JIP1-Mediated JNK Activation Negatively Regulates Synaptic Plasticity and Spatial Memory.

The c-Jun N-terminal kinase (JNK) signal transduction pathway is implicated in learning and memory. Here, we examined the role of JNK activation mediated by the JNK-interacting protein 1 (JIP1) scaffold protein. We compared male wild-type mice with a mouse model harboring a point mutation in the Jip1 gene that selectively blocks JIP1-mediated JNK activation. These male mutant mice exhibited increased NMDAR currents, increased NMDAR-mediated gene expression, and a lower threshold for induction of hippocampal long-term potentiation. The JIP1 mutant mice also displayed improved hippocampus-dependent spatial memory and enhanced associative fear conditioning. These results were confirmed using a second JIP1 mutant mouse model that suppresses JNK activity. Together, these observations establish that JIP1-mediated JNK activation contributes to the regulation of hippocampus-dependent, NMDAR-mediated synaptic plasticity and learning.SIGNIFICANCE STATEMENT The results of this study demonstrate that c-Jun N-terminal kinase (JNK) activation induced by the JNK-interacting protein 1 (JIP1) scaffold protein negatively regulates the threshold for induction of long-term synaptic plasticity through the NMDA-type glutamate receptor. This change in plasticity threshold influences learning. Indeed, mice with defects in JIP1-mediated JNK activation display enhanced memory in hippocampus-dependent tasks, such as contextual fear conditioning and Morris water maze, indicating that JIP1-JNK constrains spatial memory. This study identifies JIP1-mediated JNK activation as a novel molecular pathway that negatively regulates NMDAR-dependent synaptic plasticity and memory.

Tone identification behavior in Rattus norvegicus: muscarinic receptor blockage lowers responsiveness in nontarget selective neurons, while nicotinic receptor blockage selectively lowers target responses.

Learning to associate a stimulus with reinforcement causes plasticity in primary sensory cortex. Neural activity caused by the associated stimulus is paired with reinforcement, but population analyses have not found a selective increase in response to that stimulus. Responses to other stimuli increase as much as, or more than, responses to the associated stimulus. Here, we applied population analysis at a new time point and additionally evaluated whether cholinergic receptor blockers interacted with the plastic changes in cortex. Three days of tone identification behavior caused responsiveness to increase broadly across primary auditory cortex, and target responses strengthened less than overall responsiveness. In pharmacology studies, behaviorally impairing doses of selective acetylcholine receptor blockers were administered during behavior. Neural responses were evaluated on the following day, while the blockers were absent. The muscarinic group, blocked by scopolamine, showed lower responsiveness and an increased response to the tone identification target that exceeded both the 3-day control group and task-naïve controls. Also, a selective increase in the late phase of the response to the tone identification stimulus emerged. Nicotinic receptor antagonism, with mecamylamine, more modestly lowered responses the following day and lowered target responses more than overall responses. Control acute studies demonstrated the muscarinic block did not acutely alter response rates, but the nicotinic block did. These results lead to the hypothesis that the decrease in the proportion of the population spiking response that is selective for the target may be explained by the balance between effects modulated by muscarinic and nicotinic receptors.

Arc expression and neuroplasticity in primary auditory cortex during initial learning are inversely related to neural activity.

Models of learning-dependent sensory cortex plasticity require local activity and reinforcement. An alternative proposes that neural activity involved in anticipation of a sensory stimulus, or the preparatory set, can direct plasticity so that changes could occur in regions of sensory cortex lacking activity. To test the necessity of target-induced activity for initial sensory learning, we trained rats to detect a low-frequency sound. After learning, Arc expression and physiologically measured neuroplasticity were strong in a high-frequency auditory cortex region with very weak target-induced activity in control animals. After 14 sessions, Arc and neuroplasticity were aligned with target-induced activity. The temporal and topographic correspondence between Arc and neuroplasticity suggests Arc may be intrinsic to the neuroplasticity underlying perceptual learning. Furthermore, not all neuroplasticity could be explained by activity-dependent models but can be explained if the neural activity involved in the preparatory set directs plasticity.

Retrograde regulation of motoneuron differentiation by muscle beta-catenin.

Synapse formation requires proper interaction between pre- and postsynaptic cells. In anterograde signaling, neurons release factors to guide postsynaptic differentiation. However, less is known about how postsynaptic targets retrogradely regulate presynaptic differentiation or function. We found that muscle-specific conditional knockout of beta-catenin (Ctnnb1, also known as beta-cat) in mice caused both morphologic and functional defects in motoneuron terminals of neuromuscular junctions (NMJs). In the absence of muscle beta-catenin, acetylcholine receptor clusters were increased in size and distributed throughout a wider region. Primary nerve branches were mislocated, whereas secondary or intramuscular nerve branches were elongated and reduced in number. Both spontaneous and evoked neurotransmitter release was reduced at the mutant NMJs. Furthermore, short-term plasticity and calcium sensitivity of neurotransmitter release were compromised in beta-catenin-deficient muscle. In contrast, the NMJ was normal in morphology and function in motoneuron-specific beta-catenin-deficient mice. Taken together, these observations indicate a role for muscle beta-catenin in presynaptic differentiation and function, identifying a previously unknown retrograde signaling in the synapse formation and synaptic plasticity.

Neuregulin-1 enhances depolarization-induced GABA release.

Neuregulin-1 (NRG1), a regulator of neural development, has been shown to regulate neurotransmission at excitatory synapses. Although ErbB4, a key NRG1 receptor, is expressed in glutamic acid decarboxylase (GAD)-positive neurons, little is known about its role in GABAergic transmission. We show that ErbB4 is localized at GABAergic terminals of the prefrontal cortex. Our data indicate a role of NRG1, both endogenous and exogenous, in regulation of GABAergic transmission. This effect was blocked by inhibition or mutation of ErbB4, suggesting the involvement of ErbB4. Together, these results indicate that NRG1 regulates GABAergic transmission via presynaptic ErbB4 receptors, identifying a novel function of NRG1. Because both NRG1 and ErbB4 have emerged as susceptibility genes of schizophrenia, these observations may suggest a mechanism for abnormal GABAergic neurotransmission in this disorder.

Glutamate transporters regulate extrasynaptic NMDA receptor modulation of Kv2.1 potassium channels.

Delayed-rectifier Kv2.1 potassium channels regulate somatodendritic excitability during periods of repetitive, high-frequency activity. Recent evidence suggests that Kv2.1 channel modulation is linked to glutamatergic neurotransmission. Because NMDA-type glutamate receptors are critical regulators of synaptic plasticity, we investigated NMDA receptor modulation of Kv2.1 channels in rodent hippocampus and cortex. Bath application of NMDA potently unclustered and dephosphorylated Kv2.1 and produced a hyperpolarizing shift in voltage-dependent activation of voltage-sensitive potassium currents (I(K)). In contrast, driving synaptic activity in Mg2+-free media to hyperactivate synaptic NMDA receptors had no effect on Kv2.1 channels, and moderate pentylenetetrazole-induced seizure activity in adult mice did not dephosphorylate hippocampal Kv2.1 channels. Selective activation of extrasynaptic NMDA receptors unclustered and dephosphorylated Kv2.1 channels and produced a hyperpolarizing shift in neuronal I(K). In addition, inhibition of glutamate uptake rapidly activated NMDA receptors and dephosphorylated Kv2.1 channels. These observations demonstrate that regulation of intrinsic neuronal activity by Kv2.1 is coupled to extrasynaptic but not synaptic NMDA receptors. These data support a novel mechanism for glutamate transporters in regulation of neuronal excitability and plasticity through extrasynaptic NMDA receptor modulation of Kv2.1 channels.

Adaptive plasticity of NMDA receptors and dendritic spines: implications for enhanced vulnerability of the adolescent brain to alcohol addiction.

It is now known that brain development continues into adolescence and early adulthood and is highly influenced by experience-dependent adaptive plasticity during this time. Behaviorally, this period is also characterized by increased novelty seeking and risk-taking. This heightened plasticity appears to be important in shaping behaviors and cognitive processes that contribute to proper development of an adult phenotype. However, increasing evidence has linked these same experience-dependent learning mechanisms with processes that underlie drug addiction. As such, the adolescent brain appears to be particularly susceptible to experience-dependent learning processes associated with consumption of alcohol and addictive drugs. At the level of the synapse, homeostatic changes during ethanol consumption are invoked to counter the destabilizing effects of ethanol on neural networks. This homeostatic response may be especially pronounced in the adolescent and young adult brain due to its heightened capacity to undergo experience-dependent changes, and appears to involve increased synaptic targeting of NMDA receptors. Interestingly, recent work from our lab also indicates that the enhanced synaptic localization of NMDA receptors promotes increases in the size of dendritic spines. This increase may represent a structural-based mechanism that supports the formation and stabilization of maladapted synaptic connections that, in a sense, "fix" the addictive behavior in the adolescent and young adult brain.

Epileptic pilocarpine-treated rats exhibit aberrant hippocampal EPSP-spike potentiation but retain long-term potentiation.

Hippocampal neuron plasticity is strongly associated with learning, memory, and cognition. In addition to modification of synaptic function and connectivity, the capacity of hippocampal neurons to undergo plasticity involves the ability to change nonsynaptic excitability. This includes altering the probability that EPSPs will generate action potentials (E-S plasticity). Epilepsy is a prevalent neurological disorder commonly associated with neuronal hyperexcitability and cognitive dysfunction. We examined E-S plasticity in chronically epileptic Sprague-Dawley rats 3-10 weeks after pilocarpine-induced status epilepticus CA1 neurons in hippocampal slices were assayed by whole-cell current clamp to measure EPSPs evoked by Schaffer collateral stimulation. Using a weak spike-timing-dependent protocol to induce plasticity, we found robust E-S potentiation in conjunction with weak long-term potentiation (LTP) in saline-treated rats. In pilocarpine-treated rats, a similar degree of LTP was found, but E-S potentiation was reduced. Additionally, the degree of E-S potentiation was not correlated with the degree of LTP for either group, suggesting that they independently contribute to neuronal plasticity. E-S potentiation also differed from LTP in that E-S plasticity could be induced solely from action potentials generated by postsynaptic current injection. The calcium chelating agent BAPTA in the intracellular solution blocked LTP and E-S potentiation, revealing the calcium dependence of both processes. These findings suggest that LTP and E-S potentiation have overlapping but nonidentical mechanisms of inducing neuronal plasticity that may independently contribute to cognitive disruptions observed in the chronic epileptic state.

Chronic ethanol induces synaptic but not extrasynaptic targeting of NMDA receptors.

The development of ethanol tolerance and dependence reflects neuroadaptive changes in response to continuous depression in synaptic activity. The present study used confocal imaging and electrophysiology procedures to assess the effects of prolonged ethanol exposure on NMDA receptor trafficking in cultures of hippocampal neurons. Neurons exposed to 50 mm ethanol for 4 d showed an increase in the colocalization of NMDA receptor type 1 (NR1) clusters with the presynaptic marker protein synapsin. This was accompanied by significant increases in the size and density of these synapsin-associated clusters with no change observed in nonsynapsin-associated NR1 clusters. Similar effects were observed with NR2B clustering after chronic ethanol exposure. The increase in synaptic NMDA receptor clustering was prevented by addition of a protein kinase A inhibitor or by coexposure to a low concentration of NMDA and was reversed when ethanol was removed from the cultures. No changes were observed in the synaptic content, cluster size, or density of AMPA receptors after ethanol exposure. Electrophysiological measurements on ethanol-treated neurons revealed a similar enhancement in synaptic NMDA currents with no change in AMPA-mediated events. After isolation of extrasynaptic NMDA receptors by MK801 (+)-5-methyl-10,11-dihydro-5H-dibenzo [a,d] cyclohepten-5,10-imine maleate (/) trapping, whole-cell responses to NMDA were not different between control and ethanol-treated neurons These observations demonstrate that neuroadaptive changes in NMDA receptors in response to prolonged ethanol exposure occur through activity-dependent processes that regulate their synaptic targeting and localization.

Ethanol disrupts NMDA receptor and astroglial EAAT2 modulation of Kv2.1 potassium channels in hippocampus.

Delayed-rectifier Kv2.1 channels are the principal component of voltage-sensitive K+ currents (I(K)) in hippocampal neurons and are critical regulators of somatodendritic excitability. In a recent study, we demonstrated that surface trafficking and phosphorylation of Kv2.1 channels is modulated by NMDA-type glutamate receptors and that astroglial excitatory amino acid transporters 2 (EAAT2) regulate the coupling of NMDA receptors and Kv2.1 channels. Because ethanol is known to acutely inhibit NMDA receptors, we sought to determine if NMDA receptor and astroglial EAAT2 modulation of Kv2.1 channels is impaired by ethanol in the rodent hippocampus. As expected, bath application of NMDA to hippocampal cultures reduced the size of Kv2.1 clusters and produced a hyperpolarizing shift in the voltage-dependent activation of I(K) that was associated with dephosphorylated Kv2.1 channels. Ethanol, applied acutely, prevented the hyperpolarizing shift in activation of I(K) induced by NMDA and restored Kv2.1 clustering and phosphorylation to near control levels. Ethanol also attenuated the dephosphorylation of Kv2.1 channels produced by the EAAT2 selective inhibitor dihydrokainic acid. These data demonstrate that acute ethanol disrupts changes in Kv2.1 channels that follow NMDA receptor activation and impairs astroglial regulation of the functional coupling between NMDA receptors and Kv2.1 channels.

Homeostatic plasticity during alcohol exposure promotes enlargement of dendritic spines.

Modifications of the size, shape and number of dendritic spines is thought to be an important component of activity-dependent changes of neuronal circuits, and may play an important role in the plasticity of drug addiction. The present study examined whether homeostatic increases in synaptic N-methyl-d-aspartate (NMDA) receptors in response to chronic ethanol exposure is associated with corresponding morphological changes in dendritic spines. Prolonged exposure of rat hippocampal cultures to either the NMDA receptor antagonist d(-)-2-amino-5-phosphono-pentanoic acid or to ethanol increased punctate staining of F-actin and the postsynaptic density protein-95 (PSD-95). The increase in dendritic F-actin occurred only with clusters that co-localized with PSD-95 clusters, indicating that these actin structures likely represent dendritic spines. The ethanol-induced increases in PSD-95 and F-actin clusters were activity-dependent and reversible. Finally, inhibition of protein palmitoylation prevented ethanol-induced increases in synaptic NMDA receptor clustering and F-actin without altering the basal clustering of either F-actin or PSD-95. These observations support a model in which chronic ethanol exposure induces homeostatic increases of NR2B-containing NMDA receptors and PSD-95 to the postsynaptic density. This in turn may provide a scaffolding platform for the subsequent recruitment of actin signaling cascades that alter actin cycling and promote spine enlargement.

Structural and functional modifications in glutamateric synapses following prolonged ethanol exposure.

This article summarizes the proceedings of a symposium presented at the 2005 annual meeting of the Research Society on Alcoholism in Santa Barbara, California, USA. The organizer and chair was L. Judson Chandler. The presentations were (1) Chronic Ethanol Exposure, N-Methyl-D-Aspartate (NMDA) Receptor Dynamics, and Withdrawal Hyperexcitability, by Adam Hendricson, Regina Maldve, and Richard Morrisett; (2) Ethanol-Induced Synaptic Targeting of NMDA Receptors Is Associated With Enhanced Postsynaptic Density-95 Clustering and Spine Size, by Judson Chandler and Ezekiel Carpenter-Hyland; (3) Presynaptic and Postsynaptic Alterations in the Nucleus Accumbens Following Chronic Alcohol Exposure, by Feng Zhou, Youssef Sari, and Richard Bell; and (4) An Active Role for Accumbens Homer2 Expression in Alcohol-Induced Neural Plasticity, by Karen Szumlinski.