Fig 2: Synaptic NMDAR potentiation critically depends on MMP-3 and integrin signaling but not MMP-2/9. (a) Isolation of synaptic NMDAR-mediated fEPSPs (fEPSPNMDA). Upper, Sample traces of fEPSPs recorded in Mg2+-free solution before (gray) and after (black) application of the AMPA/kainate antagonist DNQX (20 µM) and l-type channel blocker nifedipine (20 µM). Lower, fEPSPNMDA (gray) was completely abolished upon bath application of APV (50 µM) (black). (b 1) Exemplary traces of fEPSPNMDA before HFS (1) and 90 min after HFS (colors match (b 2) legend). Scale bar = 0.2 mV, 20 ms. (b 2) Upper, Time-course of synaptic NMDAR-mediated fEPSP under control conditions (black) and upon bath application of MMP-3 inhibitor before HFS (red), 15 min post-HFS (green), or 30 min post-HFS (yellow). Lower, Time-course of synaptic NMDAR-mediated fEPSP under control conditions (black) and upon bath application of drugs before HFS: MMP-3 inhibitor UK356618 (2 µM) (magenta), MMP-2/9 inhibitor (10 µM) (cyan), or integrin-blocking peptide GRGDSP (30 min post-HFS) (gray). The zero value on the time bars represents the moment of tetanization (HFS, 4 × 100 Hz). The horizontal colored bars represent drug application. (c) Statistics of effects of drugs shown in (b 2) measured 60 min post-HFS. The numbers on the graphs refer to the number of experiments. *p < 0.05

Fig 3: Temporal dependence of hippocampal E-S potentiation on NMDARs and MMP-3 activity. a Recording scheme. Two recording (REC) electrodes simultaneously monitored fEPSPs and population spikes in the CA1 region in response to Schaffer collateral stimulation (STIM). b Left, Time-course of maximal fEPSP slopes normalized to baseline values in control slices (black circles) and when bath-applied with the NMDAR antagonist APV (50 µM) at varying time points relative to HFS: before HFS (red triangles), 15 min post-HFS (green squares), or 60 min post-HFS (blue diamonds). The top shows exemplary traces of fEPSPs before HFS (1) and 90 min after HFS (colors match figure legend). Scale bar = 0.5 mV, 20 ms. Right, Time-course of population spike (PS) amplitudes normalized to baseline values in control slices (black circles) and when bath-applied with the NMDAR antagonist APV (50 µM) at varying time points relative to HFS: before HFS (red triangles), 15 min post-HFS (green squares), or 60 min post-HFS (blue diamonds). The top shows exemplary and normalized traces of PS before HFS (1) and 90 min after HFS (colors match figure legend). Scale bar = 0.5 mV, 10 ms. c Statistics of average PS/fEPSP ratio presented in b at 90 min post-HFS. The asterisk indicates a significant difference vs. slices in which HFS was applied in the absence of APV. Notice that APV that was applied up to 30 min post-HFS (green) significantly attenuated the PS/fEPSP upregulation following HFS. d Impact of APV on E-S coupling before HFS (circles) and 90 min after HFS (triangles). Relationships between stimulus strength and the PS/fEPSP slope ratio are shown. Notice that APV that was applied up to 30 min post-HFS (yellow) significantly attenuated the PS/fEPSP upregulation following HFS that was observed in CTR (black). e Effect of MMP-3 inhibitor NNGH (10 µM) on E-S potentiation. Middle and right, fEPSP slope and population spike amplitude time-course, respectively, as in b, except instead of APV, an MMP-3 inhibitor was applied at varying time points with regards to HFS. Left, Exemplary traces of fEPSPs (top) and population spikes (bottom) before and 90 min after HFS in the presence of NNGH (colors match figure legends). Scale bars as in b. f Statistics of average PS/fEPSP ratio presented in e at 90 min post-HFS. The asterisk indicates a significant difference vs. slices in which HFS was applied in the absence of NNGH. Notice that NNGH that was applied up to 15 min post-HFS (green) significantly attenuated the PS/fEPSP upregulation following HFS. g Impact of NNGH on E-S coupling before HFS (circles) and 90 min after HFS (triangles). Relationships between stimulus strength and the PS/fEPSP slope ratio are shown. Notice that NNGH that was applied up to 15 min post-HFS (green) significantly attenuated the PS/fEPSP upregulation following HFS that was observed in CTR (black). The zero value on the time bars represents the moment of tetanization (HFS, 4 × 100 Hz). The horizontal colored bars represent drug application. The numbers on the graphs refer to the number of experiments. *p < 0.05

Fig 4: MMP-3 or NMDAR inhibition affects c-Fos expression following patterned neuronal activity. a Exemplary confocal immunofluorescence images of CA1 hippocampal region (×40 magnification) in acute brain slices fixed immediately following cessation of the electrophysiological recordings. Basally stimulated slices (0.1 Hz) (upper panels) were compared with slices fixed 90 min post-E-S potentiation induction in the absence (middle panel) or presence (lower panel) of APV. The sections were stained against cFos (green channel) and NeuN (red channel), and the colocalization of both markers was analyzed (merged channel). b Statistics of the effects of electrical stimulation and drug treatment on cFos expression in the CA1 neuronal population stained with NeuN. Slices were basally stimulated (0.1 Hz) (gray), or E-S potentiation was induced with HFS (4 × 100 Hz) in the absence (LTP) (black) or presence of the NMDAR antagonist APV applied before or 15 and 30 min after HFS. c Exemplary immunofluorescence images of CA1 sections following the induction of synaptic LTPNMDA (see Fig. 2). Basally stimulated slices (0.1 Hz) (upper panels) were compared with slices that were fixed 90 min post-LTPNMDA induction with HFS in the absence (middle panel) or presence (lower panel) of the MMP-3 inhibitor NNGH. See a for channel description. d Statistics of the effects of electrical stimulation and NNGH treatment on cFos expression in CA1 neurons following recordings of synaptic NMDAR-mediated responses (shown in Fig 2). Slices were basally stimulated (0.1 Hz) (gray), or LTPNMDA was induced with HFS (4 × 100 Hz) in the absence (LTP) (black) or presence of the MMP-3 inhibitor NNGH applied before or 15 and 30 min after HFS. The numbers on the graphs refer to the number of sections analyzed. *p < 0.05 vs. basally stimulated slices