Input-specific presynaptic spike timing-dependent LTD at lateral and medial perforant pathway-dentate gyrus granule cell synapses.

(a, k) Schemes show the general set-up for the electrophysiological recordings and pairing protocol (Δt = -18 ms, time between the peak of the postsynaptic spike and the onset of the EPSP) at both LPP- and MPP-GC synapses: CA, cornus ammonis; DG, dentate gyrus; EC, entorhinal cortex; S1 and S2, stimulating electrodes; R, recording electrode. A post-before-pre pairing protocol induced t-LTD at both LPP-GC (b) and MPP-GC (l) synapses. The EPSP slopes monitored in the paired (LPP-GC synapses black triangles, n = 14; MPP- GC synapses open black triangles, n = 14) and unpaired (LPP-GC synapses red symbols; MPP-GC synapses open red symbols) pathways are shown. The traces show the EPSP before (1) and 30 minutes after (2) pairing. Depression was only observed in the paired pathways. (c, m) Summary of the results. At the lateral (d-j) and medial (n-t) perforant pathway synapses onto dentate gyrus granule cells t-LTD is expressed presynaptically. The number of failures increases after t-LTD induction at both LPP- (n = 8, d) and MPP-GC (n = 6, n) synapses. Traces show the EPSPs before (black) and 30 min after pairing (red). PPR increases after t-LTD pairing protocol at LPP-GC (e) and MPP-GC (o) synapses. The traces show the EPSP before (black) and 30 min after t- LTD induction (red). (f, p) Normalized plots of CV−2 versus mean EPSP slopes yield data points mainly below the diagonal after t-LTD induction at both LPP- (n = 14) and MPP-GC (n = 13) synapses. The traces show the EPSPs before (black) and 30 min after t-LTD induction (red). Miniature EPSPs (mEPSPs) were monitored at baseline and after t-LTD induction in the presence of TTX (300 nM) at LPP- (g-j, n = 6) and MPP-GC synapses (q-t, n = 6). Cumulative graphs and histograms show that after t-LTD induction, the frequency of mEPSPs decreases at both types of synapses, whereas the amplitude of the mEPSPs remains constant. Scale bar for the % failures - 5 mV, 50 ms; scale bar for PPR - 5 mV, 80 ms. Error bars indicate the S.E.M.: * p < 0.05, **p < 0.01, *** p < 0.001, two-sided Student’s t-test.

The t-LTD at lateral and medial perforant pathway-dentate gyrus granule cell synapses has distinct requirements for glutamate receptors.

The t-LTD at LPP-GC synapses requires mGluR5. (a) The addition of D-AP5 (50 μM) or MK-801 (500 μM) to the superfusion fluid does not prevent the induction of t-LTD at LPP-GC synapses. The EPSP slopes shown are from D-AP5-treated (red triangles, n = 8), MK- 801-treated (grey triangles, n = 6) and untreated cells (black triangles, n = 8). The traces show EPSPs before (1) and 30 min after (2) pairing. (b) Summary of the results. (c) The t-LTD at LPP-GC synapses requires mGluR5. The EPSP slopes are shown from control slices (black triangles, n = 9), and slices treated with the mGluR antagonist LY341495 (100 µM, red squares, n = 8), the mGluR1 antagonist LY367385 (100 µM, grey triangles, n = 6) or the mGluR5 antagonist MPEP (20 µM, blue squares, n = 8). The traces show the EPSPs before (1) and 30 min after (2) pairing. (d) Summary of the results. (e) The t-LTD at MPP-GC synapses requires NMDARs containing GluN2A subunits and mGluR1. The addition of D-AP5 (50 µM) or MK-801 (500 µM) to the superfusion fluid prevented t-LTD induction at MPP-GC synapses, whereas postsynaptic loading of MK-801 (500 µM) does not block t-LTD induction. The EPSP slopes are shown from D-AP5 (open red squares, n = 10) or MK-801 treated cells (batch, open grey squares, n = 6; loaded postsynaptic cell, open blue triangles, n = 9), and untreated cells (open black triangles, n = 8). The traces show the EPSPs before (1) and 30 min after (2) pairing. (f) Summary of the results. (g) The NMDARs involved in t- LTD at MPP-GC synapses contain GluN2A subunits. The t-LTD at MPP-GC synapses was completely blocked in slices exposed to Zn2+ (300 nM), while it remained unaffected in slices treated with Ro 25-6981 (0.5 µM), ifenprodil (3 µM) or PPDA. The EPSP slopes shown are from control slices (open black triangles, n = 9) and slices treated with Zn2+ (open red squares, n = 7), Ro-25-6981 (open grey triangles, n = 6), ifenprodil (open blue triangles, n = 6) or PPDA (open pink triangles, n = 6). The traces show the EPSPs before (1) and 30 min after (2) pairing. (h) Summary of the results. (i) The t-LTD at MPP-GC synapses requires mGluR1. The EPSP slopes shown are from control slices (open black triangles, n = 9), or slices treated with LY341495 (100 µM, open red squares, n = 10), LY367385 (100 µM, open grey squares, n = 7) or MPEP (20 µM, open blue triangles, n = 6). The traces show the EPSPs before (1) and 30 min after (2) pairing. (j) Summary of the results. The error bars indicate the S.E.M.: *p < 0.05, ** p < 0.01, ***p < 0.001, One-way ANOVA + Holm–Sidak test.

The t-LTD at lateral and medial perforant pathway-dentate gyrus granule cell synapses requires postsynaptic calcium and eCB signalling.

(a) The induction of t-LTD at LPP-GC synapses in slices was unaffected by exposure to nimodipine (10 µM) but it was prevented by treatment with thapsigargin (10 µM), either in the superfusion fluid or when loaded into the postsynaptic cell. The EPSP slopes shown are from control slices (black triangles, n = 9) and slices treated with nimodipine (red triangles, n = 6), thapsigargin in the superfusion fluid (grey squares, n = 6) or thapsigargin loaded into the postsynaptic cell (blue squares, n = 9). The traces show EPSPs before (1) and 30 min after (2) pairing. (b) Summary of the results. (c) The t- LTD at LPP-GC synapses requires endocannabinoids and CB1 receptors, as it was blocked when THL was loaded into the postsynaptic neuron (pTHL, 5 µM) and in slices treated with AM251 (3 µM). The EPSP slopes shown are from control slices (black triangles, n = 14) and slices with THL loaded into the postsynaptic neuron (red squares, n = 8) or treated with AM251 (grey squares, n = 14). The traces show the EPSPs before (1) and 30 min after (2) pairing. (d) Summary of the results. (e) The induction of t-LTD at MPP-GC synapses was unaffected in nimodipine-treated slices (10 µM) but prevented in slices treated with thapsigargin (10 µM), either in the superfusion fluid or when loaded into the postsynaptic cell. The EPSP slopes shown are from control slices (open black triangles, n = 9) and slices treated with nimodipine (open red triangles, n = 6), thapsigargin in the superfusion fluid (open grey squares, n = 6) or thapsigargin loaded into the postsynaptic cell (open blue squares, n = 7). The traces show EPSPs before (1) and 30 min after (2) pairing. (f) Summary of the results. (g) The t-LTD at MPP-GC synapses requires endocannabinoids and CB1 receptors, and it was blocked when THL was loaded into the postsynaptic neuron (pTHL, 5 µM) and in slices treated with AM251 (3 µM). The EPSP slopes shown are from control slices (open black triangles, n = 8) and slices with THL loaded into the postsynaptic neuron (open red squares, n = 6) or treated with AM251 (open grey squares, n = 6). The traces show EPSPs before (1) and 30 min after (2) pairing. (h) Summary of the results. The error bars indicate S.E.M. *p < 0.05, ** p < 0.01, *** p < 0.001, One-way ANOVA + Holm– Sidak test.

The t-LTD at lateral and medial perforant pathway-dentate gyrus granule cell synapses requires astrocytes and glutamate.

(a, f) Schemes showing the general experimental set-up: A, astrocyte; CA, cornus ammonis; DG, dentate gyrus; EC, entorhinal cortex; GC, granule cell; S1 and S2, stimulating electrodes; R1 and R2, recording electrodes. Inset: voltage responses of an astrocyte shown in current clamp configuration. (b) The t-LTD at LPP-GC synapses requires astrocytes as it was prevented by loading astrocytes with BAPTA (20 mM) and in slices from dnSNARE mice. The EPSP slopes shown are from control slices (black triangles, n = 7), slices with BAPTA-loaded astrocytes (red squares, n = 6) and slices from dnSNARE mice (grey circles, n = 9). The traces show the EPSPs before (1) and 30 min after (2) pairing. (c) Summary of the results. (d) The t-LTD at LPP-GC synapses requires glutamate as it is absent in slices from dnSNARE mice but it is restored by glutamate puffs (100 µM). The EPSP slopes shown are from control slices (black triangles, n = 7), slices from dnSNARE mice (red circles, n = 7) and slices from dnSNARE mice administered glutamate puffs (grey triangles, n = 6). The traces show the EPSPs before (1) and 30 min after (2) pairing. (e) Summary of the results. (g) The t-LTD at MPP-GC synapses requires astrocytes as t-LTD induction at MPP-GC synapses was prevented by loading astrocytes with BAPTA (20 mM) and in slices from dnSNARE mice. The EPSP slopes shown are from control slices (open black triangles, n = 6), slices with BAPTA-loaded astrocytes (open red squares, n = 6) and slices from dnSNARE mice (open grey circles, n = 7). The traces show the EPSPs before (1) and 30 min after (2) pairing. (h) Summary of the results. (i) The t-LTD at MPP-GC synapses requires glutamate as it is absent in slices from dnSNARE mice but is restored by glutamate puffs (100 µM). The EPSP slopes shown are from control slices (open black triangles, n = 7), slices from dnSNARE mice (open red circles, n = 6) and slices from dnSNARE mice administered to glutamate puffs (open grey triangles, n = 6). The traces show the EPSPs before (1) and 30 min after (2) pairing. (j) Summary of the results. The error bars indicate the S.E.M.: *p < 0.05, **p < 0.01, ***P < 0.001, One-way ANOVA + Holm–Sidak test.