Protein Kinase Mζ Research Articles

Improving Old Memories

Can memory be improved? In vivo experiments using pharmacological inhibition of the enzyme protein kinase Mζ (PKMζ) in the insular cortex have shown that long-term conditioned taste aversion memory can be blocked. Now, Shema et al. (p. [1207][1]) show that overexpression of PKMζ in the rat insular cortex enhances memory, including memory that was formed long before the enzyme was overexpressed. The enhancement appears to affect more than a single memory. Thus, modulation of PKMζ can alter memories, even months after the initial encoding. [1]: /lookup/doi/10.1126/science.1200215

Enhancement of Consolidated Long-Term Memory by Overexpression of Protein Kinase Mζ in the Neocortex

Memories are more easily disrupted than improved. Many agents can impair memories during encoding and consolidation. In contrast, the armamentarium of potential memory enhancers is so far rather modest. Moreover, the effect of the latter appears to be limited to enhancing new memories during encoding and the initial period of cellular consolidation, which can last from a few minutes to hours after learning. Here, we report that overexpression in the rat neocortex of the protein kinase C isozyme protein kinase Mζ (PKMζ) enhances long-term memory, whereas a dominant negative PKMζ disrupts memory, even long after memory has been formed.

Metaplasticity governs compartmentalization of synaptic tagging and capture through brain-derived neurotrophic factor (BDNF) and protein kinase Mζ (PKMζ)

Activity-dependent synaptic plasticity is widely accepted to be the cellular correlate of learning and memory. It is believed that associativity between different synaptic inputs can transform short-lasting forms of synaptic plasticity (<3 h) to long-lasting ones. Synaptic tagging and capture (STC) might be able to explain this heterosynaptic support, because it distinguishes between local mechanisms of synaptic tags and cell-wide mechanisms responsible for the synthesis of plasticity-related proteins (PRPs). STC initiate storage processes only when the strength of the synaptic tag and the local concentration of essential proteins are above a certain plasticity threshold. We present evidence that priming stimulation through the activation of metabotropic glutamate receptors substantially increases the "range of threshold" for functional plasticity by producing protein kinase Mζ (PKMζ) as a PRP through local protein synthesis. In addition, our results implicate BDNF as a PRP which is mandatory for establishing cross-capture between synaptic strengthening and weakening, whereas the newly generated PKMζ specifically establishes synaptic tagging of long-term potentiation. Most intriguingly, we show here that STC are confined to specific dendritic compartments and that these compartments contain "synaptic clusters" with different plasticity thresholds. Our results suggest that within a dendritic compartment itself a homeostatic process exists to adjust plasticity thresholds. The range in which these clusters operate can be altered by processes of metaplasticity, which will operate on the cluster independently of other clusters at the same dendrite. These clusters will then prepare the synaptic network to form long-term memories.

Neuroplasticity of the Sensorimotor Cortex during Learning

We will discuss some of the current issues in understanding plasticity in the sensorimotor (SM) cortices on the behavioral, neurophysiological, and synaptic levels. We will focus our paper on reaching and grasping movements in the rat. In addition, we will discuss our preliminary work utilizing inhibition of protein kinase Mζ (PKMζ), which has recently been shown necessary and sufficient for the maintenance of long-term potentiation (LTP) (Ling et al., 2002). With this new knowledge and inhibitors to this system, as well as the ability to overexpress this system, we can start to directly modulate LTP and determine its influence on behavior as well as network level processing dependent at least in part due to this form of LTP. We will also briefly introduce the use of brain machine interface (BMI) paradigms to ask questions about sensorimotor plasticity and discuss current analysis techniques that may help in our understanding of neuroplasticity.

The protein kinase Mζ network as a bistable switch to store neuronal memory

BackgroundProtein kinase Mζ (PKMζ), the brain-specific, atypical protein kinase C isoform, plays a key role in long-term maintenance of memory. This molecule is essential for long-term potentiation of the neuron and various modalities of learning such as spatial memory and fear conditioning. It is unknown, however, how PKMζ stores information for long periods of time despite molecular turnover.ResultsWe hypothesized that PKMζ forms a bistable switch because it appears to constitute a positive feedback loop (PKMζ induces its local synthesis) part of which is ultrasensitive (PKMζ stimulates its synthesis through dual pathways). To examine this hypothesis, we modeled the biochemical network of PKMζ with realistic kinetic parameters. Bifurcation analyses of the model showed that the system maintains either the up state or the down state according to previous inputs. Furthermore, the model was able to reproduce a variety of previous experimental results regarding synaptic plasticity and learning, which suggested that it captures the essential mechanism for neuronal memory. We proposed in vitro and in vivo experiments that would critically examine the validity of the model and illuminate the pivotal role of PKMζ in synaptic plasticity and learning.ConclusionsThis study revealed bistability of the PKMζ network and supported its pivotal role in long-term storage of memory.

Adenylate cyclase 1 (ADCY1; AC1); protein kinase Mζ (PKMζ)

Adenylate cyclase 1 (ADCY1; AC1); protein kinase Mζ (PKMζ)

How does PKMζ maintain long-term memory?

Most of the molecular mechanisms contributing to long-term memory have been found to consolidate information within a brief time window after learning, but not to maintain information during memory storage. However, with the discovery that synaptic long-term potentiation is maintained by the persistently active protein kinase, protein kinase Mζ (PKMζ), a possible mechanism of memory storage has been identified. Recent research shows how PKMζ might perpetuate information both at synapses and during long-term memory.

Protein kinase Mζ is essential for the induction and maintenance of dopamine-induced long-term potentiation in apical CA1 dendrites

Dopaminergic D1/D5-receptor-mediated processes are important for certain forms of memory as well as for a cellular model of memory, hippocampal long-term potentiation (LTP) in the CA1 region of the hippocampus. D1/D5-receptor function is required for the induction of the protein synthesis-dependent maintenance of CA1-LTP (L-LTP) through activation of the cAMP/PKA-pathway. In earlier studies we had reported a synergistic interaction of D1/D5-receptor function and N-methyl-D-aspartate (NMDA)-receptors for L-LTP. Furthermore, we have found the requirement of the atypical protein kinase C isoform, protein kinase Mζ (PKMζ) for conventional electrically induced L-LTP, in which PKMζ has been identified as a LTP-specific plasticity-related protein (PRP) in apical CA1-dendrites. Here, we investigated whether the dopaminergic pathway activates PKMζ. We found that application of dopamine (DA) evokes a protein synthesis-dependent LTP that requires synergistic NMDA-receptor activation and protein synthesis in apical CA1-dendrites. We identified PKMζ as a DA-induced PRP, which exerted its action at activated synaptic inputs by processes of synaptic tagging.

Neurexin-Neuroligin Cell Adhesion Complexes Contribute to Synaptotropic Dendritogenesis via Growth Stabilization Mechanisms In Vivo

Cell adhesion molecules are well characterized for mediating synapse initiation, specification, differentiation, and maturation, yet their contribution to directing dendritic arborization during early brain circuit formation remains unclear. Using two-photon time-lapse imaging of growing neurons within intact and awake embryonic Xenopus brain, we examine roles of β-neurexin (NRX) and neuroligin-1 (NLG1) in dendritic arbor development. Using methods of dynamic morphometrics for comprehensive 3D quantification of rapid dendritogenesis, we find initial trans-synaptic NRX-NLG1 adhesions confer transient morphologic stabilization independent of NMDA receptor activity, whereas persistent stabilization requires NMDA receptor-dependent synapse maturation. Disrupting NRX-NLG1 function destabilizes filopodia while reducing synaptic density and AMPA receptor mEPSC frequency. Altered dynamic growth culminates in reduced dendritic arbor complexity as neurons mature over days. These results expand the synaptotropic model of dendritogenesis to incorporate cell adhesion molecule-mediated morphological stabilization necessary for directing normal dendritic arborization, providing a potential morphological substrate for developmental cognitive impairment associated with cell adhesion molecule mutations.

Erasing sensorimotor memories via PKMzeta inhibition.

Sensorimotor cortex has a role in procedural learning. Previous studies suggested that this learning is subserved by long-term potentiation (LTP), which is in turn maintained by the persistently active kinase, protein kinase Mzeta (PKMζ). Whereas the role of PKMζ in animal models of declarative knowledge is established, its effect on procedural knowledge is not well understood. Here we show that PKMζ inhibition, via injection of zeta inhibitory peptide (ZIP) into the rat sensorimotor cortex, disrupts sensorimotor memories for a skilled reaching task even after several weeks of training. The rate of relearning the task after the memory disruption by ZIP was indistinguishable from the rate of initial learning, suggesting no significant savings after the memory loss. These results indicate a shared molecular mechanism of storage for declarative and procedural forms of memory.

PKMζ Inhibition Reverses Learning-Induced Increases in Hippocampal Synaptic Strength and Memory during Trace Eyeblink Conditioning

A leading candidate in the process of memory formation is hippocampal long-term potentiation (LTP), a persistent enhancement in synaptic strength evoked by the repetitive activation of excitatory synapses, either by experimental high-frequency stimulation (HFS) or, as recently shown, during actual learning. But are the molecular mechanisms for maintaining synaptic potentiation induced by HFS and by experience the same? Protein kinase Mzeta (PKMζ), an autonomously active atypical protein kinase C isoform, plays a key role in the maintenance of LTP induced by tetanic stimulation and the storage of long-term memory. To test whether the persistent action of PKMζ is necessary for the maintenance of synaptic potentiation induced after learning, the effects of ZIP (zeta inhibitory peptide), a PKMζ inhibitor, on eyeblink-conditioned mice were studied. PKMζ inhibition in the hippocampus disrupted both the correct retrieval of conditioned responses (CRs) and the experience-dependent persistent increase in synaptic strength observed at CA3-CA1 synapses. In addition, the effects of ZIP on the same associative test were examined when tetanic LTP was induced at the hippocampal CA3-CA1 synapse before conditioning. In this case, PKMζ inhibition both reversed tetanic LTP and prevented the expected LTP-mediated deleterious effects on eyeblink conditioning. Thus, PKMζ inhibition in the CA1 area is able to reverse both the expression of trace eyeblink conditioned memories and the underlying changes in CA3-CA1 synaptic strength, as well as the anterograde effects of LTP on associative learning.

PKMζ maintains memories by regulating GluR2-dependent AMPA receptor trafficking

The maintenance of long-term memory in hippocampus, neocortex and amygdala requires the persistent action of the atypical protein kinase C isoform, protein kinase Mzeta (PKMzeta). We found that inactivating PKMzeta in the amygdala impaired fear memory in rats and that the extent of the impairment was positively correlated with a decrease in postsynaptic GluR2. Blocking the GluR2-dependent removal of postsynaptic AMPA receptors abolished the behavioral impairment caused by PKMzeta inhibition and the associated decrease in postsynaptic GluR2 expression, which correlated with performance. Similarly, blocking this pathway for removal of GluR2-containing receptors from postsynaptic sites in amygdala slices prevented the reversal of long-term potentiation caused by inactivating PKMzeta. Similar behavioral results were obtained in the hippocampus for unreinforced recognition memory of object location. Together, these findings indicate that PKMzeta maintains long-term memory by regulating the trafficking of GluR2-containing AMPA receptors, the postsynaptic expression of which directly predicts memory retention.

Functional Differences between and across Different Regions of the Apical Branch of Hippocampal CA1 Dendrites with Respect to Long-Term Depression Induction and Synaptic Cross-Tagging

The hippocampus is an ideal system to study synaptic plasticity in the context of learning and memory. The induction, expression, and interaction of long-term potentiation (LTP) as well as long-term depression (LTD) are essential elements for the functioning of complex networks in information processing and storage. Here we investigated whether different loci at the apical dendritic branch of CA1 pyramidal neurons are characterized by different capabilities to induce, express, and interact with LTP and LTD in hippocampal slices in vitro. We found that high-frequency stimulation resulted in longer-lasting forms of LTP in proximal and distal parts of the apical dendrites, whereas low-frequency stimulation induced longer-lasting LTD in distal but not at proximal parts. Interestingly, processes of "synaptic cross-tagging" could be described for any form of LTP transformation from early-stage LTP (E-LTP) into late-phase LTP (L-LTP) in distal and proximal parts, but for LTD, only at the distal part but not for the proximal part, although low-frequency stimulation at the proximal input, which resulted here only in a short-term depression, was paradoxically able to reinforce E-LTP into L-LTP at distal parts. We have identified protein kinase Mzeta (PKMzeta) as the LTP-specific, synthesized plasticity-related protein transforming E-LTP into L-LTP by strong low-frequency stimulation in the apical CA dendrite by cross-tagging mechanisms.

The protein kinase Mζ network as a bistable switch to store neuronal memory

Protein kinase Mζ (PKMζ), the brain-specific, atypical protein kinase C isoform, plays a key role in long-term maintenance of memory. This molecule is essential for long-term potentiation of the neuron and various modalities of learning such as spatial memory and fear conditioning. It is unknown, however, how PKMζ stores information for long periods of time despite molecular turnover. We hypothesized that PKMζ forms a bistable switch because it appears to constitute a positive feedback loop (PKMζ induces its local synthesis) part of which is ultrasensitive (PKMζ stimulates its synthesis through dual pathways). To examine this hypothesis, we modeled the biochemical network of PKMζ with realistic kinetic parameters. Bifurcation analyses of the model showed that the system maintains either the up state or the down state according to previous inputs. Furthermore, the model was able to reproduce a variety of previous experimental results regarding synaptic plasticity and learning, which suggested that it captures the essential mechanism for neuronal memory. We proposed in vitro and in vivo experiments that would critically examine the validity of the model and illuminate the pivotal role of PKMζ in synaptic plasticity and learning. This study revealed bistability of the PKMζ network and supported its pivotal role in long-term storage of memory.

PKMζ Restricts Dendritic Arbor Growth by Filopodial and Branch Stabilization within the Intact and Awake Developing Brain

The molecular mechanisms underlying activity-dependent neural circuit growth and plasticity during early brain development remain poorly understood. Protein kinase Mzeta (PKMz), an endogenous constitutively active kinase associated with late-phase long-term synaptic potentiation and memory in the mature brain, is expressed in the embryonic Xenopus retinotectal system with heightened levels during peak periods of dendrite growth and synaptogenesis. In vivo rapid time-lapse imaging of actively growing tectal neurons and comprehensive three-dimensional tracking of dynamic dendritic growth behavior finds that altered PKMz activity affects morphologic stabilization. Exogenous expression of PKMz within single neurons stabilizes dendritic filopodia by increasing dendritic filopodial lifetimes and decreasing filopodial additions, eliminations, and motility, whereas long-term in vivo imaging demonstrates restricted expansion of the dendritic arbor. Alternatively, blocking endogenous PKMz activity in individual growing tectal neurons with an inhibitory peptide (zeta-inhibitory peptide) destabilizes dendritic filopodia and over long periods promotes excessive arbor expansion. Furthermore, inhibiting endogenous PKMz throughout the tectum decreases colocalization of immunostained presynaptic and postsynaptic markers, SNAP-25 and PSD-95, respectively, suggesting impaired synapse maintenance. Together, these results implicate PKMz activity in restricting dendritic arborization during embryonic brain circuit development through synaptotropic stabilization of dynamic processes.

Separating kindling and LTP: Lessons from studies of PKM zeta in developing and adult rats

The kindling model of temporal lobe epilepsy (TLE) and the memory model of long-term potentiation (LTP) may have common underlying mechanisms. This is evident by the demonstration that certain signaling molecules play a key role in both. Recently, a brain specific isoform of protein kinase C (PKMζ) has been shown to play a significant role in both maintaining LTP and memory storage. We were interested in determining if this kinase had a crossover role in kindling-induced epileptogenesis. Using developing and adult rats we examined the role of PKMζ in kindling. In developing (P15) rats we determined the effect of PKMζ on retention of amygdala kindling and kindling rate by intra-amygdala administration of a selective PKMζ antagonist, ZIP (10nmol). In adult rats we examined the effect of PKMζ inhibition, ZIP (10nmol), on after discharge (AD) thresholds and kindling retention using rapid hippocampal kindling. Inhibition of PKMζ by the antagonist ZIP did not affect kindling rate or retention in developing rats. In addition there was also no observed effect on AD thresholds and kindling retention in adult rats. Our results show that, despite the similarities between kindling and LTP in their induction, there is dissociation in the role that PKMζ plays within the two in maintenance. This may be of importance in establishing a separation between the pathophysiological processes involved in sustaining kindling and the physiological mechanisms involved in maintaining LTP and memory storage.

Priming of short-term potentiation and synaptic tagging/capture mechanisms by ryanodine receptor activation in rat hippocampal CA1

Activity-dependent changes in synaptic strength such as long-term potentiation (LTP) and long-term depression (LTD) are considered to be cellular mechanisms underlying learning and memory. Strengthening of a synapse for a few seconds or minutes is termed short-term potentiation (STP) and is normally unable to take part in the processes of synaptic tagging/capture due to its inability to set the "synaptic tags." Here, we report that priming of synapses with ryanodine receptor agonists such as ryanodine (10 microM) or caffeine (10 mM) facilitates subsequent synaptic tagging/capture, enabling an STP protocol to establish a late-LTP in response to strong tetanization of a heterosynaptic input. We identified calcium/calmodulin-dependent protein kinase II (CaMKII) as mediating the primed synaptic tag setting, which persisted for 1 h. We also identified protein kinase Mzeta (PKMzeta), presumably captured from the strongly tetanized heterosynaptic input, as a plasticity-related protein maintaining the LTP at the tagged synapses. In addition, synaptic tags in primed STP were erased or interfered with by delivering low-frequency depotentiating stimulation 5 or 10 min after its induction, thus preventing capture of newly synthesized proteins. These data reveal a novel form of metaplasticity, whereby ryanodine receptor activation lowers the threshold for subsequent synaptic tagging/capture, thus priming weakly activated synapses for heterosynaptic interactions that promote long-term functional plasticity.

KIBRA Co-Localizes with Protein Kinase Mζ (PKMζ) in the Mouse Hippocampus

KIBRA is a WW domain-containing protein that can bind to protein kinase Czeta (PKCzeta). The SNP of the ninth intron of the KIBRA gene is associated with human episodic memory performance. Protein kinase Mzeta (PKMzeta), a brain-specific variant of PKCzeta, plays important roles in memory formation. Here we examined the interaction of KIBRA and PKMzeta in the adult mouse brain. Immunoprecipitation using newly-raised anti-KIBRA antibody revealed the interaction between KIBRA and PKMzeta in the brain. KIBRA was co-localized with PKMzeta in a single cultured neuron. Distribution analysis by immunohistochemistry and in situ hybridization indicated that KIBRA was highly localized with PKMzeta in the hippocampal CA1, CA3, and dentate gyrus. These results suggest that KIBRA functions in memory performance via interaction with PKMzeta.

A dynamical system analysis of a putative long-term memory trace, protein kinase Mζ

A dynamical system analysis of a putative long-term memory trace, protein kinase Mζ

Protein kinase Mzeta maintains fear memory in the amygdala but not in the hippocampus.

Recent work on the long-term stability of memory and synaptic plasticity has identified a potentially critical role for protein kinase Mzeta (PKMzeta). PKMzeta is a constitutively active, atypical isoform of protein kinase C that is believed to maintain long term potentiation at hippocampal synapses in vitro. In behaving animals, local inhibition of PKMzeta disrupts spatial memory in the hippocampus and conditioned taste aversion memory in the insular cortex. The role of PKMzeta in context fear memory is less clear. This study examined the role of PKMzeta in amygdala and hippocampal neurons following a standard fear conditioning protocol. The results indicate that PKMzeta inhibition in the amygdala, but not in the hippocampus, can disrupt fear memory. This suggests that PKMzeta may only maintain select forms of memory in specific brain structures and does not participate in a universal memory storage mechanism.