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, 121 (2), 609-619

LTD at Mossy Fiber Synapses Onto Stratum Lucidum Interneurons Requires TrkB and Retrograde Endocannabinoid Signaling

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LTD at Mossy Fiber Synapses Onto Stratum Lucidum Interneurons Requires TrkB and Retrograde Endocannabinoid Signaling

Enhui Pan et al. J Neurophysiol.

Abstract

Hippocampal mossy fiber axons simultaneously activate CA3 pyramidal cells and stratum lucidum interneurons (SLINs), the latter providing feedforward inhibition to control CA3 pyramidal cell excitability. Filopodial extensions of giant boutons of mossy fibers provide excitatory synaptic input to the SLIN. These filopodia undergo extraordinary structural plasticity causally linked to execution of memory tasks, leading us to seek the mechanisms by which activity regulates these synapses. High-frequency stimulation of the mossy fibers induces long-term depression (LTD) of their calcium-permeable AMPA receptor synapses with SLINs; previous work localized the site of induction to be postsynaptic and the site of expression to be presynaptic. Yet, the underlying signaling events and the identity of the retrograde signal are incompletely understood. We used whole cell recordings of SLINs in hippocampal slices from wild-type and mutant mice to explore the mechanisms. Genetic and pharmacologic perturbations revealed a requirement for both the receptor tyrosine kinase TrkB and its agonist, brain-derived neurotrophic factor (BDNF), for induction of LTD. Inclusion of inhibitors of Trk receptor kinase and PLC in the patch pipette prevented LTD. Endocannabinoid receptor antagonists and genetic deletion of the CB1 receptor prevented LTD. We propose a model whereby release of BDNF from mossy fiber filopodia activates TrkB and PLCγ1 signaling postsynaptically within SLINs, triggering synthesis and release of an endocannabinoid that serves as a retrograde signal, culminating in reduced glutamate release. Insights into the signaling pathways by which activity modifies function of these synapses will facilitate an understanding of their contribution to the local circuit and behavioral consequences of hippocampal granule cell activity. NEW & NOTEWORTHY We investigated signaling mechanisms underlying plasticity of the hippocampal mossy fiber filopodial synapse with interneurons in stratum lucidum. High-frequency stimulation of the mossy fibers induces long-term depression of this synapse. Our findings are consistent with a model in which brain-derived neurotrophic factor released from filopodia activates TrkB of a stratum lucidum interneuron; the ensuing activation of PLCγ1 induces synthesis of an endocannabinoid, which provides a retrograde signal leading to reduced release of glutamate presynaptically.

Keywords: BDNF; TrkB; hippocampus; interneuron; synaptic plasticity.

Conflict of interest statement

No conflicts of interest, financial or otherwise, are declared by the authors.

Figures

Fig. 1.
Fig. 1.
TrkB signaling is required for mossy fiber calcium permeable-AMPA receptor (mossy fiber-CP-AMPAR) long-term depression (LTD). A, top: representative traces (inset) show individual excitatory postsynaptic currents (EPSCs) at time points collected 10 min before (1, black) and between 10 and 20 min after (2, gray) application of high-frequency stimulation (HFS) in either wild-type (WT; □) or TrkB−/− (■) mice. Top left: HFS produced LTD of mossy fiber-CP-AMPAR stratum lucidum interneuron synapses in WT (65 ± 7%, n = 6, paired t-test, P = 0.003) but not in TrkB null mice (144 ± 30%, n = 8, paired t-test, P = 0.2). Top right: results of individual cells are plotted. Comparison of WT and TrkB−/− groups with Student’s t-test revealed a significant difference (*P = 0.01). Bottom: repeated-measures two-way ANOVA revealed a P value of 0.005 for paired-pulse ratio (PPR) data in the two groups. Post hoc paired t-tests revealed increased PPR in cells from WT (PPR was 1.5 ± 0.2 before and 1.9 ± 0.4 after HFS, n = 6, *P = 0.03) but not TrkB−/− mice (PPR was 1.3 ± 0.1 before and 1.0 ± 0.1 after HFS, n = 8, P = 0.1). B: chemical-genetic approach reveals requirement of TrkB kinase for mossy fiber-CP-AMPAR LTD in slices from TrkBF616A.mice. Inset presents representative traces of EPSCs collected 10 min before (1, black) and between 10 and 20 min after (2, gray) HFS of mossy fibers (□, vehicle group; ■, 2 μM 1NMPP1). Top left: treatment with 1NMPP1 impaired mossy fiber-CP-AMPAR LTD (105 ± 7%, n = 10, paired t-test, P = 0.3) compared with vehicle (68 ± 7%, n = 10, paired t-test, P = 0.001). Top right: results of individual animals are plotted. *P = 0.01, vehicle vs. 1NMPP1 groups (Student’s t-test). Bottom: repeated-measures two-way ANOVA revealed P = 0.089 for PPR data in the two groups.
Fig. 2.
Fig. 2.
Synaptically released zinc is not required for mossy fiber calcium permeable-AMPA receptor (mossy fiber-CP-AMPAR) long-term depression (LTD). A, top: representative traces (inset) show individual excitatory postsynaptic currents (EPSCs) at time points before (1, black) and after (2, gray) high-frequency stimulation (HFS) of mossy fibers. Top left: HFS induced similar amounts of LTD in the presence of vehicle (63 ± 9%, n = 5, P = 0.02, paired t-test) or ZX1 (59 ± 6%, n = 9, P = 0.0002, paired t-test). Top right: values from individual cells are plotted. Comparison of vehicle- and ZX1-treated groups with Student’s t-test revealed no significant differences (P = 0.4). Bottom: repeated-measures two-way ANOVA revealed a P value of 0.227 for paired-pulse ratio (PPR) data in the two groups. B, top: representative traces (inset) of mossy fiber-CP-AMPAR EPSC were collected before (1, black) and after (2, gray) HFS of mossy fibers. Top left: HFS induced LTD of mossy fiber-CP-AMPAR EPSC in slices from both WT (77 ± 8%, n = 5, paired t-test, P = 0.02) and ZnT3−/− mice (76 ± 4%, n = 10, paired t-test, P = 0.002). Top right: values from individual cells are plotted. Comparison of WT and ZnT3−/− mice with Student’s t-test revealed no significant difference (P = 0.5). Bottom: repeated-measures two-way ANOVA revealed a P value of 0.564 for PPR data in the two groups.
Fig. 3.
Fig. 3.
Brain-derived neurotrophic factor (BDNF) is required for mossy fiber calcium permeable-AMPA receptor (mossy fiber-CP-AMPAR) long-term depression (LTD). A, top: representative traces (inset) of mossy fiber-CP-AMPAR excitatory postsynaptic current (EPSC) were collected before (1, black) and after (2, gray) high-frequency stimulation (HFS) of mossy fibers. Top left: HFS induced LTD of mossy fiber-CP-AMPAR synapses in the presence of control (72 ± 6%, n = 10, paired t-test, P = 0.005) but not in TrkB-Ig groups (98 ± 7%, n = 7, paired t-test, P = 0.4). Controls included 5 human IgG and 5 vehicle experiments; these were pooled for presentation because similar results were obtained with each. Top right: values from individual experiments are plotted. Comparison of control vs. TrkB-Ig with Student’s t-test revealed significant differences (*P = 0.005). Bottom, repeated-measures two-way ANOVA revealed a P value of 0.368 for paired-pulse ratio (PPR) data in the two groups. B, top: representative traces (inset) of mossy fiber-CP-AMPAR EPSC were collected before (1, black) and after HFS (2, gray). Top left: HFS induced LTD of mossy fiber-CP-AMPA synapse in slices from WT (63 ± 8%, n = 7, paired t-test, P = 0.002) and conditional BDNF mutant mice (88 ± 6%, n = 6, paired t-test, P = 0.04). Top right: values from individual experiments are plotted. Comparison of the LTD evident in WT vs. conditional BDNF mutant mice with Student’s t-test revealed a significant difference (P = 0.01). Bottom: repeated-measures two-way ANOVA revealed a P value of 0.06 for PPR data in the two groups.
Fig. 4.
Fig. 4.
Inclusion of inhibitors of Trk/PLC signaling in stratum lucidum interneurons inhibits mossy fiber calcium permeable-AMPA receptor (mossy fiber-CP-AMPAR) long-term depression (LTD). A, top left: inclusion of K252a (200 nM) in the recording pipette prevented high-frequency stimulation (HFS)-induced mossy fiber-CP-AMPAR LTD (vehicle: 57 ± 2%, n = 6, paired t-test, P = 0.00002; K252a: 92 ± 9%, paired t-test, P = 0.2). Top right: results of individual cells are plotted. Comparison of vehicle vs. K252a groups with Student’s t-test revealed a significant difference (*P = 0.007). Bottom: repeated-measures two-way ANOVA revealed a P value of 0.029 for paired-pulse ratio (PPR) data in the two groups. Post hoc paired t-tests revealed increased PPR in cells from vehicle (1.7 ± 0.2 before and 2.1 ± 0.4 after, paired t-test, P = 0.03) but not K2525a (1.5 ± 0.2 before and 1.3 ± 0.3 after, paired t-test, P = 0.2). B, top left: inclusion of U-73122 (5 μM) in recording pipette prevented HFS-induced mossy fiber-CP-AMPAR LTD (vehicle: 74 ± 5%, n = 5, paired t-test, P = 0.003; U-73122: 109 ± 7%, n = 10, paired t-test, P = 0.11). Top right: results of individual cells are plotted. Comparison of vehicle vs. U-73122 groups with Student’s t-test revealed a significant difference (*P = 0.0005). Bottom: repeated-measures two-way ANOVA revealed a P value of 0.058 for PPR data in the two groups. C, top left: inclusion of pY816 peptide (10 µM) in recording pipette prevented HFS-induced mossy fiber CP-AMPAR LTD (Scramble peptide: 73 ± 5, n = 6, paired t-test, P = 0.003; pY816 peptide: 96 ± 9, n = 10, paired t-test, P = 0.5). Top right: results of individual cells are plotted. Comparison of scrambled vs. pY816 peptide groups with Student’s t-test revealed a significant difference (*P = 0.04). Bottom: repeated-measures two-way ANOVA revealed a P value of 0.125 for PPR data in the two groups.
Fig. 5.
Fig. 5.
Bath application of CB1 receptor antagonists prevents mossy fiber calcium permeable-AMPA receptor (mossy fiber-CP-AMPAR) long-term depression (LTD). A, top left: inclusion of AM251 (5 μM) prevented high-frequency stimulation (HFS)-induced mossy fiber-CP-AMPAR LTD (vehicle: 61 ± 10%, n = 8, paired t-test, P = 0.003; AM251: 102 ± 6%, n = 10, paired t-test, P = 0.2). Top right: results of individual cells are plotted. Comparison of vehicle vs. AM251 groups with Student’s t-test revealed a significant difference (*P = 0.002). Bottom: repeated-measures two-way ANOVA revealed a P value of 0.124 for paired-pulse ratio (PPR) data in the two groups. B, top left: inclusion of SR141716A (5 μM) prevented HFS-induced mossy fiber-CP-AMPAR LTD (vehicle: 66 ± 5%, n = 9, paired t-test, P = 0.0001; SR141716A: 109 ± 6%, n = 8, paired t-test, P = 0.08). Top right: results of individual cells are plotted. Comparison of vehicle vs. SR141716A groups with Student’s t-test revealed a significant difference (*P = 0.00005). Bottom: repeated-measures two-way ANOVA revealed a P value of 0.205 for PPR data in the two groups.
Fig. 6.
Fig. 6.
High-frequency stimulation (HFS)-mediated induction of mossy fiber calcium permeable-AMPA receptor (mossy fiber-CP-AMPAR) long-term depression (LTD) is impaired in slices from CB1 receptor knockout (CB1R−/−) mice. Top left: HFS induced LTD in slices from wild-type (WT) but not CB1R−/− mice (WT: 73 ± 8%, n = 7, paired t-test, P = 0.006; CB1R−/−: 129 ± 21%, n = 6, paired t-test, P = 0.1). Top right: results of individual cells are plotted. Comparison of vehicle vs. CB1R−/− mice with Student’s t-test revealed a significant difference (*P = 0.02). Bottom: repeated-measures two-way ANOVA revealed a P value of 0.309 for paired-pulse ratio (PPR) data in the two groups.

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