Ionotropic NMDA receptor signaling is required for the induction of long-term depression in the mouse hippocampal CA1 region

Abstract

Previous studies have provided strong support for the notion that NMDAR-mediated increases in postsynaptic Ca(2+) have a crucial role in the induction of long-term depression (LTD). This view has recently been challenged, however, by findings suggesting that LTD induction is instead attributable to an ion channel-independent, metabotropic form of NMDAR signaling. Thus, to explore the role of ionotropic versus metabotropic NMDAR signaling in LTD, we examined the effects of varying extracellular Ca(2+) levels or blocking NMDAR channel ion fluxes with MK-801 on LTD and NMDAR signaling in the mouse hippocampal CA1 region. We find that the induction of LTD in the adult hippocampus is highly sensitive to extracellular Ca(2+) levels and that MK-801 blocks NMDAR-dependent LTD in the hippocampus of both adult and immature mice. Moreover, MK-801 inhibits NMDAR-mediated activation of p38-MAPK and dephosphorylation of AMPAR GluA1 subunits at sites implicated in LTD. Thus, our results indicate that the induction of LTD in the hippocampal CA1 region is dependent on ionotropic, rather than metabotropic, NMDAR signaling.

Keywords: MK-801; NMDA receptor; hippocampus; long-term depression.

Figure 1.

MK-801 blocks LTD in the hippocampal CA1 region of adult mice. A, LTD induced by 1 Hz LFS (open circles, n = 6) is blocked by 10 μm MK-801 (filled circles, n = 8) or 50 μm d-APV (triangles, n = 6). MK-801 or d-APV was present throughout the experiment. Traces are superimposed fEPSPs recorded during baseline and 45 min after LFS. B, Changes in synaptic strength induced by LFS in slices bathed in ACSF containing 2 mm CaCl₂ (open bar, n = 8) or 4 mm CalCl₂ (filled bars, *p < 0.005 compared with pre-LFS baseline). C, HFS-induced LTP (open circles, n = 9) is inhibited by MK-801 (filled circles, n = 8) or d-APV (triangles, n = 6). Traces show superimposed fEPSPs recorded during baseline and 45 min after HFS. D, chem-LTD is blocked by MK-801. Bath application of 20 μm NMDA induced LTD in interleaved control experiments (open circles, n = 6) but had no effect on synaptic strength in MK-801-treated slices (filled circles, n = 4, p < 0.001 compared with control). Traces are superimposed fEPSPs recorded during baseline and 50 min after NMDA application. E, chem-LTD is blocked when NMDA is applied in Ca²⁺-free ACSF (no added CaCl₂). A 15 min bath application of Ca²⁺-free ACSF alone (shaded region) transiently abolished synaptic transmission but had no lasting effect on synaptic strength (squares, n = 8). Bath application of NMDA induced LTD in interleaved control experiments (open circles, n = 6) but had no lasting effect on synaptic strength when applied in Ca²⁺-free ACSF (filled circles, n = 6). Traces show superimposed fEPSPs recorded during baseline and 50 min after NMDA application in Ca²⁺-free ACSF (right) in control experiments (left). Calibration: A, D, E, 1 mV, 5 ms; C, 2 mV, 5 ms.

Figure 2.

MK-801 blocks LTD induction in the hippocampal CA1 region of slices from young animals. A, MK-801 blocks 1 Hz LFS-induced LTD in slices from young C57BL/6 mice (open circles, interleaved control experiments, n = 7; filled circles, LFS in 10 μm MK-801, n = 6, p < 0.05 compared with control). B, MK-801 blocks 2 Hz LFS-induced LTD in slices obtained from young CD1 mice. Slices were incubated in ACSF containing 50 μm MK-801 for 2–6 h before 2 Hz LFS, and MK-801 was washed out after LFS (filled symbols, n = 8, p < 0.05 compared with control). Open symbols indicate results from interleaved control experiments (n = 8). Traces in A and B show superimposed fEPSPs recorded during baseline and after LFS in control (bottom) and MK-801 (top) experiments. Calibration: A, 1 mV, 10 ms; B, 0.5 mV, 10 ms.

Figure 3.

MK-801 blocks activation of p38-MAPK and AMPAR GluA1 subunit dephosphorylation induced by NMDAR activation. A, MK-801 blocks NMDA-induced increases in p38-MAPK phosphorylation and GluA1 dephosphorylation at serine 845 and threonine 840 in CA1 mini slices from adult mice ([Ca²⁺]_o = 4.0 mm, n = 6). Although basal levels of GluA1 S845 phosphorylation tended to be elevated in MK-801-treated slices, this difference was not statistically significant (p = 0.065). UT, Untreated slices; N, NMDA treated slices. B, NMDA-induced increases in p38-MAPK phosphorylation and GluA1 phosphorylation are blocked by MK-801 in hippocampal slices from young mice ([Ca²⁺]_o = 2.0 mm, n = 5). *p < 0.05 compared with untreated control slices.