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. 2010 Jun 22;107(25):11591-6.
doi: 10.1073/pnas.1002262107. Epub 2010 Jun 7.

Characterization and reversal of synaptic defects in the amygdala in a mouse model of fragile X syndrome

Aparna Suvrathan  1 , Charles A Hoeffer, Helen Wong, Eric Klann, Sumantra Chattarji
Affiliations

Characterization and reversal of synaptic defects in the amygdala in a mouse model of fragile X syndrome

Aparna Suvrathan et al. Proc Natl Acad Sci U S A. .

Abstract

Fragile X syndrome (FXS), a common inherited form of mental impairment and autism, is caused by transcriptional silencing of the fragile X mental retardation 1 (FMR1) gene. Earlier studies have identified a role for aberrant synaptic plasticity mediated by the metabotropic glutamate receptors (mGluRs) in FXS. However, many of these observations are derived primarily from studies in the hippocampus. The strong emotional symptoms of FXS, on the other hand, are likely to involve the amygdala. Unfortunately, little is known about how exactly FXS affects synaptic function in the amygdala. Here, using whole-cell recordings in brain slices from adult Fmr1 knockout mice, we find mGluR-dependent long-term potentiation to be impaired at thalamic inputs to principal neurons in the lateral amygdala. Consistent with this long-term potentiation deficit, surface expression of the AMPA receptor subunit, GluR1, is reduced in the lateral amygdala of knockout mice. In addition to these postsynaptic deficits, lower presynaptic release was manifested by a decrease in the frequency of spontaneous miniature excitatory postsynaptic currents (mEPSCs), increased paired-pulse ratio, and slower use-dependent block of NMDA receptor currents. Strikingly, pharmacological inactivation of mGluR5 with 2-methyl-6-phenylethynyl-pyridine (MPEP) fails to rescue either the deficit in long-term potentiation or surface GluR1. However, the same acute MPEP treatment reverses the decrease in mEPSC frequency, a finding of potential therapeutic relevance. Therefore, our results suggest that synaptic defects in the amygdala of knockout mice are still amenable to pharmacological interventions against mGluR5, albeit in a manner not envisioned in the original hippocampal framework.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.
mGluR5-dependent LTP is absent and surface GluR1 is reduced in the amygdala of Fmr1 KO mice. (A) High-frequency stimulation at 30 Hz (arrow) induces LTP in LA slices from WT mice but not Fmr1 KO siblings. (Insets) Average of 10 traces before tetanus and up to 45th minute: (i) WT (ii) KO. (Scale bars: 5 mV, 50 ms.) (B) Mean values of LTP from minutes 40 to 45, normalized to the 5-min pretetanus baseline. *P = 0.033 (number of cells). (C) Surface expression of GluR1, determined by pull-down of biotin-labeled receptor, is reduced in KO mice. *P = 0.024, (number of mice). (D) Total GluR1 is not different. (E) Representative Western blots are shown. Surface GluR1 is normalized to avidin, and total GluR1 to GAPDH. FMRP is absent in tissue from KO mice.
Fig. 2.
Fig. 2.
Neither the LTP deficit nor reduction in surface GluR1 is rescued by MPEP. (A) MPEP (40 μM) treatment fails to rescue deficit in 30 Hz-LTP in KO cells. Mean values of LTP from minutes 30 to 35, normalized to the 5-min baseline. *P = 0.019. (Number of cells) (B) MPEP preincubation does not change surface or total GluR1 levels in KO mice. (Number of mice) (C) LTP induced by 100-Hz tetanic stimuli (arrow) in WT slices is greater than KO slices; this deficit is not reversed by MPEP treatment. (Insets) Average of 10 traces before tetanus and up to 35th minute: (i) WT (ii) KO. (Scale bars: 5 mV, 50 ms.) (D) Mean values of LTP from minutes 30 to 35, normalized to 5-min pretetanus. One-way ANOVA *P = 0.022, post hoc Tukey's HSD P values: WT-KO = 0.034, WT-KO+MPEP = 0.042 (number of cells).
Fig. 3.
Fig. 3.
LA cells exhibit reduced frequency and amplitude of mEPSCs, enhanced paired-pulse facilitation, and slower MK-801-induced decays in Fmr1 KO mice. (A) Sample mEPSC traces. (Scale bar: 20 pA, 1s.) (B) Summary of change in frequency. **P = 0.002. (C) Summary of change in amplitude. **P = 0.005. (D) Strontium-desynchronized events evoked by stimulation of thalamic inputs to the LA show reduced amplitude in KO mice. *P = 0.029. (Inset) Sample traces. (Scale bars: 50 pA, 0.02 s.) (E) Enhanced paired-pulse facilitation in KO (○) compared with WT cell (●). **P = 0.006, (repeated measures ANOVA). (Inset) Sample of averaged EPSC traces at 100-ms interstimulus interval. (Scale bars: 20 pA, 0.05 s.) (Number of cells). (F) Release probability is lower at thalamic inputs to LA in KO mice. Normalized amplitude of the NMDA-EPSC is plotted after bath application of 40 μM MK-801. Progressive block of the NMDA-EPSC by MK-801 is slower at KO synapses (gray) compared with WT synapses (black) in the LA. (Inset) Sample traces of first five EPSCs in MK-801. (Scale bars: 50 pA, 0.02 s.)
Fig. 4.
Fig. 4.
LA Deficits in presynaptic release are reversed by acute MPEP treatment. (A) Preincubation of LA slices from Fmr1 KO mice in 40 μM MPEP and reversal of decreased mEPSC frequency: cumulative probability of mEPSC interevent intervals (IEI). The x axis truncated at IEI of 50,000 ms. IEIs averaged within 100-ms bins, then across cells, and normalized. K-S test, significance levels P < 0.0001 between WT-KO and MPEP-KO, P = 0.999 between WT-MPEP. (B) Cumulative probability of mEPSC amplitudes. Bin size 0.5 pA. K-S test, P < 0.0001 between WT-KO, P = 0.016 between WT-MPEP, and P = 0.163 between KO and MPEP. (C) Summary of across-cell comparisons: MPEP preincubation reverses reduction in mEPSC frequency in KO cells. One-way ANOVA with Welch's statistic P = 0.001. Post hoc pairwise Games-Howell test: Fmr1 KO-WT **P = 0.005; MPEP-treated KO-KO *P = 0.029; MPEP-treated KO-WT: P = 0.439. (D) LA cells in KO slices have lower mEPSC amplitude. One-way ANOVA P = 0.018 (Post hoc pairwise Games-Howell test *, P = 0.015). Reduction in mEPSC amplitude is not rescued by MPEP preincubation (number of cells).
Fig. 5.
Fig. 5.
Summary of results in the LA of Fmr1 KO mice and how they differ from those reported earlier in the hippocampus. MPEP treatment reverses some phenotypes (ticks), but fails to rescue others (cross); absence of data, ?. **Contradictory findings (6, 28, 32).
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