EphB2 activation in neural stem cells in the basolateral amygdala facilitates neurogenesis and enhances long-term memory.

Many brain diseases lead to a reduction in the number of functional neurons and it would be of value to be able to increase the number of neurons in the affected brain areas. In this study, we examined whether we can promote neural stem cells to produce mature neurons and whether an increase in the mature neurons can affect cognitive performance. We detected that the EphB2 receptor is localized in immature basolateral amygdala (BLA) neurons. We therefore aimed to increase the level of EphB2 activity in neural stem cells (NSCs) in the BLA and examine the effects on the production of mature neurons and cognition. Toward that end, we utilized a photoactivatable EphB2 construct (optoEphB2) to increase EphB2 forward signaling in NSCs in the BLA. We revealed that the activation of optoEphB2 in NSCs in the BLA increased the level of immature and mature neurons in the BLA. We further found that activation of optoEphB2 in BLA NSCs enhanced auditory, but not contextual, long-term fear memory formation. Impairing EphB2 forward signaling did not affect the level of immature and mature neurons in the BLA. This study provides evidence that NSCs can be promoted to produce mature neurons by activating EphB2 to enhance specific brain functions.

Learning-induced bidirectional enhancement of inhibitory synaptic metaplasticity.

Training rodents in a particularly difficult olfactory-discrimination (OD) task results in the acquisition of the ability to perform the task well, termed 'rule learning'. In addition to enhanced intrinsic excitability and synaptic excitation in piriform cortex pyramidal neurons, rule learning results in increased synaptic inhibition across the whole cortical network to the point where it precisely maintains the balance between inhibition and excitation. The mechanism underlying such precise inhibitory enhancement remains to be explored. Here, we use brain slices from transgenic mice (VGAT-ChR2-EYFP), enabling optogenetic stimulation of single GABAergic neurons and recordings of unitary synaptic events in pyramidal neurons. Quantal analysis revealed that learning-induced enhanced inhibition is mediated by increased quantal size of the evoked inhibitory events. Next, we examined the plasticity of synaptic inhibition induced by long-lasting, intrinsically evoked spike firing in post-synaptic neurons. Repetitive depolarizing current pulses from depolarized (-70 mV) or hyperpolarized (-90 mV) membrane potentials induced long-term depression (LTD) and long-term potentiation (LTP) of synaptic inhibition, respectively. We found a profound bidirectional increase in the ability to induce both LTD, mediated by L-type calcium channels, and LTP, mediated by R-type calcium channels after rule learning. Blocking the GABAB receptor reversed the effect of intrinsic stimulation at -90 mV from LTP to LTD. We suggest that learning greatly enhances the ability to modify the strength of synaptic inhibition of principal neurons in both directions. Such plasticity of synaptic plasticity allows fine-tuning of inhibition on each particular neuron, thereby stabilizing the network while maintaining the memory of the rule. KEY POINTS: Olfactory discrimination rule learning results in long-lasting enhancement of synaptic inhibition on piriform cortex pyramidal neurons. Quantal analysis of unitary inhibitory synaptic events, evoked by optogenetic minimal stimulation, revealed that enhanced synaptic inhibition is mediated by increased quantal size. Surprisingly, metaplasticity of synaptic inhibition, induced by intrinsically evoked repetitive spike firing, is increased bidirectionally. The susceptibility to both long-term depression (LTD) and long-term potentiation (LTP) of inhibition is enhanced after learning. LTD of synaptic inhibition is mediated by L-type calcium channels and LTP by R-type calcium channels. LTP is also dependent on activation of GABAB receptors. We suggest that learning-induced changes in the metaplasticity of synaptic inhibition enable the fine-tuning of inhibition on each particular neuron, thereby stabilizing the network while maintaining the memory of the rule.

The role of p21-activated kinase in maintaining the fear learning-induced modulation of excitation/inhibition ratio in lateral amygdala.

We study the relations between different learning paradigms and enduring changes in excitatory synaptic transmission. Here we show that auditory fear conditioning (AFC), but not olfactory fear conditioning (OFC) training, led to enduring enhancement in AMPA-mediated miniature EPSCs (mEPSCs). Moreover, olfactory unpaired training led to a stable significant reduction in excitatory synaptic transmission. However, olfactory discrimination learning (OD) did not modulate postsynaptic AMPA-mediated mEPSCs in LA. The p21-activated kinase (PAK) activity, previously shown to have a key role in maintaining persistent long-lasting enhancement in synaptic inhibition after OFC, has an opposing effect on excitatory synaptic transmission. PAK maintained the level of excitatory synaptic transmission in the amygdala in all experimental groups, except in neurons in the OFC trained rats. PAK also maintained excitatory synaptic transmission in all neurons of auditory fear conditioning and naïve training groups except in neurons of the auditory safety learning. Safety learning was previously shown in our study to enhance synaptic inhibition. We thus suggest that PAK maintains inhibitory synaptic transmission in a learning-dependent manner and on the other hand affects excitatory synaptic transmission only in groups where learning has not affected inhibitory transmission. Thus, PAK controls learning-induced changes in the excitation/inhibition balance.

A Quasi-Static Quantitative Ultrasound Elastography Algorithm Using Optical Flow.

Ultrasound elastography is a constantly developing imaging technique which is capable of displaying the elastic properties of tissue. The measured characteristics could help to refine physiological tissue models, but also indicate pathological changes. Therefore, elastography data give valuable insights into tissue properties. This paper presents an algorithm that measures the spatially resolved Young's modulus of inhomogeneous gelatin phantoms using a CINE sequence of a quasi-static compression and a load cell measuring the compressing force. An optical flow algorithm evaluates the resulting images, the stresses and strains are computed, and, conclusively, the Young's modulus and the Poisson's ratio are calculated. The whole algorithm and its results are evaluated by a performance descriptor, which determines the subsequent calculation and gives the user a trustability index of the modulus estimation. The algorithm shows a good match between the mechanically measured modulus and the elastography result-more precisely, the relative error of the Young's modulus estimation with a maximum error 35%. Therefore, this study presents a new algorithm that is capable of measuring the elastic properties of gelatin specimens in a quantitative way using only the image data. Further, the computation is monitored and evaluated by a performance descriptor, which measures the trustability of the results.

Dynamic Biomechanical Analysis of Vocal Folds Using Pipette Aspiration Technique.

The voice producing process is a complex interplay between glottal pressure, vocal folds, their elasticity and tension. The material properties of vocal folds are still insufficiently studied, because the determination of material properties in soft tissues is often difficult and connected to extensive experimental setups. To shed light on this less researched area, in this work, a dynamic pipette aspiration technique is utilized to measure the elasticity in a frequency range of 100-1000 Hz. The complex elasticity could be assessed with the phase shift between exciting pressure and tissue movement. The dynamic pipette aspiration setup has been miniaturized with regard to a future in-vivo application. The techniques were applied on 3 different porcine larynges 4 h and 1 d postmortem, in order to investigate the deterioration of the tissue over time and analyze correlation in elasticity values between vocal fold pairs. It was found that vocal fold pairs do have different absolute elasticity values but similar trends. This leads to the assumption that those trends are more important for phonation than having same absolute values.

Learning-induced enduring changes in inhibitory synaptic transmission in lateral amygdala are mediated by p21-activated kinase.

In this study we explored whether learning leads to enduring changes in inhibitory synaptic transmission in lateral amygdala (LA). We revealed that olfactory discrimination (OD) learning in rats led to a long-lasting increase in postsynaptic GABAA channel-mediated miniature inhibitory postsynaptic currents (mIPSCs) in LA. Olfactory fear conditioning, but not auditory fear conditioning, also led to enduring enhancement in GABAA-mediated mIPSCs. Auditory fear conditioning, but not olfactory fear conditioning or OD learning, induced an enduring reduction in the frequency but not the current of mIPSC events. We found that p21-activated kinase (PAK) activity is needed to maintain OD and olfactory fear conditioning learning-induced enduring enhancement of mIPSCs. Further analysis revealed that OD led to an increase in GABAA channel conductance whereas olfactory fear conditioning increased the number of GABAA channels. These alterations in GABAA channels conductance and level are controlled by PAK activity. Our study shows that the learning-induced increase in postsynaptic inhibitory transmission in LA is specific to the sensory modality. However, the mechanism that mediates the increase in inhibitory transmission, namely the increase in the conductance or in the level of GABAA channel, is learning specific.NEW & NOTEWORTHY Here we studied whether learning leads to long-lasting alterations in inhibitory synaptic transmission in lateral amygdala (LA). We revealed that learning led to enduring changes in inhibitory synaptic transmission in LA that are affected by the sensory modality (auditory or olfaction) used during learning. However, the mechanism that mediated the changes in inhibitory transmission (alterations in GABAA channel level or conductance) depended on the type of learning. These long-lasting alterations are maintained by p21-activated kinase.

Long-term memory is maintained by continuous activity of Arp2/3 in lateral amygdala.

Evidence indicates that long-term memory formation involves alterations in synaptic efficacy produced by modifications in neural transmission and morphology. However, it is not clear how such changes induced by learning, that encode memory, are maintained over long period of time to preserve long-term memory. It has been shown that the actin nucleating protein Arp2/3 is essential for supporting neuronal morphology and synaptic transmission. We therefore hypothesized that continuous Arp2/3 activity is needed to maintain long-term memory over time. To test this hypothesis we microinjected into lateral amygdala (LA) of rats CK-666, a specific inhibitor of Arp2/3, two days after fear conditioning and tested the effect on long-term fear memory maintenance a day afterward. We found that injection of CK-666 two days after training abolished fear conditioning memory. Fear conditioning could be formed when a control compound CK-689 was applied two days after training. Microinjection of CK-666 a day before fear conditioning training had no effect on fear conditioning learning and long-term memory formation. We revealed that Arp2/3 is also needed to maintain long-term conditioned taste aversion (CTA) memory in LA. Microinjection of CK-666 two days after CTA training impaired long-term memory tested a day afterwards. We conclude that continuous activity of Arp2/3 in LA is essential for the maintenance of long-term memory.

Learning-induced modulation of the effect of endocannabinoids on inhibitory synaptic transmission.

are key modulators that regulate central brain functions and behaviors, including learning and memory. At the cellular and molecular levels, endocannabinoids are potent modulators of excitatory and inhibitory synaptic function. Most effects of cannabinoids are thought to be mediated via G protein-coupled cannabinoid receptors. In particular, cannabinoids released from postsynaptic neurons are suggested to act as retrograde messengers, activating presynaptic type-1 cannabinoid receptors (CB1Rs), thereby inducing suppression of synaptic release. Another central mechanism of cannabinoid-induced action requires activation of astroglial CB1Rs. CB1Rs are also implicated in self-modulation of cortical neurons. Rats that are trained in a particularly difficult olfactory-discrimination task show a dramatic increased ability to acquire memories of new odors. The memory of the acquired high-skill acquisition, termed "rule learning" or "learning set," lasts for many months. Using this behavioral paradigm, we show a novel function of action for CB1Rs, supporting long-term memory by maintaining persistent enhancement of inhibitory synaptic transmission. Long-lasting enhancement of inhibitory synaptic transmission is blocked by a CB1R inverse agonist. This effect is mediated by a novel purely postsynaptic mechanism, obtained by enhancing the single GABAA channel conductance that is PKA dependent. The significant role that CB1R has in maintaining learning-induced long-term strengthening of synaptic inhibition suggests that endocannabinoids have a key role in maintaining long-term memory by enhancing synaptic inhibition. NEW & NOTEWORTHY In this study we show a novel function and mechanism of action for cannabinoids in neurons, mediated by activation of type-1 cannabinoid receptors, supporting long-term memory by maintaining persistent enhancement of inhibitory synaptic transmission on excitatory neurons. This effect is mediated by a novel purely postsynaptic mechanism, obtained by enhancing the single GABAA channel conductance that is PKA dependent. Thus we report for the first time that endocannabinoids have a key role maintaining learning-induced synaptic modification.

Persistent CaMKII activation mediates learning-induced long-lasting enhancement of synaptic inhibition.

Training rats in a particularly difficult olfactory-discrimination task results in acquisition of high skill to perform the task superbly, termed "rule learning" or "learning set." Such complex learning results in enhanced intrinsic neuronal excitability of piriform cortex pyramidal neurons, and in their excitatory synaptic interconnections. These changes, while subserving memory maintenance, must be counterbalanced by modifications that prevent overspreading of activity and uncontrolled synaptic strengthening. Indeed, we have previously shown that the average amplitude of GABAA-mediated miniature IPSCs (mIPSCs) in these neurons is enhanced for several days after learning, an enhancement mediated via a postsynaptic mechanism. To unravel the molecular mechanism of this long-term inhibition enhancement, we tested the role of key second-messenger systems in maintaining such long-lasting modulation. The calcium/calmodulin-dependent kinase II (CaMKII) blocker, KN93, significantly reduced the average mIPSC amplitude in neurons from trained rats only to the average pretraining level. A similar effect was obtained by the CaMKII peptide inhibitor, tatCN21. Such reduction resulted from decreased single-channel conductance and not in the number of activated channels. The PKC inhibitor, GF109203X, reduced the average mIPSC amplitude in neurons from naive, pseudo-trained, and trained animals, and the difference between the trained and control groups remained. Such reduction resulted from a decrease in the number of activated channels. The PKA inhibitor H89 dihydrochloride did not affect the average mIPSC amplitude in neurons from any of the three groups. We conclude that learning-induced enhancement of GABAA-mediated synaptic inhibition is maintained by persistent CaMKII activation.

Calcium/calmodulin-dependent kinase II activity is required for maintaining learning-induced enhancement of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor-mediated synaptic excitation.

Learning leads to changes in α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR)-mediated synaptic excitation. The mechanisms for maintaining such alterations needed for memory persistence remain to be clarified. Here, we report a novel molecular mechanism for maintaining learning-induced AMPAR-mediated enhancement of synaptic excitation. We show that training rats in a complex olfactory discrimination task, such that requires rule learning, leads to the enhancement of averaged amplitude of AMPAR-mediated miniature excitatory post-synaptic currents (mEPSCs) in piriform cortex pyramidal neurons for days after learning. Inhibiting calcium/calmodulin-dependent kinase II (CaMKII) using KN93 or tatCN21 days after learning, reduced the averaged mEPSC amplitude in neurons in piriform cortex of trained rats to the level where they are not significantly different from mEPSC of control animals. CaMKII inhibition leads to a decrease in single channel conductance and not to changes in the number of synaptic-activated channels. We conclude that the maintenance of learning-induced enhancement of AMPAR-mediated synaptic excitation requires the activity of calcium/calmodulin-dependent kinase II. We show that training rats in a complex olfactory discrimination task leads to the enhancement of averaged amplitude of AMPA receptor-mediated miniature excitatory post-synaptic currents (mEPSCs) in piriform cortex pyramidal neurons for days after learning. Inhibiting calcium/calmodulin-dependent kinase II (CaMKII) using KN93 or tatCN21 days after learning, reduced the averaged mEPSC amplitude in neurons in piriform cortex of trained rats to the level where they are not significantly different from mEPSC of control animals. CaMKII inhibition leads to a decrease in AMPAR single channel conductance. We conclude that the maintenance of learning-induced enhancement of AMPAR-mediated synaptic excitation requires the activity of CaMKII.

The Role of Ephs and Ephrins in Memory Formation.

The ability to efficiently store memories in the brain is a fundamental process and its impairment is associated with multiple human mental disorders. Evidence indicates that long-term memory formation involves alterations of synaptic efficacy produced by modifications in neural transmission and morphology. The Eph receptors and their cognate ephrin ligands have been shown to be involved in these key neuronal processes by regulating events such as presynaptic transmitter release, postsynaptic glutamate receptor conductance and trafficking, synaptic glutamate reuptake, and dendritic spine morphogenesis. Recent findings show that Ephs and ephrins are needed for memory formation in different organisms. These proteins participate in the formation of various types of memories that are subserved by different neurons and brain regions. Ephs and ephrins are involved in brain disorders and diseases with memory impairment symptoms, including Alzheimer's disease and anxiety. Drugs that agonize or antagonize Ephs/ephrins signaling have been developed and could serve as therapeutic agents to treat such diseases. Ephs and ephrins may therefore induce cellular alterations mandatory for memory formation and serve as a target for pharmacological intervention for treatment of memory-related brain diseases.

Fear conditioning leads to alteration in specific genes expression in cortical and thalamic neurons that project to the lateral amygdala.

RNA transcription is needed for memory formation. However, the ability to identify genes whose expression is altered by learning is greatly impaired because of methodological difficulties in profiling gene expression in specific neurons involved in memory formation. Here, we report a novel approach to monitor the expression of genes after learning in neurons in specific brain pathways needed for memory formation. In this study, we aimed to monitor gene expression after fear learning. We retrogradely labeled discrete thalamic neurons that project to the lateral amygdala (LA) of rats. The labeled neurons were dissected, using laser microdissection microscopy, after fear conditioning learning or unpaired training. The RNAs from the dissected neurons were subjected to microarray analysis. The levels of selected RNAs detected by the microarray analysis to be altered by fear conditioning were also assessed by nanostring analysis. We observed that the expression of genes involved in the regulation of translation, maturation and degradation of proteins was increased 6 h after fear conditioning compared to unpaired or naïve trained rats. These genes were not expressed 24 h after training or in cortical neurons that project to the LA. The expression of genes involved in transcription regulation and neuronal development was altered after fear conditioning learning in the cortical-LA pathway. The present study provides key information on the identity of genes expressed in discrete thalamic and cortical neurons that project to the LA after fear conditioning. Such an approach could also serve to identify gene products as targets for the development of a new generation of therapeutic agents that could be aimed to functionally identified brain circuits to treat memory-related disorders.

The roles of Eph receptors in contextual fear conditioning memory formation.

Eph receptors regulate glutamate receptors functions, neuronal morphology and synaptic plasticity, cellular events believed to be involved in memory formation. In this study we aim to explore the roles of Eph receptors in learning and memory. Toward that end, we examined the roles of EphB2 and EphA4 receptors, key regulators of synaptic functions, in fear conditioning memory formation. We show that mice lacking EphB2 (EphB2(-/-)) are impaired in short- and long-term contextual fear conditioning memory. Mice that express a carboxy-terminally truncated form of EphB2 that lacks forward signaling, instead of the full EphB2, are impaired in long-term, but not short-term, contextual fear conditioning memory. Long-term contextual fear conditioning memory is attenuated in CaMKII-cre;EphA4(lx/-) mice where EphA4 is removed from all pyramidal neurons of the forebrain. Mutant mice with targeted kinase-dead EphA4 (EphA4(KD)) exhibit intact long-term contextual fear conditioning memory showing that EphA4 kinase-mediated forward signaling is not needed for contextual fear memory formation. The ability to form long-term conditioned taste aversion (CTA) memory is not impaired in the EphB2(-/-) and CaMKII-cre;EphA4(lx/-) mice. We conclude that EphB2 forward signaling is required for long-term contextual fear conditioning memory formation. In contrast, EphB2 mediates short-term contextual fear conditioning memory formation in a forward signaling-independent manner. EphA4 mediates long-term contextual fear conditioning memory formation in a kinase-independent manner.

What kind of phonation causes the strongest vocal fold collision? - A hemi-larynx phonation contact pressure study.

This paper presents a measurement setup which is able to measure the distribution of small scale pressure on an area of 15.2 mm × 30.4 mm with a sample rate up to 1.2 kHz. It was used to investigate the contact pressures of vocal folds during phonation. This was performed in ex vivo experiments of 11 porcine larynges. The contact pressure at the medial surface and other phonation parameters, as the glottal resistance and the closing velocity of the vocal fold, were measured at different adduction and elongation levels and air flow rates. A statistical analysis was carried out. It could be shown that the contact pressure rises, when the vocal fold is manipulated or when the flow rate is increased.

Fear Conditioning Leads to Enduring Alterations in RNA Transcripts in Hippocampal Neuropil that are Dependent on EphB2 Forward Signaling.

Alterations in mRNA transcription have been associated with changes in brain functions. We wanted to examine if fear conditioning causes long-term changes in transcriptome profiles in the basolateral amygdala (BLA) and hippocampus using RNA-Seq and laser microdissection microscopy. We further aimed to uncover whether these changes are involved in memory formation by monitoring their levels in EphB2lacZ/lacZ mice, which lack EphB2 forward signaling and can form short-term fear conditioning memory but not long-term fear conditioning memory. We found transcriptome signatures unique to each brain region that are comprise of specific cellular pathways. We also revealed that fear conditioning leads to alterations in mRNAs levels 24 h after training in hippocampal neuropil, but not in hippocampal cell layers or BLA. The two main groups of altered mRNAs encode proteins involved in neuronal transmission, neuronal morphogenesis and neuronal development and the vast majority are known to be enriched in neurons. None of these mRNAs levels were altered by fear conditioning in EphB2lacZ/lacZ mice, which were also impaired in long-term fear memory. We show here that fear conditioning leads to an enduring alteration in mRNAs levels in hippocampal neuropil that is dependent on processes mediated by EphB2 that are needed for long-term memory formation.

Rac1 GTPase activation impairs fear conditioning-induced structural changes in basolateral amygdala neurons and long-term fear memory formation.

Long-term memory formation leads to enduring alterations in synaptic efficacy and neuronal responses that may be created by changes in neuronal morphology. We show that fear conditioning leads to a long-lasting increase in the volume of the primary and secondary dendritic branches, but not of distal branches, of neurons located at the basolateral amygdala (BLA). The length of the dendritic branches is not affected by fear conditioning. Fear conditioning leads to an enduring increase in the length and volume of dendritic spines, especially in the length of the spine neck and the volume of the spine head. Fear conditioning does not affect dendritic spine density. We further reveal that activation of Rac1 in BLA during fear conditioning impairs long-term auditory, but not contextual, fear conditioning memory. Activation of Rac1 during fear conditioning prevents the enduring increase in the dendritic primary branch volume and dendritic spines length and volume. Rac1 activation per se has no effect on neuronal morphology. These results show that fear conditioning induces changes known to reduce the inhibition of signal propagation along the dendrite and the increase in synaptic efficacy whereas preventing these changes, by Rac1 activation, impairs fear memory formation.

Behind the Complex Interplay of Phonation: Investigating Elasticity of Vocal Folds With Pipette Aspiration Technique During Ex Vivo Phonation Experiments.

OBJECTIVES: The vibration of the vocal folds produces the primary sound for the human speech. The vibration depends mainly on the pressure, airflow of the lungs, and the material properties of the vocal folds. In order to change them, muscles in the larynx stretch the vocal folds. This interplay is rarely investigated, but can give insight in the complex process of speech production. Most material properties studies are damaging the tissue; therefore, a nondestructive one is desired. METHODS: An ex vivo phonation experiment combined with the dynamic Pipette Aspiration Technique is used to investigate 10 porcine larynges, under manipulations of different adduction and elongation levels. For each manipulation, the near surface material properties of the vocal folds are measured as well as different phonation parameters like the subglottal pressure, glottal resistance, frequency, and stiffness. Thereby, a high-speed camera was used to record the vocal fold movement. RESULTS: On most of the measured parameters, the manipulations do show an effect. Both manipulations lead to a higher phonation frequency and an increase of the stiffness of the tissue. Comparing both manipulations, the elongation results in higher elasticity values than the adduction. Different measurement parameters have been compared with each other and correlations could be found. Where the strongest correlation are found among the elasticity values of different frequencies. But it can also be seen that the elasticity values correlate with phonation parameters. CONCLUSION: It was possible to produce a data set of 560 measurements in total. To our knowledge, this is the first time Pipette Aspiration Technique was combined with ex vivo phonation measurements for combined measurements. The amount of measurement data made it possible to carry out statistic investigations. The effect of the manipulations on material properties as well as on phonation parameters could be measured and different correlations could be found. The results lead to the hypothesis that the stretch does not have a huge effect on the material properties of the lamina propria, but more on the underlying muscle.

Rho-associated kinase in the gustatory cortex is involved in conditioned taste aversion memory formation but not in memory retrieval or relearning.

Rho-associated kinase (ROCK) is intimately involved in cortical neuronal morphogenesis. The present study explores the roles of ROCK in conditioned taste aversion (CTA) memory formation in gustatory cortex (GC) in adult rat. Microinjection of the ROCK inhibitor Y-27632 into the GC 30 min before CTA training or 10 min after the conditioned stimulus (CS) impaired long-term CTA memory (LTM) formation. ROCK inhibitor had no effect on taste aversion when injected before the first LTM test day and did not alter taste aversion on subsequent test days. Microinjection of ROCK inhibitor into GC 30 min before preexposure to the taste CS had no effect on latent inhibition of CTA learning suggesting that ROCK is involved in CS-US association rather than taste learning per se. Cumulatively, these results show that ROCK is needed for normal CTA memory formation but not retrieval, relearning or incidental taste learning.

EphrinA4 mimetic peptide impairs fear conditioning memory reconsolidation in lateral amygdala.

Fear memory may undergo a process after memory reactivation called reconsolidation. To examine the roles of ephrinA4 in fear memory reconsolidation an inhibitory ephrinA4 mimetic peptide (pep-ephrinA4), that targets the EphA binding site and inhibits EphA activation, was used. Pep-ephrinA4 was microinjected into the lateral amygdala (LA) of fear-conditioned rats 24 h after training and 30 min before tone CS memory retrieval. Memory retrieval was unaffected by pep-ephrinA4. However, the animals were impaired in fear memory tested 1 h or 24 h afterward when compared to controls. Fear-conditioned animals injected with pep-ephrinA4 into LA immediately after long-term memory retrieval were unaffected when tested 24 h afterward. Microinjection into LA of a peptide originated from an ephrinA4 site that does not interact with EphA did not affect fear memory reconsolidation. Rats that were administrated with pep-ephrinA4 systemically 24 h after fear conditioning and 30 min before CS memory retrieval were impaired in long-term fear conditioning memory tested 24 h afterward when compared to the control peptide. These results show that ephrinA4 binding sites are needed for long-term fear memory reconsolidation in LA and may serve as a target for the treatment of fear-related disorders by blocking reconsolidation.

Nck1 activity in lateral amygdala regulates long-term fear memory formation.

Fear conditioning leads to long-term fear memory formation and is a model for studying fear-related psychopathological conditions such as phobias and post-traumatic stress disorder. Long-term fear memory formation is believed to involve alterations of synaptic efficacy mediated by changes in synaptic transmission and morphology in lateral amygdala (LA). Nck1 is a key neuronal adaptor protein involved in the regulation of the actin cytoskeleton and the neuronal processes believed to be involved in memory formation. However, the role of Nck1 in memory formation is not known. Here we explored the role of Nck1 in fear memory formation in lateral amygdala (LA). Reduction of Nck1 in excitatory neurons in LA enhanced long-term, but not short-term, auditory fear conditioning memory. Activation of Nck1, by using a photoactivatable Nck1 (PA-Nck1), during auditory fear conditioning in excitatory neurons in LA impaired long-term, but not short-term, fear memory. Activation of Nck1 immediately or a day after fear conditioning did not affect fear memory. The hippocampal-mediated contextual fear memory was not affected by the reduction or activation of Nck1 in LA. We show that Nck1 is localized to the presynapses in LA. Nck1 activation in LA excitatory neurons decreased the frequency of AMPA receptors-mediated miniature excitatory synaptic currents (mEPSCs). Nck1 activation did not affect GABA receptor-mediated inhibitory synaptic currents (mIPSCs). These results show that Nck1 activity in excitatory neurons in LA regulates glutamate release and sets the threshold for fear memory formation. Moreover, our research shows that Nck1 may serve as a target for pharmacological treatment of fear and anxiety disorders.