Long-Term Depression in the Cerebellum, Hippocampus, and Neocortex

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Long-term depression (LTD) is a type of synaptic plasticity in which the efficacy of signal transmission across a synapse is persistently reduced after a certain triggering activity. LTD in the cerebellum was proposed as a theoretical possibility around 1970 and was detected a decade later (Ito, Sakurai, and Tongroach, 1982). So far, several subtypes of LTD varying in cellular and molecular mechanisms have been found in the cerebellum, hippocampus, and neocortex. LTD occurs not only in excitatory synapses, but also in inhibitory ones. LTD may weaken or functionally interrupt useless or erroneous synaptic connections between neurons, providing an opposing mechanism against long-term potentiation (LTP) in various forms of learning and memory.

Induction and Observation of LTD

The various forms of LTD revealed to occur in the cerebellum, cerebellum-like structures in fish, hippocampus and neocortex have the following characteristic features. In this article, references for the cerebellum are largely omitted because they can be found in recent review articles (Ito, 2001; Hansel, Linden, and D'Angelo, 2001).

In the Cerebellum and Cerebellum-Like Structures

In the cerebellar cortex each Purkinje cell receives two distinct types of excitatory synapses, one from parallel fibers and the other from a (normally single) climbing fiber. LTD occurs when these two types of synapses are activated repeatedly in approximate synchrony, leading to an enduring decrease in synaptic strength of the parallel fibers. In in vitro cerebellar slices, LTD is typically induced by conjunctive stimulation at 1 hertz for five minutes (300 stimuli), which reduces the transmission efficacy by 30 to 40 percent (see Figure 1). To induce LTD, climbing fiber stimulation is often replaced by current-induced membrane depolarization that enhances entry of Ca2+ ions into Purkinje cell dendrites as in the case of climbing fiber stimulation (see below). Conjunctive LTD can be followed for one to three hours without sign of recovery. It is robust in the sense that its induction does not depend critically on the timing between climbing fiber and parallel fiber stimulations.

Strong stimulation of parallel fibers alone causes homosynaptic LTD. Conjunctive LTD is accompanied by LTD in the neighboring synapses located within a distance of 100 micrometers, which are not involved in conjunctive stimulation. Other types of LTD also occur in the cerebellum.

Brains of certain fish species contain a cerebellum-like structure that lacks climbing fibers. LTD occurs in parallel fiber-evoked excitatory postsynaptic potentials (EPSPs) in Purkinje cell-like neurons when EPSPs repeatedly precede postsynaptic spikes within 60 milliseconds (Bell, Han, Sugawara, and Grant, 1997).

In the Hippocampus

In a pyramidal cell of the hippocampal CA1 region, a low-frequency stimulation (LFS) of a bundle of presynaptic fibers (1 hertz for five to fifteen minutes) typically induces LTD, while a high-frequency stimulation (five stimuli at 100 hertz repeated at 200-millisecond intervals for two seconds) induces LTP. Associative stimulation of presynaptic fibers with postsynaptic membrane depolarization (5 hertz for sixteen seconds) also induces LTD or LTP depending on the time from pre-to postsynaptic activities (Nishiyama et al., 2000). While LTP is induced when pre-synaptic stimulation falls within the window from -2 to 12 milliseconds after postsynaptic spikes, LTD occurs when presynaptic stimulation either precedes the postsynaptic spikes by 16 to 28 milliseconds or follows with a delay of 15 to 20 milliseconds (see Figure 2). Associative LTD occurs in both stimulated (homosynaptic LTD) and unstimulated (heterosynaptic LTD) synapses, while LTP occurs only homosynaptically. As revealed in a triple chain of cultured hippocampal neurons, induction of associative LTD in a synapse is accompanied by back-propagated induction of LTD in a synapse on a presynaptic cell (Fitzsimonds, Song, and Poo, 1997). Through these LTD/LTP inductions, temporal information coded in the timing of individual spikes may be converted into spatially distributed patterns of persistent synaptic modifications in a neural network (Bi and Poo, 1999).

In the Neocortex

LFS-induced LTD occurs in layer III neurons of the visual cortex following stimulation of the white matter or layer IV (Kirkwood and Bear, 1994). Associative LTP or LTD is induced when presynaptic stimulation is paired with membrane depolarization. A relatively large membrane depolarization induces LTP, while a relatively small membrane depolarization causes LTD (Artola, Brocher, and Singer, 1990). This dichotomy depends on the amount of Ca2+ influx evoked by the depolarization (see below). LTD has also been observed in the sensorimotor (Bindman, Murphy, and Pockett, 1988) and prefrontal cortex (Hirsch and Crepel, 1990).

Signal Transduction Underlying LTD

Conjunctive LTD is a purely postsynaptic event, but in other LTD subtypes, the contribution of pre-synaptic factors is not excluded. The following post-synaptic signal transduction processes have been analyzed in synaptically induced as well as in various reduced forms of LTD induced with chemical or electrical stimulation of neurons in place of synaptic activation (Ito, 2001).


LTD in excitatory synapses is, at least in part, due to a reduced number of functional AMPA receptor molecules in the postsynaptic membrane. Other types of receptors play roles in eventually inducing this change. Activation of NMDA receptors is required in LTD induction in excitatory synapses, but not in inhibitory synapses, of hippocampal neurons (Fitzsimonds, Song, and Poo, 1997). NMDA receptors are not functional in adult Purkinje cells. Parallel fiber synapses on Purkinje cells contain, besides AMPA receptors, type-1 metabotropic glutamate receptors (mGluR1s) and δ 2 glutamate receptors, whereas climbing fiber synapses contain AMPA receptors, type-1 corticotropin-releasing factor receptors (CRFR1s), and type-1 insulin-like growth factor receptors (IGF-1Rs). Inhibition of any of these receptors results in the blockage of LTD. The δ 2 receptor is an orphan receptor with an unknown function. mGluR1s (see below), CRFR1s, and IGF-1Rs are associated with G-proteins which are coupled with certain second messenger processes.

Calcium Entry and Release from Intracellular Stores

Induction of conjunctive LTD requires enhancement of the intracellular Ca2+ concentration, for it is blocked by injection of a Ca2+ chelator, EGTA, into Purkinje cells. Climbing fiber impulses evoke Ca2+ influx into dendrites of Purkinje cells through voltage-sensitive Ca2+ channels. As underlying homosynaptic LTD, Ca2+ influx also occurs in association with parallel fiber-induced EPSPs, if the EPSPs are sufficiently large to activate voltage-sensitive Ca2+ channels. Ca2+ ions are also released from intracellular stores in endoplasmic reticula, when inositol trisphosphate (IP3) receptors or ryanodine receptors on the reticula are activated. Inhibition of IP3 receptors, depletion of Ca2+ stores, or genetically induced loss of endoplasmic reticulum in dendritic spines results in the blockage of conjunctive LTD. However, intracellular Ca2+ release is not required for the reduced form of LTD in isolated Purkinje cells.

In hippocampal and neocortical neurons, LTD induction depends on the membrane potential, which determines the entry of Ca2+ ions into neurons through cation channels associated with a NMDA receptors. These channels are normally blocked by Mg2+ ions that are removed at a depolarized membrane potential level. Modest and strong activations of NMDA receptors lead to LTD and LTP, respectively. Reduction of postsynaptic Ca2+ entry by partial blockage of NMDA receptors converts LTP to LTD (Nishiyama et al., 2000). LTP is also converted to LTD by injecting EGTA into neocortical neurons (Kimura, Tsumoto, Nishigori, and Yoshimura, 1990). Induction of homosynaptic and heterosynaptic LTD in the hippocampus requires functional ryanodine and IP3 receptors, respectively. Functional blockade or genetic deletion of type-1 IP3 receptors leads to a conversion of LTD to LTP and elimination of heterosynaptic LTD, while blockage of ryanodine receptors eliminated only homosynaptic LTD (Nishiyama et al., 2000).

Metabotropic Glutamate Receptor-Driven Processes

Activation of mGluR1s in Purkinje cells results in activation of phopholipase C (PLC), which generates IP3 (see above) and diacylglycerol (DAG) from membrane phospholipid. DAG activates PKC. mGluR1s could also be coupled with phospholipase A2 (PLA2), which produces arachidonic acid and oleic acid from membrane phospholipid. Inhibition of mGluR1s, PKC, PLA2 or receptors of IP3 prevents LTD from induction, indicating that these are involved in LTD induction. Application of an mGluR5 agonist, DHPG, effectively induces LTD in hippocampal CA1 neurons (Fitzjohn, Kingston, Lodge, and Collingridge, 1999), which has been studied as a reduced form of hippocampal LTD.

Nitric Oxide and cGMP

Cerebellar granule cells contain a neuronal type of nitric oxide synthase (nNOS), and parallel fiber terminals release NO upon stimulation. NO, so released, diffuses into Purkinje cells, and activates guanylate cyclase to increase the level of cyclic GMP (cGMP). cGMP in turn activates cGMP-dependent protein kinase (PKG). The role of NO in the induction of conjunctive LTD is evident because it is blocked when an inhibitor of NO synthase, L-NMMA, or hemoglobin (HB) which absorbs NO, is applied to cerebellar slices, or in those mice deficient of nNOS. Furthermore, sodium nitroprusside that releases NO or a membrane-soluble derivative of cGMP induces an LTD-like phenomenon when applied to cerebellar slices together with AMPA. However, there is no evidence that a reduced form of LTD in isolated Purkinje cells requires a NO-cGMP pathway.

Protein Kinases and Phosphatases

Induction of cerebellar conjunctive LTD requires activities of PKC, PKG, protein tyrosin kinase (PTK) and mitogen-activated protein kinase (MAPK) but not PKA. PKC phosphorylates serine-880 of AMPA receptors (see below), and PTK interacts with PKC. PKG acts on a PKG-specific substrate, G-substance, which is richly contained in Purkinje cells. The phosphorylated G-substrate was found to be a potent inhibitor of protein phosphatases, presumably type 2A. Inhibitors of PP1/2A induce LTD when combined with stimulation of AMPA receptors.

PKC and PTK, but not PKA, are involved in mGluR-induced LTD in neurons of hippocampal dentate gyrus (Camodeca, Breakwell, Rowan, and Anwyl, 1999). A Ca2+/ Calmodulin-dependent protein phosphatase, calcineurin (or PP2B), mediates LTD induction in the hippocampus because a calcineurin inhibitor, FK506, blocks LFS induction of LTD in the visual cortex (Torii, Kamishita, Otsu, and Tsumoto, 1995). Mice deficient in the calcineurin catalytic unit B1 exhibited significantly diminished LFS-induced LTD in the hippocampus (Zeng et al., 2001). This is in contrast to the observation in Purkinje cells in which inhibition of PP2A facilitates conjunctive LTD. Calcineurin is also involved in LTD induction downregulating GABAA receptors in inhibitory synapses of the hippocampus (Lu, Mansuy, Kandel, and Roder, 2000).

Protein Synthesis

Translational inhibitors depressed the late phase of LTD in cultured Purkinje cells, but they abolished the entire LTD including its early phase in cerebellar slices. Studies using five-minute pulses of translational inhibitors revealed that a quickly turned over protein synthesis was required for LTD induction only during the fifteen-minute period from the onset of conjunctive stimulation (Karacho et al., 2001). A five-minute pulse application of transcriptional inhibitors also effectively blocked LTD induction with a delay of thirty minutes. Protein synthesis also plays roles in hippocampal neurons, in which a translational inhibitor blocked the late phase of LFS-induced LTD (Kauderer and Kandel, 2000) as well as the entire course of DHPG-induced LTD (Huber, Kayser, and Bear, 2000).

Phosphorylation and Inactivation of AMPA Receptors

Among subunits constituting AMPA receptors, the GluR2 subunits are dominant in parallel fiber-Purkinje cell synapses. Evidence suggests that at the end stage of cerebellar conjunctive LTD, GluR2 is phosphorylated at its serine-880 anchorage to GRIP, and freed AMPA receptors are removed from post-synaptic membrane by internalization via endocytosis. Similar consideration applies to associative LTD in hippocampal neurons (Luscher, Nicoli, Malenka, and Muller, 2000; Man et al., 2000).

Functional Roles of LTD

In the cerebellum-like structure of fish, LTD plays a clear role of lessening the effects of sensory inputs coinciding with command signals in order to allow only unpredicted sensory inputs to stand out, thus forming an adaptive sensory processor. Involvement of LTD in certain forms of cerebellar learning has been demonstrated using pharmacological or genetic means that impair LTD (Ito, 2001). Adaptation of reflexive eye movements was abolished by applying an NO scavenger or transfecting a pseudo inhibitor of PKC to the cerebellum. An NO scavenger or NOS inhibitor also blocked adaptation in smooth pursuit eye movement to repeated sudden increases in the velocity of the moving spotlight. Mice deficient in glial fibrillary acidic protein or phospholipase C β 4 (Miyata et al., 2001) lacked LTD and did not exhibit the classical conditioning of eye blinking. A walking cat or mouse normally adapted to a sudden change in the running belt conditions, but not under the influences of NO scavenger or NOS inhibitor or mGluR1 deficiency, which block LTD.

Mice in which hippocampal LTD was diminished due to conditional knockout of calcineurin were specifically impaired in working memory and episodic-like memory tasks, including the delayed matching-to-place task and the radial maze task (Zeng et al., 2001).

Accumulating evidence indicates that LTD is a major type of synaptic plasticity that plays roles in various forms of learning and memory. It will be a future task to confirm whether LTD is converted to a permanent memory, and if so, by what mechanism is it converted. Researchers should also determine the specific functional roles of LTD in various forms of learning and memory.



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