MEF2C, a transcription factor that facilitates learning and memory by negative regulation of synapse numbers and function

Abstract

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 hippocampal-dependent learning and memory.

Fig. 1.

Hippocampal-dependent learning deficits in Mef2c^BKO/KO mice. Contextual and cued conditioning are two forms of associative learning that induce effective memory for either the context or the cue after a single training session. Memory for the conditioned stimulus was measured as freezing, an absence of visible movement, when presented with the conditioned context or cue in a 24-h retention test. (A) Mef2c^BKO/KO (n = 18) showed impaired context-dependent memory compared to WT littermates (n = 17). In contrast, there was no difference in cue-dependent fear conditioning in Mef2c^BKO/KO compared to WT mice. (B) The threshold responses of Mef2c^BKO/KO and WT mice to foot shocks were indistinguishable, which indicates that the impairment in context conditioning represents a learning deficit and not a difference in pain sensitivity.

Fig. 2.

Basal synaptic transmission and short-term plasticity in dentate granule neurons of Mef2c^BKO/KO. (A) Schematic representation of electrophysiological recordings performed on granule cells of dentate gyrus region of mice hippocampal slices. (B) (Upper) Representative traces of whole-cell patch–clamp recordings demonstrating spontaneous EPSCs from dentate granule cells (n = 23 for WT and n = 20 for Mef2c^BKO/KO). Increased mEPSC frequency for dentate gyrus neurons in Mef2C^BKO/KO mice. **, Statistical significance in pairwise comparison; P = 0.005649. Spontaneuous EPSC amplitudes are unaffected in dentate gyrus neurons of Mef2C^BKO/KO mice. (C) Paired-pulse ratios of the first two responses during various stimulation frequencies were measured in WT (filled triangle) and Mef2c^BKO/KO (open triangle) dentate gyrus neurons. Paired-pulse facilitation is decreased in Mef2c^BKO/KO dentate gyrus when compared to WT mice.

Fig. 3.

Evoked perforant path synaptic transmission in Mef2C^BKO/KO. Lateral perforant path was stimulated by a 10- to 40-μA current pulse to evoke ESPCs at perforant path-dentate granule cell synapses. Representative examples of current voltage of glutamate-evoked response from single AMPA (A Upper) or NMDA (B Upper) synapses from WT and MEF2C-deficient neurons. Bar graph showing increased amplitude in both AMPA (A; V_m = −70 mV) and NMDA (B; V_m = +40 mV) mediated evoked EPSCs in granule cells of dentate gyrus of Mef2C^BKO/KO mice. Asterisk indicates statistical significance in pairwise comparison: P < 0.0005. (C) The AMPA/NMDA ratios were similar in WT and Mef2C^BKO/KO mice. (D) Input–output curves showing increase in the saturation of AMPA receptor in dentate gyrus neurons lacking MEF2C (open triangles).

Fig. 4.

Synaptic function is altered in NSE-MEF2C-VP16 transgenic mice. (A) (Upper) Representative traces of whole-cell patch–clamp recordings demonstrating spontaneous EPSCs from dentate granule cells (n = 24 for WT and n = 26 for NSE-MEF2C-VP16 transgenic mice). Bar graph showing a decrease in mEPSC frequency for dentate gyrus neurons in NSE-MEF2C-VP16 transgenic mice when compared to WT littermate controls. mEPSC amplitudes are unaffected in dentate gyrus neurons of the transgenic mice. (B) Paired-pulse facilitation at perforant path synapses also appeared normal in the transgenic mice. (C) Input–output curves showing decreased saturation of AMPA receptor in dentate gyrus neurons expressing the superactive MEF2C-VP16.

Fig. 5.

Effects of MEF2C on excitatory synapse number in dentate granule neurons. Spine density of dentate gyrus dendrites was determined from Golgi-stained sections of (A) Mef2c^+/+, Mef2c^BKO/KO and MEF2C-VP6 Tg mice. (Scale bar, 10 μm.) (B) Mef2c^BKO/KO animals have a significant increase in spine density when compared to WT littermates. *, Statistical significance in pairwise comparison; P = 0.0001. MEF2C-VP16 transgenic mice show a decrease in spine density when compared to WT littermates. **, Statistical significance in pairwise comparison; P = 0.018. For Mef2c^BKO/KO mice; P = 0.0001 using two-tailed two-sample equal variance t test; n = 30 dendritic segments from 26 neurons of two mice and n = 27 dendritic segments from 25 neurons of two mice, respectively. For MEF2C-VP16 transgenic mice; P = 0.018 using two-tailed two-sample equal variance t test; n = 28 dendritic segments from 26 neurons of two mice and n = 25 dendritic segments from 23 neurons of two mice, respectively. (C) Average density of synapsin-1-PSD-95 coclusters along the dendrites of cultured hippocampal neurons. There was no significant difference in the number of excitatory synapses between Mef2c^BKO/KO and WT neurons.