Raphael Lamprecht

Structural plasticity and memory.

Much evidence indicates that, after learning, memories are created by alterations in glutamate-dependent excitatory synaptic transmission. These modifications are then actively stabilized, over hours or days, by structural changes at postsynaptic sites on dendritic spines. The mechanisms of this structural plasticity are poorly understood, but recent findings are beginning to provide clues. The changes in synaptic transmission are initiated by elevations in intracellular calcium and consequent activation of second messenger signalling pathways in the postsynaptic neuron. These pathways involve intracellular kinases and GTPases, downstream from glutamate receptors, that regulate and coordinate both cytoskeletal and adhesion remodelling, leading to new synaptic connections. Rapid changes in cytoskeletal and adhesion molecules after learning contribute to short-term plasticity and memory, whereas later changes, which depend on de novo protein synthesis as well as the early modifications, seem to be required for the persistence of long-term memory.

Fear Memory Formation Involves p190 RhoGAP and ROCK Proteins through a GRB2-Mediated Complex

We used fear conditioning, which is known to alter synaptic efficacy in lateral amygdala (LA), to study molecular mechanisms underlying long-term memory. Following fear conditioning, the tyrosine phosphorylated protein p190 RhoGAP becomes associated with GRB2 in LA significantly more in conditioned than in control rats. RasGAP and Shc were also found to associate with GRB2 in LA significantly more in the conditioned animals. Inhibition of the p190 RhoGAP-downstream kinase ROCK in LA during fear conditioning impaired long- but not short-term memory. Thus, the p190 RhoGAP/ROCK pathway, which regulates the morphology of dendrites and axons during neural development, plays a central role, through a GRB2-mediated molecular complex, in fear memory formation in the lateral amygdala.

Fear conditioning drives profilin into amygdala dendritic spines

Changes in spine morphology may underlie memory formation, but the molecular mechanisms that subserve such alterations are poorly understood. Here we show that fear conditioning in rats leads to the movement of profilin, an actin polymerization–regulatory protein, into dendritic spines in the lateral amygdala and that these spines undergo enlargements in their postsynaptic densities (PSDs). A greater proportion of profilin-containing spines with enlarged PSDs could contribute to the enhancement of associatively induced synaptic responses in the lateral amygdala following fear learning.

A-kinase anchoring proteins in amygdala are involved in auditory fear memory

A-kinase anchoring proteins (AKAPs) constitute a family of scaffolding proteins that bind the regulatory subunits of protein kinase A (PKA). AKAP binding to PKA regulates the phosphorylation of various proteins, some of which have been implicated in synaptic plasticity and memory consolidation. Here we show that the regulatory subunits of PKA are colocalized with AKAP150 (an AKAP isoform that is expressed in the brain) in the lateral amygdala (LA) and that infusion to the LA of the peptide St-Ht31, which blocks PKA anchoring onto AKAPs, impairs memory consolidation of auditory fear conditioning.

Actin polymerization in lateral amygdala is essential for fear memory formation.

Actin polymerization is involved in key neuronal functions such as intracellular trafficking and morphogenesis. In this study, we examined the role of actin polymerization in lateral amygdala (LA) in fear conditioning memory formation. Microinfusion of cytochalasin D, an actin polymerization inhibitor, into rat LA immediately before fear conditioning training impaired the formation of long-term fear memory (LTM) but not short-term fear memory (STM). Microinfusion of cytochalasin D into rat LA immediately after fear conditioning impaired LTM. Cytochalasin D had no effect on fear conditioning memory retrieval when injected immediately before LTM test. These results show that actin cytoskeleton rearrangement is essential for fear memory consolidation.

Associative Pavlovian conditioning leads to an increase in spinophilin-immunoreactive dendritic spines in the lateral amygdala

Changes in dendritic spine number and shape are believed to reflect structural plasticity consequent to learning. Previous studies have strongly suggested that the dorsal subnucleus of the lateral amygdala is an important site of physiological plasticity in Pavlovian fear conditioning. In the present study, we examined the effect of auditory fear conditioning on dendritic spine numbers in the dorsal subnucleus of the lateral amygdala using an immunolabelling procedure to visualize the spine‐associated protein spinophilin. Associatively conditioned rats that received paired tone and shock presentations had 35% more total spinophilin‐immunoreactive spines than animals that had unpaired stimulation, consistent with the idea that changes in the number of dendritic spines occur during learning and account in part for memory.

The actin cytoskeleton 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 (LTM) formation involves alterations of synaptic efficacy produced by modifications in neural transmission and morphology. The actin cytoskeleton has been shown to be involved in these key neuronal processes by subserving events such as presynaptic vesicle movement, postsynaptic glutamate receptors trafficking and dendritic spines morphogenesis. Actin cytoskeleton dynamics and structure underlying such cellular events can be regulated by extracellular signals through its regulatory proteins. Recent findings show that the actin cytoskeleton and its regulatory proteins are needed for memory formation and extinction in different organisms throughout the phyla from invertebrates such as Caenorhabditis elegans and Drosophila to mammalians. The actin cytoskeleton and its regulatory proteins participate in the formation of various types of memories that are subserved by different neurons and brain regions. The actin cytoskeleton may therefore mediate between synaptic transmission during learning and long-term cellular alterations mandatory for memory formation.

Fear conditioning induces distinct patterns of gene expression in lateral amygdala

The lateral nucleus of the amygdala (LA) has been implicated in the formation of long‐term associative memory (LTM) of stimuli associated with danger through fear conditioning. The current study aims to detect genes that are expressed in LA following associative fear conditioning. Using oligonucleotide microarrays, we monitored gene expression in rats subjected to paired training where a tone co‐terminates with a footshock, or unpaired training where the tone and footshock are presented in a non‐overlapping manner. The paired protocol consistently leads to auditory fear conditioning memory formation, whereas the unpaired protocol does not. When the paired group was compared with the unpaired group 5 h after training, the expression of genes coding for the limbic system‐associated membrane protein (Lsamp), kinesin heavy chain member 2 (Kif2), N‐ethylmaleimide‐sensitive fusion protein (NSF) and Hippocalcin‐like 4 protein (Hpcal4) was higher in the paired group. These genes encode proteins that regulate neuronal axonal morphology (Lsamp, Kif2), presynaptic vesicle cycling and release (Hpcal4 and NSF), and AMPA receptor maintenance in synapses (NSF). Quantitative real‐time PCR (qPCR) showed that Kif2 and Lsamp are expressed hours following fear conditioning but minutes after unpaired training. Hpcal4 is induced by paired stimulation only 5 h after the training. These results show that fear conditioning induces a unique temporal activation of molecular pathways involved in regulating synaptic transmission and axonal morphology in LA, which is different from non‐associative stimulation.

Myosin light chain kinase regulates synaptic plasticity and fear learning in the lateral amygdala.

Learning and memory depend on signaling molecules that affect synaptic efficacy. The cytoskeleton has been implicated in regulating synaptic transmission but its role in learning and memory is poorly understood. Fear learning depends on plasticity in the lateral nucleus of the amygdala. We therefore examined whether the cytoskeletal-regulatory protein, myosin light chain kinase, might contribute to fear learning in the rat lateral amygdala. Microinjection of ML-7, a specific inhibitor of myosin light chain kinase, into the lateral nucleus of the amygdala before fear conditioning, but not immediately afterward, enhanced both short-term memory and long-term memory, suggesting that myosin light chain kinase is involved specifically in memory acquisition rather than in posttraining consolidation of memory. Myosin light chain kinase inhibitor had no effect on memory retrieval. Furthermore, ML-7 had no effect on behavior when the training stimuli were presented in a non-associative manner. Anatomical studies showed that myosin light chain kinase is present in cells throughout lateral nucleus of the amygdala and is localized to dendritic shafts and spines that are postsynaptic to the projections from the auditory thalamus to lateral nucleus of the amygdala, a pathway specifically implicated in fear learning. Inhibition of myosin light chain kinase enhanced long-term potentiation, a physiological model of learning, in the auditory thalamic pathway to the lateral nucleus of the amygdala. When ML-7 was applied without associative tetanic stimulation it had no effect on synaptic responses in lateral nucleus of the amygdala. Thus, myosin light chain kinase activity in lateral nucleus of the amygdala appears to normally suppress synaptic plasticity in the circuits underlying fear learning, suggesting that myosin light chain kinase may help prevent the acquisition of irrelevant fears. Impairment of this mechanism could contribute to pathological fear learning.

The Roles of the Actin Cytoskeleton in Fear Memory Formation

The formation and storage of fear memory is needed to adapt behavior and avoid danger during subsequent fearful events. However, fear memory may also play a significant role in stress and anxiety disorders. When fear becomes disproportionate to that necessary to cope with a given stimulus, or begins to occur in inappropriate situations, a fear or anxiety disorder exists. Thus, the study of cellular and molecular mechanisms underpinning fear memory may shed light on the formation of memory and on anxiety and stress related disorders. Evidence indicates that fear learning leads to changes in neuronal synaptic transmission and morphology in brain areas underlying fear memory formation including the amygdala and hippocampus. The actin cytoskeleton has been shown to participate in these key neuronal processes. Recent findings show that the actin cytoskeleton is needed for fear memory formation and extinction. Moreover, the actin cytoskeleton is involved in synaptic plasticity and in neuronal morphogenesis in brain areas that mediate fear memory. The actin cytoskeleton may therefore mediate between synaptic transmission during fear learning and long-term cellular alterations mandatory for fear memory formation.