Persistent synaptic scaling independent of AMPA receptor subunit composition

Abstract

Despite long-standing evidence that the specific intracellular domains of AMPA-type glutamate receptor (AMPAR) subunits are critical for trafficking, it has recently been demonstrated that there is no absolute requirement for any AMPAR subunit for the receptor insertion underlying LTP. It is unclear whether this holds true to other forms of plasticity. Homeostatic synaptic plasticity (HSP) is an important form of negative feedback that provides stability to neuronal networks, and results at least in part from the insertion of AMPARs into glutamatergic synapses following chronic reductions in neuronal activity. Similar to LTP, the GluA1 subunit has been suggested to be the requisite subunit for HSP-induced AMPAR insertion and acute treatment with signaling molecules that underlie some forms of HSP results in the preferential incorporation of GluA2-lacking receptors. However, knockdown experiments have instead implicated a requirement for the GluA2 subunit. Here we re-examined the requirement for specific AMPAR subunit during chronic tetrodotoxin-induced HSP using hippocampal cultures derived from AMPAR subunit knock-out mice. We observed HSP in cultures from GluA1⁻/⁻, GluA2⁻/⁻, and GluA2⁻/⁻ GluA3⁻/⁻ mice, and conclude that, as with LTP, there is no subunit requirement for HSP.

Figure 1.

AMPAR surface trafficking in response to inactivity is independent of subunit identity. A, Example images of cell-surface labeling of GluA2 in wild-type and GluA1^−/− neurons, control or treated for 2 d with TTX. B, Summary data showing an increase in AMPAR surface levels with 2 d TTX treatment over control in wild-type and GluA1^−/− cultures (p < 0.01, n = 60 images per condition). C, Example images of cell-surface labeling of GluA1 in GluA3^−/−, and GluA2^−/− GluA3^−/− neurons, control and treated for 2 d with TTX. D, Summary data showing an increase in AMPAR surface levels with TTX treatment over control in GluA3^−/− (p < 0.01), and GluA2^−/− GluA3^−/− cultures (p < 0.001, n = 30–61 images per condition).

Figure 2.

Synaptic scaling is independent of AMPAR subunit identity. A, Example traces from wild-type, GluA1^−/−, and GluA2^−/− neurons, control or treated for 2 d with TTX. B, Mean mEPSC amplitudes for wild-type, GluA1^−/−, and GluA2^−/− increased significantly with 2 d TTX treatment (wild-type control n = 18, TTX n = 20, p < 0.01; GluA1^−/− control n = 10, TTX n = 10, p = 0.01; GluA2^−/− control n = 17, TTX n = 19, p = 0.04).

Figure 3.

Synaptic scaling remains multiplicative in the absence of GluA1 or GluA2. A, Randomly selected mEPSCs for 2 d TTX-treated were ranked and averaged, and compared against control. Linear regression yielded slope factors of: TTX = control × 1.54 for wild-type, TTX = 2.46 × control for GluA1^−/−, and TTX = 2.01 × control for GluA2^−/−. B, Cumulative fraction for ranked averaged mEPSC amplitudes (±SEM) was plotted for control (gray line) and TTX (black line) for each genotype; K–S testing of TTX versus control yielded p values of < 0.001 for all genotypes. The TTX distribution for each individual genotype was then scaled down by respective slope factor (dashed black line), and when compared versus control yielded p values of 0.62 for wild-type, 1.00 for GluA1^−/−, and 1.00 for GluA2^−/−.