Isolation of CA1 Nuclear Enriched Fractions from Hippocampal Slices to Study Activity-dependent Nuclear Import of Synapto-nuclear Messenger Proteins

Studying activity dependent protein expression, subcellular translocation, or phosphorylation is essential to understand the underlying cellular mechanisms of synaptic plasticity. Long-term potentiation (LTP) and long-term depression (LTD) induced in acute hippocampal slices are widely accepted as cellular models of learning and memory. There are numerous studies that use live cell imaging or immunohistochemistry approaches to visualize activity dependent protein dynamics. However these methods rely on the suitability of antibodies for immunocytochemistry or overexpression of fluorescence-tagged proteins in single neurons. Immunoblotting of proteins is an alternative method providing independent confirmation of the findings. The first limiting factor in preparation of subcellular fractions from individual tetanized hippocampal slices is the low amount of material. Second, the handling procedure is crucial because even very short and minor manipulations of living slices might induce activation of certain signaling cascades. Here we describe an optimized workflow in order to obtain sufficient quantity of nuclear enriched fraction of sufficient purity from the CA1 region of acute hippocampal slices from rat brain. As a representative example we show that the ERK1/2 phosphorylated form of the synapto-nuclear protein messenger Jacob actively translocates to the nucleus upon induction of LTP and can be detected in a nuclear enriched fraction from CA1 neurons.     

Calcium signals in long-term potentiation and long-term depression

We describe postsynaptic Ca2+ signals that subserve induction of two forms of neuronal plasticity, long-term potentiation (LTP) and long-term depression (LTD), in rat hippocampal neurons. The common induction protocol for LTP, a 1-s, 50-Hz tetanus, generates Ca2+ increases of about 50-Hz in dendritic spines of CA1 neurons. These very large increases, measured using a low affinity indicator (Mg fura 5), were found only in the spines and tertiary dendrites, and were dependent upon influx through N-methyl-D-aspartate (NMDA) gated channels. High affinity Ca2+ indicators (e.g., fura 2) are unable to demonstrate these events. In acute slices, neighboring dendritic branches often showed very different responses to a tetanus, and in some instances, neighboring spines on the same dendrite responded differently. LTD in mature CA1 neurons was induced by a low frequency stimulus protocol (2 Hz, 900 pulses), in the presence of GABA- and NMDA-receptor blockers. This LTD protocol produced dendritic Ca2+ increases of <1 microM. Duration of the Ca2+ increase was approximately 30 s and was due to voltage-gated Ca2+ influx. Finally, the ability of synaptically addressed Ca2+ stores to release Ca2+ was studied in CA3 neurons and was found to require immediate preloading and high intensity presynaptic stimulation, conditions unlike normal LTP-LTD protocols.     

Unconjugated Bilirubin Exposure Impairs Hippocampal Long-Term Synaptic Plasticity

Background

Jaundice is one of the most common problems encountered in newborn infants, due to immaturity of hepatic conjugation and transport processes for bilirubin. Although the majority of neonatal jaundice is benign, some neonates with severe hyperbilirubinemia develop bilirubin encephalopathy or kernicterus. Accumulation of unconjugated bilirubin (UCB) in selected brain regions may result in temporary or permanent impairments of auditory, motor, or cognitive function; however, the molecular mechanisms by which UCB elicits such neurotoxicity are still poorly understood. The present study is undertaken to investigate whether prolonged exposure of rat organotypic hippocampal slice cultures to UCB alters the induction of long-term synaptic plasticity.

Methodology/Principal Findings

Using electrophysiological recording techniques, we find that exposure of hippocampal slice cultures to clinically relevant concentrations of UCB for 24 or 48 h results in an impairment of CA1 long-term potentiation (LTP) and long-term depression (LTD) induction in a time- and concentration-dependent manner. Hippocampal slice cultures stimulated with UCB show no changes in the secretion profiles of the pro-inflammatory cytokines, interleukin-1β and tumor necrosis factor-α, or the propidium ioide uptake. UCB treatment produced a significant decrease in the levels of NR1, NR2A and NR2B subunits of N-methyl-D-aspartate (NMDA) receptors through a calpain-mediated proteolytic cleavage mechanism. Pretreatment of the hippocampal slice cultures with NMDA receptor antagonist or calpain inhibitors effectively prevented the UCB-induced impairment of LTP and LTD.

Conclusion/Significance

Our results indicate that the proteolytic cleavage of NMDA receptor subunits by calpain may play a critical role in mediating the UCB-induced impairment of long-term synaptic plasticity in the hippocampus. These observations provide new insights into the molecular mechanisms underlying UCB-induced impairment of hippocampal synaptic plasticity which, in turn, might provide opportunities for the development of novel therapeutic strategies that targets these pathways for treatment.     

Control of Cerebellar Long-Term Potentiation by P-Rex-Family Guanine-Nucleotide Exchange Factors and Phosphoinositide 3-Kinase

Background

Long-term potentiation (LTP) at the parallel fibre–Purkinje cell synapse in the cerebellum is a recently described and poorly characterized form of synaptic plasticity. The induction mechanism for LTP at this synapse is considered reciprocal to “classical” LTP at hippocampal CA1 pyramidal neurons: kinases promote increased trafficking of AMPA receptors into the postsynaptic density in the hippocampus, whereas phosphatases decrease internalization of AMPA receptors in the cerebellum. In the hippocampus, LTP occurs in overlapping phases, with the transition from early to late phases requiring the consolidation of initial induction processes by structural re-arrangements at the synapse. Many signalling pathways have been implicated in this process, including PI3 kinases and Rho GTPases.

Principal Findings

We hypothesized that analogous phases are present in cerebellar LTP, and took as the starting point for investigation our recent discovery that P-Rex – a Rac guanine nucleotide exchange factor which is activated by PtdIns(3,4,5)P₃ – is highly expressed in mouse cerebellar Purkinje neurons and plays a role in motor coordination. We found that LTP evoked at parallel fibre synapses by 1 Hz stimulation or by NO donors was not sustained beyond 30 min when P-Rex was eliminated or Rac inhibited, suggesting that cerebellar LTP exhibits a late phase analogous to hippocampal LTP. In contrast, inhibition of PI3 kinase activity eliminated LTP at the induction stage.

Conclusions

Our data suggest that a PI3K/P-Rex/Rac pathway is required for late phase LTP in the mouse cerebellum, and that other PI3K targets, which remain to be discovered, control LTP induction.     

Nitric oxide and carbon monoxide as possible retrograde messengers in hgippocampal long-term potentiation

We have been investigating the hypothesis that the membrane-permeant molecules nitric oxide (NO) and carbon monoxide(CO) may act as retrograde messengers during long-term potentiation (LTP). Inhibitors of either NO synthase or heme oxygenase, the enzyme that produces CO, blocked induction of LTP in the CA1 region of hippocampal slices. Brief application of either NO or CO to slices produced a rapid and long-lasting increase in the size of synaptic potentials if, and only if, the application occurred at the same time as weak tetanic stimulation of the presynaptic fibers. The long-term enhancement by NO or CO was spatially restricted to synapses from active presynaptic fibers and appeared to involve mechanisms utilized by LTP, occluding the subsequent induction of LTP by strong tetanic stimulation. The enhancement by No or CO was not blocked by the NMDA receptor blocker APV, suggesting that NO and CO act downstream for the NMDA receptor. In other systems, both NO and CO produce many of their effects by activation of soluble guanylyl cyclase nd cGMP-dependent protein kinase. An inhibitor of soluble guabylyl cyclase blocked the induction of normal LTP. Conversely, membrane-permeabel analog 8-Br-cGMP produced a rapid onset and long-lasting synaptic enhancement if, and only if, it was applied at the same time as weak presynaptic stimulation. Similarly, two inhibitors of cGMP-dependent protein kinase blocked the induction of normal LTP, and a selective activator of cGMP-dependent protein kinase produced activity-dependent long-lasting synaptic enhancement. 8-Br-cGMP also produced and activity-dependent, long-lasting increase in the amplitude of evoked synaptic current between pairs of hippocampal neurons in dissociated cell culture. In addition, 8-Br-cGMP, like NO, produced a long-lasting increase in the frequency of spontaneous miniature synaptic currents. These results are consistent with the hypothesis that NO and CO, either alone or in combination, serve as retrograde messengers that produce activity-dependent presynaptic enhancement, perhaps by stimulating soluble guanbylyl cyclase and cGMP-dependent protein kinase, during LTP in hippocampus. 1994 John Wiley & Sons, Inc.     

Coincident activity of converging pathways enables simultaneous long-term potentiation and long-term depression in hippocampal CA1 network in vivo

Memory is believed to depend on activity-dependent changes in the strength of synapses, e.g. long-term potentiation (LTP) and long-term depression (LTD), which can be determined by the sequence of coincident pre- and postsynaptic activity, respectively. It remains unclear, however, whether and how coincident activity of converging efferent pathways can enable LTP and LTD in the pathways simultaneously. Here, we report that, in pentobarbital-anesthetized rats, stimulation (600 pulses, 5 Hz) to Schaffer preceding to commissural pathway within a 40-ms timing window induced similar magnitudes of LTP in both pathways onto synapses of CA1 neurons, with varied LTP magnitudes after reversal of the stimulation sequence. In contrast, in urethane-anesthetized or freely-moving rats, the stimulation to Schaffer preceding to commissural pathway induced Schaffer LTP and commissural LTD simultaneously within a 40-ms timing window, without affecting synaptic efficacy in the reversed stimulation sequence. Coincident activity of Schaffer pathways confirmed the above findings under pentobarbital and urethane anesthesia. Thus, coincident activity of converging afferent pathways tends to switch the pathways to be LTP only or LTP/LTD depending on the activity states of the hippocampus. This network rule strengthens the view that activity-dependent synaptic plasticity may well contribute to memory process of the hippocampal network with flexibility or stability from one state to another.     

Persistent Synapse Loss Induced by Repetitive LTD in Developing Rat Hippocampal Neurons

Synaptic pruning is a physiological event that eliminates excessive or inappropriate synapses to form proper synaptic connections during development of neurons. Appropriate synaptic pruning is required for normal neural development. However, the mechanism of synaptic pruning is not fully understood. Strength of synaptic activity under competitive circumstances is thought to act as a selective force for synaptic pruning. Long-term depression (LTD) is a synaptic plasticity showing persistent decreased synaptic efficacy, which is accompanied by morphological changes of dendritic spines including transient retraction. Repetitive induction of LTD has been shown to cause persistent loss of synapses in mature neurons. Here, we show that multiple, but not single, induction of LTD caused a persistent reduction in the number of dendritic synapses in cultured rat developing hippocampal neurons. When LTD was induced in 14 days in vitro cultures by application of (RS)-3,5-dihydroxyphenylglycine (DHPG), a group I metabotropic glutamate receptor (mGluR) agonist, and repeated three times with a one day interval, there was a significant decrease in the number of dendritic synapses. This effect continued up to at least two weeks after the triple LTD induction. The persistent reduction in synapse number occurred in the proximal dendrites, but not the distal dendrites, and was prevented by simultaneous application of the group I/II mGluR antagonist (S)-a-methyl-4-carboxyphenylglycine (MCPG). In conclusion, we found that repetitive LTD induction in developing neurons elicits synaptic pruning and contributes to activity-dependent regulation of synapse number in rat hippocampal neurons.     

Phosphorylation of proteins involved in activity-dependent forms of synaptic plasticity is altered in hippocampal slices maintained in vitro

The acute hippocampal slice preparation has been widely used to study the cellular mechanisms underlying activity-dependent forms of synaptic plasticity such as long-term potentiation (LTP) and long-term depression (LTD). Although protein phosphorylation has a key role in LTP and LTD, little is known about how protein phosphorylation might be altered in hippocampal slices maintained in vitro. To begin to address this issue, we examined the effects of slicing and in vitro maintenance on phosphorylation of six proteins involved in LTP and/or LTD. We found that AMPA receptor (AMPAR) glutamate receptor 1 (GluR1) subunits are persistently dephosphorylated in slices maintained in vitro for up to 8 h. alpha calcium/calmodulin-dependent kinase II (alphaCamKII) was also strongly dephosphorylated during the first 3 h in vitro but thereafter recovered to near control levels. In contrast, phosphorylation of the extracellular signal-regulated kinase ERK2, the ERK kinase MEK, proline-rich tyrosine kinase 2 (Pyk2), and Src family kinases was significantly, but transiently, increased. Electrophysiological experiments revealed that the induction of LTD by low-frequency synaptic stimulation was sensitive to time in vitro. These findings indicate that phosphorylation of proteins involved in N-methyl-D-aspartate (NMDA) receptor-dependent forms of synaptic plasticity is altered in hippocampal slices and suggest that some of these changes can significantly influence the induction of LTD.     

The Role of Growth Retardation in Lasting Effects of Neonatal Dexamethasone Treatment on Hippocampal Synaptic Function

Background

Dexamethasone (DEX), a synthetic glucocorticoid, is commonly used to prevent or lessen the morbidity of chronic lung disease in preterm infants. However, evidence is now increasing that this clinical practice negatively affects somatic growth and may result in long-lasting neurodevelopmental deficits. We therefore hypothesized that supporting normal somatic growth may overcome the lasting adverse effects of neonatal DEX treatment on hippocampal function.

Methodology/Principal Findings

To test this hypothesis, we developed a rat model using a schedule of tapering doses of DEX similar to that used in premature infants and examined whether the lasting influence of neonatal DEX treatment on hippocampal synaptic plasticity and memory performance are correlated with the deficits in somatic growth. We confirmed that neonatal DEX treatment switched the direction of synaptic plasticity in hippocampal CA1 region, favoring low-frequency stimulation- and group I metabotropic glutamate receptor agonist (S)-3,5,-dihydroxyphenylglycine-induced long-term depression (LTD), and opposing the induction of long-term potentiation (LTP) by high-frequency stimulation in the adolescent period. The effects of DEX on LTP and LTD were correlated with an increase in the autophosphorylation of Ca²⁺/calmodulin-dependent protein kinase II at threonine-286 and a decrease in the protein phosphatase 1 expression. Neonatal DEX treatment resulted in a disruption of memory retention subjected to object recognition task and passive avoidance learning. The adverse effects of neonatal DEX treatment on hippocampal synaptic plasticity and memory performance of the animals from litters culled to 4 pups were significantly less than those for the 8-pup litters. However, there was no significant difference in maternal care between groups.

Conclusion/Significance

Our results demonstrate that growth retardation plays a crucial role in DEX-induced long-lasting influence of hippocampal function. Our findings suggest that therapeutic strategies designed to support normal development and somatic growth may exert beneficial effects to reduce lasting adverse effects following neonatal DEX treatment.     

Insulin modulates hippocampal activity-dependent synaptic plasticity in a N-methyl-d-aspartate receptor and phosphatidyl-inositol-3-kinase-dependent manner

Insulin and its receptor are both present in the central nervous system and are implicated in neuronal survival and hippocampal synaptic plasticity. Here we show that insulin activates phosphatidylinositol 3-kinase (PI3K) and protein kinase B (PKB), and results in an induction of long-term depression (LTD) in hippocampal CA1 neurones. Evaluation of the frequency-response curve of synaptic plasticity revealed that insulin induced LTD at 0.033 Hz and LTP at 10 Hz, whereas in the absence of insulin, 1 Hz induced LTD and 100 Hz induced LTP. LTD induction in the presence of insulin required low frequency synaptic stimulation (0.033 Hz) and blockade of GABAergic transmission. The LTD or LTP induced in the presence of insulin was N-methyl-d-aspartate (NMDA) receptor specific as it could be inhibited by alpha-amino-5-phosphonopentanoic acid (APV), a specific NMDA receptor antagonist. LTD induction was also facilitated by lowering the extracellular Mg(2+) concentration, indicating an involvement of NMDA receptors. Inhibition of PI3K signalling or discontinuing synaptic stimulation also prevented this LTD. These results show that insulin modulates activity-dependent synaptic plasticity, which requires activation of NMDA receptors and the PI3K pathway. The results obtained provide a mechanistic link between insulin and synaptic plasticity, and explain how insulin functions as a neuromodulator.     

Cholinergic Neurotransmission and Synaptic Plasticity Concerning Memory Processing

The brain is able to change the synaptic strength in response to stimuli that leave a memory trace. Long-term potentiation (LTP) and long-term depression (LTD) are forms of activity-dependent synaptic plasticity proposed to underlie memory. The induction of LTP appears mediated by glutamate acting on AMPA and then on NMDA receptors. Cholinergic muscarinic agonists facilitate learning and memory. Acetylcholine depolarizes pyramidal neurons, reduces inhibition, upregulates NMDA channels and activates the phosphoinositide cascade. Postsynaptic Ca²⁺ rises and stimulates Ca-dependent PK, promoting synaptic changes. Electroencephalographic desynchronization and hippocampal theta rhythm are related to learning and memory, are inducible by Cholinergic agonists and elicited by hippocampal Cholinergic terminals. Their loss results in memory deficits. Hence, Cholinergic pathways may act synergically with glutamatergic transmission, regulating and leading to synaptic plasticity. The stimulation that induces plasticity in vivo has not been established. The patterns for LTP/LTD induction in vitro may be due to the loss of ascending Cholinergic inputs. As a rat explores pyramidal cells fire bursts that could be relevant to plasticity.     

Neurabin contributes to hippocampal long-term potentiation and contextual fear memory

Neurabin is a scaffolding protein that interacts with actin and protein phosphatase-1. Highly enriched in the dendritic spine, neurabin is important for spine morphogenesis and synaptic formation. However, less is known about the role of neurabin in hippocampal plasticity and its possible effect on behavioral functions. Using neurabin knockout (KO) mice, here we studied the function of neurabin in hippocampal synaptic transmission, plasticity and behavioral memory. We demonstrated that neurabin KO mice showed a deficit in contextual fear memory but not auditory fear memory. Whole-cell patch clamp recordings in the hippocampal CA1 neurons showed that long-term potentiation (LTP) was significantly reduced, whereas long-term depression (LTD) was unaltered in neurabin KO mice. Moreover, increased AMPA receptor but not NMDA receptor-mediated synaptic transmission was found in neurabin KO mice, and is accompanied by decreased phosphorylation of GluR1 at the PKA site (Ser845) but no change at the CaMKII/PKC site (Ser831). Pre-conditioning with LTD induction rescued the following LTP in neurabin KO mice, suggesting the loss of LTP may be due to the saturated synaptic transmission. Our results indicate that neurabin regulates contextual fear memory and LTP in hippocampal CA1 pyramidal neurons.     

Heterogeneous nuclear ribonucleoprotein k interacts with Abi-1 at postsynaptic sites and modulates dendritic spine morphology

Background

Abelson-interacting protein 1 (Abi-1) plays an important role for dendritic branching and synapse formation in the central nervous system. It is localized at the postsynaptic density (PSD) and rapidly translocates to the nucleus upon synaptic stimulation. At PSDs Abi-1 is in a complex with several other proteins including WASP/WAVE or cortactin thereby regulating the actin cytoskeleton via the Arp 2/3 complex.

Principal Findings

We identified heterogeneous nuclear ribonucleoprotein K (hnRNPK), a 65 kDa ssDNA/RNA-binding-protein that is involved in multiple intracellular signaling cascades, as a binding partner of Abi-1 at postsynaptic sites. The interaction with the Abi-1 SH3 domain is mediated by the hnRNPK-interaction (KI) domain. We further show that during brain development, hnRNPK expression becomes more and more restricted to granule cells of the cerebellum and hippocampal neurons where it localizes in the cell nucleus as well as in the spine/dendritic compartment. The downregulation of hnRNPK in cultured hippocampal neurons by RNAi results in an enlarged dendritic tree and a significant increase in filopodia formation. This is accompanied by a decrease in the number of mature synapses. Both effects therefore mimic the neuronal morphology after downregulation of Abi-1 mRNA in neurons.

Conclusions

Our findings demonstrate a novel interplay between hnRNPK and Abi-1 in the nucleus and at synaptic sites and show obvious similarities regarding both protein knockdown phenotypes. This indicates that hnRNPK and Abi-1 act synergistic in a multiprotein complex that regulates the crucial balance between filopodia formation and synaptic maturation in neurons.     

Synaptic Plasticity in Medial Vestibular Nucleus Neurons: Comparison with Computational Requirements of VOR Adaptation

Background

Vestibulo-ocular reflex (VOR) gain adaptation, a longstanding experimental model of cerebellar learning, utilizes sites of plasticity in both cerebellar cortex and brainstem. However, the mechanisms by which the activity of cortical Purkinje cells may guide synaptic plasticity in brainstem vestibular neurons are unclear. Theoretical analyses indicate that vestibular plasticity should depend upon the correlation between Purkinje cell and vestibular afferent inputs, so that, in gain-down learning for example, increased cortical activity should induce long-term depression (LTD) at vestibular synapses.

Methodology/Principal Findings

Here we expressed this correlational learning rule in its simplest form, as an anti-Hebbian, heterosynaptic spike-timing dependent plasticity interaction between excitatory (vestibular) and inhibitory (floccular) inputs converging on medial vestibular nucleus (MVN) neurons (input-spike-timing dependent plasticity, iSTDP). To test this rule, we stimulated vestibular afferents to evoke EPSCs in rat MVN neurons in vitro. Control EPSC recordings were followed by an induction protocol where membrane hyperpolarizing pulses, mimicking IPSPs evoked by flocculus inputs, were paired with single vestibular nerve stimuli. A robust LTD developed at vestibular synapses when the afferent EPSPs coincided with membrane hyperpolarisation, while EPSPs occurring before or after the simulated IPSPs induced no lasting change. Furthermore, the iSTDP rule also successfully predicted the effects of a complex protocol using EPSP trains designed to mimic classical conditioning.

Conclusions

These results, in strong support of theoretical predictions, suggest that the cerebellum alters the strength of vestibular synapses on MVN neurons through hetero-synaptic, anti-Hebbian iSTDP. Since the iSTDP rule does not depend on post-synaptic firing, it suggests a possible mechanism for VOR adaptation without compromising gaze-holding and VOR performance in vivo.     

Regulation of dendritic excitability by activity-dependent trafficking of the A-type K+ channel subunit Kv4.2 in hippocampal neurons

Voltage-gated A-type K+ channel Kv4.2 subunits are highly expressed in the dendrites of hippocampal CA1 neurons. However, little is known about the subcellular distribution and trafficking of Kv4.2-containing channels. Here we provide evidence for activity-dependent trafficking of Kv4.2 in hippocampal spines and dendrites. Live imaging and electrophysiological recordings showed that Kv4.2 internalization is induced rapidly upon glutamate receptor stimulation. Kv4.2 internalization was clathrin mediated and required NMDA receptor activation and Ca2+ influx. In dissociated hippocampal neurons, mEPSC amplitude depended on functional Kv4.2 expression level and was enhanced by stimuli that induced Kv4.2 internalization. Long-term potentiation (LTP) induced by brief glycine application resulted in synaptic insertion of GluR1-containing AMPA receptors along with Kv4.2 internalization. We also found evidence of Kv4.2 internalization upon synaptically evoked LTP in CA1 neurons of hippocampal slice cultures. These results present an additional mechanism for synaptic integration and plasticity through the activity-dependent regulation of Kv4.2 channel surface expression.     

Structural Plasticity Can Produce Metaplasticity

Background

Synaptic plasticity underlies many aspect of learning memory and development. The properties of synaptic plasticity can change as a function of previous plasticity and previous activation of synapses, a phenomenon called metaplasticity. Synaptic plasticity not only changes the functional connectivity between neurons but in some cases produces a structural change in synaptic spines; a change thought to form a basis for this observed plasticity. Here we examine to what extent structural plasticity of spines can be a cause for metaplasticity. This study is motivated by the observation that structural changes in spines are likely to affect the calcium dynamics in spines. Since calcium dynamics determine the sign and magnitude of synaptic plasticity, it is likely that structural plasticity will alter the properties of synaptic plasticity.

Methodology/Principal Findings

In this study we address the question how spine geometry and alterations of N-methyl-D-aspartic acid (NMDA) receptors conductance may affect plasticity. Based on a simplified model of the spine in combination with a calcium-dependent plasticity rule, we demonstrated that after the induction phase of plasticity a shift of the long term potentiation (LTP) or long term depression (LTD) threshold takes place. This induces a refractory period for further LTP induction and promotes depotentiation as observed experimentally. That resembles the BCM metaplasticity rule but specific for the individual synapse. In the second phase, alteration of the NMDA response may bring the synapse to a state such that further synaptic weight alterations are feasible. We show that if the enhancement of the NMDA response is proportional to the area of the post synaptic density (PSD) the plasticity curves most likely return to the initial state.

Conclusions/Significance

Using simulations of calcium dynamics in synaptic spines, coupled with a biophysically motivated calcium-dependent plasticity rule, we find under what conditions structural plasticity can form the basis of synapse specific metaplasticity.     

Endocannabinoids Differentially Modulate Synaptic Plasticity in Rat Hippocampal CA1 Pyramidal Neurons

Background

Hippocampal CA1 pyramidal neurons receive two excitatory glutamatergic synaptic inputs: their most distal dendritic regions in the stratum lacunosum-moleculare (SLM) are innervated by the perforant path (PP), originating from layer III of the entorhinal cortex, while their more proximal regions of the apical dendrites in the stratum radiatum (SR) are innervated by the Schaffer-collaterals (SC), originating from hippocampal CA3 neurons. Endocannabinoids (eCBs) are naturally occurring mediators capable of modulating both GABAergic and glutamatergic synaptic transmission and plasticity via the CB1 receptor. Previous work on eCB modulation of excitatory synapses in the CA1 region largely focuses on the SC pathway. However, little information is available on whether and how eCBs modulate glutamatergic synaptic transmission and plasticity at PP synapses.

Methodology/Principal Findings

By employing somatic and dendritic patch-clamp recordings, Ca²⁺ uncaging, and immunostaining, we demonstrate that there are significant differences in low-frequency stimulation (LFS)- or DHPG-, an agonist of group I metabotropic glutamate receptors (mGluRs), induced long-term depression (LTD) of excitatory synaptic transmission between SC and PP synapses in the same pyramidal neurons. These differences are eliminated by pharmacological inhibition with selective CB1 receptor antagonists or genetic deletion of the CB1 receptor, indicating that these differences likely result from differential modulation via a CB1 receptor-dependent mechanism. We also revealed that depolarization-induced suppression of excitation (DSE), a form of short-term synaptic plasticity, and photolysis of caged Ca²⁺-induced suppression of Excitatory postsynaptic currents (EPSCs) were less at the PP than that at the SC. In addition, application of WIN55212 (WIN) induced a more pronounced inhibition of EPSCs at the SC when compared to that at the PP.

Conclusions/Significance

Our results suggest that CB1 dependent LTD and DSE are differentially expressed at the PP versus SC synapses in the same neurons, which may have an impact on synaptic scaling, integration and plasticity of hippocampal CA1 pyramidal neurons.     

A cooperative switch determines the sign of synaptic plasticity in distal dendrites of neocortical pyramidal neurons

Pyramidal neurons in the cerebral cortex span multiple cortical layers. How the excitable properties of pyramidal neuron dendrites allow these neurons to both integrate activity and store associations between different layers is not well understood, but is thought to rely in part on dendritic backpropagation of action potentials. Here we demonstrate that the sign of synaptic plasticity in neocortical pyramidal neurons is regulated by the spread of the backpropagating action potential to the synapse. This creates a progressive gradient between LTP and LTD as the distance of the synaptic contacts from the soma increases. At distal synapses, cooperative synaptic input or dendritic depolarization can switch plasticity between LTD and LTP by boosting backpropagation of action potentials. This activity-dependent switch provides a mechanism for associative learning across different neocortical layers that process distinct types of information.     

NMDA Receptor-Dependent Long-Term Potentiation and Long-Term Depression (LTP/LTD)

Long-term potentiation and long-term depression (LTP/LTD) can be elicited by activating N-methyl-d-aspartate (NMDA)-type glutamate receptors, typically by the coincident activity of pre- and postsynaptic neurons. The early phases of expression are mediated by a redistribution of AMPA-type glutamate receptors: More receptors are added to potentiate the synapse or receptors are removed to weaken synapses. With time, structural changes become apparent, which in general require the synthesis of new proteins. The investigation of the molecular and cellular mechanisms underlying these forms of synaptic plasticity has received much attention, because NMDA receptor–dependent LTP and LTD may constitute cellular substrates of learning and memory.Long-term synaptic plasticity is a generic term that applies to a long-lasting experience-dependent change in the efficacy of synaptic transmission. Here we will focus on N-methyl-d-aspartate (NMDA) receptor–dependent synaptic potentiation (LTP) and depression (LTD), two forms of activity-dependent long-term changes in synaptic efficacy that have been extensively studied. Because both LTP and LTD are believed to represent cellular correlates of learning and memory, they have attracted considerable interest. In this article we will focus on the molecular and cellular mechanisms associated with LTP and LTD. As for other forms of long-term synaptic plasticity, a characterization of LTP and LTD involves describing the molecular mechanisms that are required to elicit the change (induction), followed by an investigation of the mechanism of expression (hours) and maintenance (days). The best-characterized form of NMDA receptor (NMDAR)-dependent LTP occurs between CA3 and CA1 pyramidal neurons of the hippocampus (Fig. 1). Throughout the chapter we will mostly refer to this specific form of LTP. At these CA3-CA1 Schaffer collateral synapses, the loci of both induction and expression are situated in the postsynaptic neuron.Open in a separate windowFigure 1.NMDAR-dependent LTD and LTP in the hippocampus. (A) Historical drawing by Ramon y Cajal (1909) of the trisynaptic pathway in the hippocampus. LTP and LTD are induced by activation of NMDARs at synapses between CA3 and CA1 pyramidal neurons (blue and red). In contrast, LTP at mossy fiber synapses onto CA3 neurons (green on blue) is NMDAR-independent. (B) This electron microscopy image shows the densely packed neuropil in the CA1 region of the hippocampus and highlights two asymmetric CA3-CA1 synapses. Note the typical “bouton en passant” configuration of synapse 1 and the prominent spine in synapse 2. The postsynaptic densities (PSDs) are visible. Scale bar, 200 nm. (Image kindly provided by Rafael Luján, Universitad de Castilla-La Mancha.) (C) Bidirectional change in CA3-CA1 synaptic efficacy by LTD and LTP in the same synapses monitored by extracellular field recordings in an acute slice preparation of the hippocampus. Note the contrasting induction protocols (Data from C Lüscher, unpubl.).