Learning and memory depend on the activity-dependent structural plasticity of synapses and changes in neuronal gene expression. We show that deletion of the MEF2C transcription factor in the CNS of mice impairs hippocampal-dependent learning and memory. Unexpectedly, these behavioral changes were accompanied by a marked increase in the number of excitatory synapses and potentiation of basal and evoked synaptic transmission. Conversely, neuronal expression of a superactivating form of MEF2C results in a reduction of excitatory postsynaptic sites without affecting learning and memory performance. We conclude that MEF2C limits excessive synapse formation during activity-dependent refinement of synaptic connectivity and thus facilitates hippocampaldependent learning and memory.synaptic transmission ͉ synaptogenesis ͉ learning deficits N eurons process and retain information by forming synaptic connections that are modified by the intensity and frequency of their activity. The capacity to regulate the efficacy of synaptic transmission is essential for the continual remodeling of neural networks required for cognitive processes such as learning and memory. Distinct molecular mechanisms control synaptic plasticity associated with the different temporal stages of memory. A short-term process lasting minutes depends on modifications of preexisting proteins, whereas a long-term process lasting hours and days depends on changes in gene expression and protein synthesis (1).Originally identified as regulators of muscle development, members of the MEF2 (Myocyte Enhancer Factor 2) family of MADS (MCM1, agamous, deficiens, serum response factor) box transcription factors are expressed in overlapping but distinct regions of the CNS that correlate with the withdrawal of neurons from the cell cycle and acquisition of a differentiated phenotype (2). Mef2c is the first of four Mef2 genes to be expressed in the CNS and, in the adult brain, is highly expressed in the frontal cortex, entorhinal cortex, dentate gyrus, and amygdala (3, 4). RNA interference-mediated knockdown of MEF2A and MEF2D in cultured hippocampal neurons increases the number of excitatory synapses and the frequency of miniature excitatory postsynaptic currents (mEPSCs) (5). These alterations depend on the ability of the MEF2 proteins to stimulate neural activitydependent transcription of target genes (5). In contrast, loss of MEF2A in cerebellar granule neurons results in a decrease in the number of dendritic claws (6).Here, we present an analysis of the neuronal functions of the Mef2 gene in vivo. Through conditional deletion of Mef2c and expression of a superactive form of MEF2C in neurons of mice, we show that this MEF2 isoform plays an essential role in hippocampal-dependent learning and memory by suppressing the number of excitatory synapses and thus regulating basal and evoked synaptic transmission. ResultsBrain-Specific Deletion of MEF2C. We deleted Mef2c specifically in the CNS by breeding Mef2c loxP/loxP females (7) to Mef2c KO/ϩ heterozygous male (8) mice h...

Reelin, a large protein that regulates neuronal migration during embryonic development, activates a conserved signaling pathway that requires its receptors, very low-density lipoprotein receptor and apolipoprotein E receptor 2, the cytoplasmic adaptor protein Disabled-1 (Dab1), and Src family kinases (SFK). Reelin also markedly enhances long-term potentiation in the adult hippocampus, suggesting that this developmental signaling pathway can physiologically modulate learning and behavior. Here, we show that Reelin can regulate NMDA-type glutamate receptor activity through a mechanism that requires SFKs and Dab1. Reelin mediates tyrosine phosphorylation of and potentiates calcium influx through NMDA receptors in primary wild-type cortical neurons but not in Dab1 knock-out neurons or in cells in which Reelin binding to its receptors is blocked by a receptor antagonist. Inhibition of SFK abolishes Reelin-induced and glutamate-dependent enhancement of calcium influx. We also show that Reelin-induced augmentation of Ca 2ϩ entry through NMDA receptors increases phosphorylation and nuclear translocation of the transcription factor cAMP-response element binding protein. Thus, Reelin may physiologically modulate learning and memory by modulating NMDA receptor functions.

In a synapse, spontaneous and action-potential-driven neurotransmitter release is assumed to activate the same set of postsynaptic receptors. Here, we tested this assumption using (ϩ)-5-methyl-10,11-dihydro-5H-dibenzo

Cholesterol is a prominent component of nerve terminals. To examine cholesterol's role in central neurotransmission, we treated hippocampal cultures with methyl-β-cyclodextrin, which reversibly binds cholesterol, or mevastatin, an inhibitor of cholesterol biosynthesis, to deplete cholesterol. We also used hippocampal cultures from Niemann-Pick type C1-deficient mice defective in intracellular cholesterol trafficking. These conditions revealed an augmentation in spontaneous neurotransmission detected electrically and an increase in spontaneous vesicle endocytosis judged by horseradish peroxidase uptake after cholesterol depletion by methyl-β-cyclodextrin. In contrast, responses evoked by action potentials and hypertonicity were severely impaired after the same treatments. The increase in spontaneous vesicle recycling and the decrease in evoked neurotransmission were reversible upon cholesterol addition. Cholesterol removal did not impact on the low level of evoked neurotransmission seen in the absence of synaptic vesicle SNARE protein synaptobrevin-2 whereas the increase in spontaneous fusion remained. These results suggest that synaptic cholesterol balances evoked and spontaneous neurotransmission by hindering spontaneous synaptic vesicle turnover and sustaining evoked exo-endocytosis.

During short-term synaptic depression, neurotransmission rapidly decreases in response to repetitive action potential firing. Here, by blocking the vacuolar ATPase, alkalinizing the extracellular pH, or exposing hippocampal slices to pH buffers, we impaired neurotransmitter refilling, and electrophysiologically tested the role of vesicle reuse in synaptic depression. Under all conditions, synapses onto hippocampal CA1 pyramidal cells showed faster depression with increasing stimulation frequencies. At 20 Hz, compromising neurotransmitter refilling increased depression within 300 ms reaching completion within 2 s, suggesting a minimal contribution of reserve vesicles to neurotransmission. In contrast, at 1 Hz, depression emerged gradually and became significant within 100 s. Moreover, the depression induced by pH buffers was reversible with a similar frequency dependence, suggesting that the frequency-dependent increase in depression was caused by impairment of rapid synaptic vesicle reuse. These results indicate that synaptic vesicle trafficking impacts the kinetics of short-term synaptic plasticity at an extremely rapid time scale.