Reactivated memory undergoes a rebuilding process that depends on de novo protein synthesis. This suggests that retrieval is dynamic and serves to incorporate new information into preexisting memories. However, little is known about whether or not protein degradation is involved in the reorganization of retrieved memory. We found that postsynaptic proteins were degraded in the hippocampus by polyubiquitination after retrieval of contextual fear memory. Moreover, the infusion of proteasome inhibitor into the CA1 region immediately after retrieval prevented anisomycin-induced memory impairment, as well as the extinction of fear memory. This suggests that ubiquitin- and proteasome-dependent protein degradation underlies destabilization processes after fear memory retrieval. It also provides strong evidence for the existence of reorganization processes whereby preexisting memory is disrupted by protein degradation, and updated memory is reconsolidated by protein synthesis.

Serotonin (5-HT) plays a critical role in modulating synaptic plasticity in the marine mollusc Aplysia and in the mammalian nervous system. In Aplysia sensory neurons, 5-HT can activate several signal cascades, including PKA and PKC, presumably via distinct types of G proteincoupled receptors. However, the molecular identities of these receptors have not yet been identified. We here report the cloning and functional characterization of a 5-HT receptor that is positively coupled to adenylyl cyclase in Aplysia neurons. The cloned receptor, 5-HT apAC1, stimulates the production of cAMP in HEK293T cells and in Xenopus oocytes. Moreover, the knockdown of 5-HT apAC1 expression by RNA interference blocked 5-HT-induced cAMP production in Aplysia sensory neurons and blocked synaptic facilitation in nondepressed or partially depressed sensory-to-motor neuron synapses. These data implicate 5-HT apAC1 as a major modulator of learning related synaptic facilitation in the direct sensory to motor neuron pathway of the gill withdrawal reflex.5-HT receptor ͉ memory ͉ cAMP ͉ protein kinase A 5-Hydroxytryptamine (5-HT), or serotonin, is a key neurotransmitter that modulates a variety of behaviors in both invertebrate and vertebrate animals and is involved in the regulation of mood and mood disorders in humans (1). Serotonin also modulates synaptic plasticity in the marine mollusc Aplysia (2, 3). Synaptic facilitation of the connections between sensory and motor neurons of the gill-withdrawal reflex is mediated by 5-HT, and this form of synaptic plasticity has been found to be a critical cellular mechanism of behavioral sensitization (4-6). A number of pharmacological studies have found that, depending on the behavioral history and pattern of sensory stimulation, 5-HT stimulates several downstream signaling pathways, including protein kinase A (PKA), protein kinase C (PKC), and mitogen-activated protein kinase (MAPK), suggesting that serotonin acts on more than one receptor type (2,3,7,8). Of these signaling cascades, the adenylyl cyclase-cAMP-PKA cascade has been most extensively investigated because of its important roles in both behavioral sensitization and synaptic facilitation (3, 4, 9, 10). Historically, this was the initially identified second-messenger system involved in the regulation of synaptic plasticity, behavior, and memory storage (4).A single pulse of 5-HT activates PKA, which phosphorylates and inactivates potassium channels (11) and subsequently increases synaptic strength at nondepressed synapses. At depressed synapses, however, PKC becomes the major downstream kinase to be activated by a single pulse of 5-HT (8). In addition, repetitive exposures to 5-HT that induce long-term facilitation result in the activation of additional kinases, including MAPK (12), that translocate to the nucleus to induce gene expression. However, the molecular mechanism for this dynamic coupling specificity of downstream signaling pathways is not known.In vertebrates, seven families of 5-HT receptors have been characterized; six of the...

The cAMP cascade and vascular endothelial growth factor (VEGF) are critical modulators of depression. Here we have tested whether the antidepressive effect of the cAMP cascade is mediated by VEGF in the adult hippocampus. We used a conditional genetic system in which the Aplysia octopamine receptor (Ap oa 1 ), a G s -coupled receptor, is transgenically expressed in the forebrain neurons of mice. Chronic activation of the heterologous Ap oa 1 by its natural ligand evoked antidepressant-like behaviors, accompanied by enhanced phosphorylation of cAMP response element-binding protein and transcription of VEGF in hippocampal dentate gyrus (DG) neurons. Selective knockdown of VEGF in these cells during the period of cAMP elevation inhibited the antidepressant-like behaviors. These findings reveal a molecular interaction between the cAMP cascade and VEGF expression, and the pronounced behavioral consequences of this interaction shed light on the mechanism underlying neuronal VEGF functions in antidepression.

Phosphodiesterases (PDEs) are known to play a key role in the compartmentalization of cAMP signaling; however, the molecular mechanisms underlying intracellular localization of different PDE isoforms are not understood. In this study, we have found that each of the supershort, short, and long forms of apPDE4 showed distinct localization in the cytoplasm, plasma membrane, and both plasma membrane and presynaptic terminals, respectively. The N-terminal 20 amino acids of the long form of apPDE4 were involved in presynaptic terminal targeting by binding to several lipids. In addition, the N terminus of the short form of apPDE4 bound to several lipids including phosphoinositols, thereby targeting the plasma membrane. Overexpression of the long and the short forms, but not the supershort form attenuated 5-HT-induced membrane hyperexcitability. Finally, the knockdown of apPDE4s in sensory neurons impaired both short-term and long-term facilitation. Thus, these results suggest that apPDE4s can participate in the regulation of cAMP signaling through specific subcellular localization by means of lipid binding activities.

Consistent evidence from pharmacological and genetic studies shows that cAMP is a critical modulator of synaptic plasticity and memory formation. However, the potential of the cAMP signaling pathway as a target for memory enhancement remains unclear because of contradictory findings from pharmacological and genetic approaches. To address these issues, we have developed a novel conditional genetic system in mice based on the heterologous expression of an Aplysia octopamine receptor, a G-protein-coupled receptor whose activation by its natural ligand octopamine leads to rapid and transient increases in cAMP. We find that activation of this receptor transgenically expressed in mouse forebrain neurons induces a rapid elevation of hippocampal cAMP levels, facilitates hippocampus synaptic plasticity, and enhances the consolidation and retrieval of fear memory. Our findings clearly demonstrate that acute increases in cAMP levels selectively in neurons facilitate synaptic plasticity and memory, and illustrate the potential of this heterologous system to study cAMP-mediated processes in mammalian systems.