts the binding of GluR2 to GRIP1 facilitating binding to PICK1 (Xia et al., 2000). Purkinje cells are enriched with GluR2/GluR3 receptors whereas it is poor in expressing GluR1 subunits. Purkinje cells lack NMDAR and thus LTD is mainly AMPAR mediated. The cerebellar LTD is the type of plasticity where

information is stored as a decrease in the effectiveness of synaptic connection.

by kinases whereas in hippocampus it is brought about by phosphatases.

It is interesting to see that the LTP and LTD induction cascades in hippocampus and cerebellar Purkinje synapses are different and exhibit a mirror image like relationship (Jorntell & Hansel, 2006). As already discussed, LTD in cerebellum, is being brought about

internalization (Collingridge et al., 2010).

evidence for plasticity in the cerebellar cortex.

alone causes LTP (Lev Ram et al., 2002).

**4.3 LTD in cerebellum** 

LTD can be induced by prolonged periods of LFS by pairing baseline synaptic stimulation with depolarization i.e. normally at a frequency of 10 Hz. LFS of 1 Hz for 15 minutes brings about LTD in CA1 area of hippocampus of anaesthetized rabbit (Bliss & Lomo, 1973). This stimulation is preceded by baseline stimulation at frequencies such as 0.1–0.05 Hz to establish a reference level for basal synaptic transmission. Electrically induced LTD is typically generated by low frequency stimulation (in the range of 1–3 Hz) given for prolonged periods of time (5–15 min) (Braunewell et al., 2001). Coincident EPSPs with action potentials (APs) evokes large calcium signals and leads to LTP whereas unpaired stimulation brings about LTD (Markram et al., 1997). LTD, like LTP possess the characteristics of longevity, input-specificity and associativity. The relative contributions of pre and postsynaptic mechanisms may vary at different times after induction and also across different classes of synapses. In cerebellum, PF-LTD can be induced by paired PF and CF stimulation (Ito et al., 1982), which is typically applied at 1–4 Hz for 5 min.

#### **4.5 Physiological functions of LTD**

Elucidating the functional role of LTD *in vivo* has been challenging due to difficulty of inducing it *in vivo* and due to the lack of selective inhibitors for the same (Collingridge et al., 2010). But still some of the physiological functions of LTD are known.

LTD has been implicated in cerebellar motor learning. For example, studies using PKC transgenic mice showed that chronic PKC inhibition restricted to cerebellar PF-PC synapses exhibited compromised LTD and defective adaptation of VOR. But still a causal relationship remains to be demonstatred between PF-PC LTD and motor learning (De Zeeuw et al., 1998). To the contrary, recent studies using knockout mice which target the expression of PF-PC LTD by blocking internalization of AMPARs show that LTD is not necessary for motor learning. The mutant mice lacked PF-PC LTD but had no difficulty in performing motor learning skills like VOR adaptation, eyeblink conditioning, and locomotion learning on the Erasmus Ladder which covers a wide range of cerebellar learning behaviors (Schonewille et al., 2011).

LTD is implicated in hippocampus dependent learning because of its property of depotentiation. Depotentiation in CA1 and DG has been shown when rats explore a novel environment or a familiar environment containing novel objects. Impairment of reversal learning in water maze was found to be associated with severely impaired hippocampal LTD in dopamine transporter knockout mice. LTD is also involved in learning regarding novelty detection. In freely moving rats, LTD is facilitated during exploration of complex environments containing novel objects (Collingridge et al., 2010).

The various molecular mechanisms involved in LTD in brain have been discussed. LTD has been implicated in several physiological processes including learning and memory and also in development of visual system. Future studies will be focused on the aspects of protein synthesis and turnover involved in LTD in detail. Also the mechanisms by which LTD could be induced by neurotransmitters other than glutamate remains to be elucidated. (Collingridge et al., 2010).

Molecular Mechanisms in Synaptic Plasticity 317

that starts even at early stages of AD, results in a condition where minimum number of synapses are not available for cortical networks and impairment of synaptic plasticity occurs. The neuropathological hallmarks of Alzheimer's disease are the presence of intracellular neurofibrillary tangles (NFT) composed of hyperphosphorylated tau protein and extracellular neuritic plaques composed of amyloid β protein (Aβ) (Montoya, 2011). Aβ is produced by the sequential cleavage of amyloid precursor protein (APP) by β-secretase and γ-secretase (Haass & Selkoe, 1993). 40-residue Aβ (Aβ40) and 42-residue Aβ (Aβ42) are the most common isoforms of Aβ (Xia, 2010). Even before plaques could be observed, significant deficits in synaptic transmission have been detected by electrophysiological recordings from the hippocampus of transgenic mice over expressing APP (Hsia et al., 1999; Mucke et al., 2000). Thus aberrations in synaptic function are the early events followed by the formation of plaques and NFT (Funato et al., 1999; Hartl et al., 2008; Selkoe, 2002; Walsh

**5.1.1 Disruption of the plasticity of glutamatergic synaptic transmission** 

The alterations of synaptic plasticity which happens before synaptic loss may be initiating neurodegeneration. The spatial working memory and LTP were normal in young APP695SWE transgenic mice. There was reduction in LTP and deficits in behavioural performance with aged transgenic mice. The deterioration of LTP in dentate gyrus and CA1 and behavioural deficit appear in a correlated manner (Chapman et al., 1999). Aβ concentrations increased with age. Many lines of investigations show that oligomeric forms of Aβ species interfere with synaptic plasticity, inhibit LTP and impairs maintenance of LTP (Barghorn et al., 2005; Klyubin et al., 2008; Shankar et al., 2008; Walsh et al., 2002; Wang et al., 2002; Stephan et al., 2001). Aβ peptide when applied before and during HFS inhibits LTP induction in the dentate medial perforant path and Schaffer colllateral-CA1 pathway (Chen et al., 2000; Chen et al., 2002). The basal synaptic transmission or short-term synaptic plasticity remained intact. Aβ inhibits maintenance phase of L-LTP and also inhibits protein synthesis in the L-LTP phase when applied after HFS. The effects of Aβ on the induction of LTP and on L-LTP are independent of each other, working through multiple mechanisms (Chen et al., 2002). Different forms of LTP affected by Aβ will be reflected as deficits in different phases of memory and also at a concentration of Aβ, below that is required to

Aβ was shown to increase intracellular Ca2+ concentrations due to potentiation of currents through L-type Ca2+ channels (Ueda et al., 1997) and blockade of fast-inactivating K+ channels that leads to prolonged membrane depolarization and Ca2+ influx (Good et al., 1996). Basal synaptic transmission and NMDAR-dependent forms of LTP are impaired in the aging hippocampus (Foster & Norris, 1997), which correlates with deficits in spatial memory (Barnes & McNaughton, 1985; Diana et al., 1995). Aβ induced Ca2+ transients activate calcineurin and cause desensitization of NMDAR channels, reducing Ca2+ influx through these channels during LTP-inducing stimulus protocol. As a result induction of NMDAR-dependent forms of LTP, E-LTP and also L-LTP are suppressed (Chen et al., 2002). Aβ also inhibited NMDAR mediated EPSCs. Activated calcineurin could impair the mechanisms underlying the components of L-LTP and long-term memory. This could be

& Selkoe, 2004, as cited in Proctor et al., 2011).

produce neurotoxicity (Chen et al., 2002).

**5.1.2 Molecular events causing disruption of LTP** 

Fig. 7. Scheme of shared post synaptic signaling pathways leading to LTP and LTD
