Loss of the fragile X mental retardation protein decouples metabotropic glutamate receptor dependent priming of long-term potentiation from protein synthesis

Abstract

Fragile X Syndrome (FXS), the most common inherited form of intellectual disability, is caused by loss of the fragile X mental retardation protein (FMRP). FMRP is a negative regulator of local mRNA translation downstream of group 1 metabotropic glutamate receptor (Gp1 mGluR) activation. In the absence of FMRP there is excessive mGluR-dependent protein synthesis, resulting in exaggerated mGluR-dependent long-term synaptic depression (LTD) in area CA1 of the hippocampus. Understanding disease pathophysiology is critical for development of therapies for FXS and the question arises of whether it is more appropriate to target excessive LTD or excessive mGluR-dependent protein synthesis. Priming of long-term potentiation (LTP) is a qualitatively different functional consequence of Gp1 mGluR-stimulated protein synthesis at the same population of CA1 synapses where LTD can be induced. Therefore we determined if LTP priming, like LTD, is also disrupted in the Fmr1 knockout (KO) mouse. We found that mGluR-dependent priming of LTP is of comparable magnitude in wild-type (WT) and Fmr1 KO mice. However, whereas LTP priming requires acute stimulation of protein synthesis in WT mice, it is no longer protein synthesis dependent in the Fmr1 KO. These experiments show that the dysregulation of mGluR-mediated protein synthesis seen in Fmr1 KO mice has multiple consequences on synaptic plasticity, even within the same population of synapses. Furthermore, it suggests that there is a bifurcation in the Gp1 mGluR signaling pathway, with one arm triggering synaptic modifications such as LTP priming and LTD and the other stimulating protein synthesis that is permissive for these modifications.

Fig. 1.

R,S-Dihydroxyphenylglycine (DHPG) application facilitates subsequently induced long-term synaptic potentiation (LTP). Field excitatory postsynaptic potentials (fEPSPs) were recorded from the CA1 region of hippocampal slices from either (A) wildtype (WT) or (B) Fmr1 knockout mice (KO). After 1 h of baseline recording, unprimed slices were administered a 1-s 100-Hz tetanus (indicated by arrow), which induced a modest level of LTP in both WT and KO slices (WT: 111.2 ± 2.1%, n = 9 slices from 9 animals, open black circles; KO: 113.8 ± 3.1%, n = 9 slices from 8 animals, open gray circles). In both genotypes, a 10-min priming application of the group 1 metabotropic glutamate receptor (Gp1 mGluR) agonist DHPG (10 μM, black bar) significantly enhanced the magnitude of subsequent LTP induced using this same 100-Hz tetanus (WT: 123.9 ± 3.8%, n = 11 slices from 11 animals, closed black circles, P < 0.02; KO: 133.7 ± 6.7%, n = 11 slices from 10 animals, closed gray circles, P < 0.02). Representative field potential traces (average of 10 sweeps) were taken at times indicated by numerals.

Fig. 2.

DHPG-induced priming of LTP does not require protein synthesis in Fmr1 KO mice. Delivery of the protein synthesis inhibitor cycloheximide (60 μM CHX, 30 min; gray bar) before and during DHPG priming prevented facilitation of LTP in slices from WT mice (A; unprimed: 117.5 ± 7.0%, n = 7 slices from 6 animals, open black circles; primed: 118.6 ± 6.0%, n = 8 slices from 6 animals, closed black circles; P = 0.94); however, this treatment had no effect on DHPG-induced priming in slices from Fmr1 KO mice (B; unprimed: 118.5 ± 6.1%, n = 7 slices from 6 animals, open gray circles; primed: 149.6 ± 11.0%, n = 8 slices from 7 animals, closed gray circles; P < 0.05). Representative field potential traces (average of 10 sweeps) were taken at times indicated by numerals.

Fig. 3.

Summary of DHPG-induced priming of LTP and its protein synthesis dependence in wildtype and Fmr1 KO mice. Bar graphs represent the average percentage LTP observed 55–60 min posttetanus. Wildtype unprimed: open black; wildtype primed: closed black; Fmr1 KO unprimed: open gray; Fmr1 KO primed: closed gray. Asterisks denote significant differences (unpaired Student's t-test, P < 0.05).

Fig. 4.

Model. The finding that LTP priming by mGluR activation occurs in the Fmr1 KO without a need for acute protein synthesis suggests a bifurcation in the signaling pathway. The priming step (1) occurs in response to mGluR activation via a mechanism involving posttranslational modification of synaptic proteins (possibly the AMPA receptor itself). In WT animals, priming is not possible without (2) concurrent mGluR activation of mRNA translation and synthesis of protein(s) that gate plasticity. In the absence of the translational repressor FMPP, the gating proteins are constitutively overexpressed, rendering priming no longer sensitive to protein synthesis inhibitors. The identity of the hypothetical gating proteins remains to be determined. AMPA, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid.