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. 2016 Mar 18;7:10923.
doi: 10.1038/ncomms10923.

Rapid Erasure of Hippocampal Memory Following Inhibition of Dentate Gyrus Granule Cells

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Free PMC article

Rapid Erasure of Hippocampal Memory Following Inhibition of Dentate Gyrus Granule Cells

Noelia Madroñal et al. Nat Commun. .
Free PMC article

Abstract

The hippocampus is critical for the acquisition and retrieval of episodic and contextual memories. Lesions of the dentate gyrus, a principal input of the hippocampus, block memory acquisition, but it remains unclear whether this region also plays a role in memory retrieval. Here we combine cell-type specific neural inhibition with electrophysiological measurements of learning-associated plasticity in behaving mice to demonstrate that dentate gyrus granule cells are not required for memory retrieval, but instead have an unexpected role in memory maintenance. Furthermore, we demonstrate the translational potential of our findings by showing that pharmacological activation of an endogenous inhibitory receptor expressed selectively in dentate gyrus granule cells can induce a rapid loss of hippocampal memory. These findings open a new avenue for the targeted erasure of episodic and contextual memories.

Figures

Figure 1
Figure 1. Rapid and selective inhibition of DG neurotransmission in vivo.
(a) The hippocampal tri-synaptic circuit receives PP inputs from entorhinal cortex to DG, CA3 and CA1. (b) A stimulating electrode was implanted in the PP and a recording electrode in CA3 pyramidal layer. (c) Strength of CA3 pyramidal layer fEPSPs evoked in anaesthetized mice by electrical stimulation of PP inputs showed fast and slow latency population spike components corresponding to direct PP-CA3 and indirect PP–DG-CA3 inputs, respectively. Systemic administration of the selective Htr1a agonist, 8-OH-DPAT (0.3 mg kg−1, subcutaneous), to Htr1aDG (Tg) mice caused a rapid and selective decrease in the long-latency component that persisted for several hours. Quantification indicated a significant decrease in DG neurotransmission following agonist treatment of Htr1aDG, but not Htr1aKO (KO) littermates or vehicle treated wild-type mice that reached 80% suppression and persisted for >2 h (mean±s.e.m.; n=10; *P<0.05; two-way analysis of variance followed by Holm–Sidak post hoc test). (d) Representative fEPSPs evoked at CA3 pyramidal layer after stimulation of PP inputs before and after agonist treatment. The fast and the slow latency population spike components are indicated (black arrow, short; grey arrow, long).
Figure 2
Figure 2. Inhibition of DG induces rapid and persistent loss of hippocampal memory and plasticity.
(a) Stimulating and recording electrodes were implanted in the SC region and CA1 pyramidal layer, respectively. Eye-blink conditioning was followed using electromyographic (EMG) measurement of orbicularis oculi (O.O.) muscle activity following electrical stimulation of the same muscle. (b) EMG was recorded during trace conditioning. Top: Conditioned Stimulus (CS), tone, 2.4 kHz, 85 dB. Middle: Unconditioned Stimulus (US), shock, 3 × threshold; 500 ms interval. Bottom: representative conditioned response in trained animal. (c) Evoked fEPSP responses were obtained to stimuli delivered to SC inputs in Htr1aDG (Tg) and Htr1aKO (KO) control mice. (d) Two days of habituation (60 presentations/day, tone only) were followed by 12 days of conditioning (60 presentations/day, 90% tone+shock and 10% tone only) during which animals developed stable conditioned eye-blink responding. A parallel increase in (e) SC plasticity was seen during conditioning of wild-type (WT), Htr1aDG and Htr1aKO mice. Injection of the Htr1a agonist 8-OH-DPAT to Htr1aDG, but not Htr1aKO littermates before conditioning on day 8 lead to a significant reduction in (d) conditioned responding and (e) SC plasticity. No change in (d) conditioned responding or (e) plasticity was seen in vehicle treated wild-type mice. Following agonist injection, conditioned behaviour and CA3-CA1 plasticity in the Htr1aDG group continued to be significantly reduced and showed a recovery (days 9–12) similar to that seen during initial conditioning (days 1–7). (f,g) Mice were trained to tone–shock presentations on days 1–8 and 14–17. After treatment with 8-OH-DPAT on day 8, mice were left in their home cages on days 9–13. Both (f) conditioned responding and (g) SC plasticity remained suppressed in the Htr1aDG group when compared with Htr1aKO mice on day 14 (mean±s.e.m.; n=9–10; *P<0.05; two-way analysis of variance followed by Holm–Sidak post hoc test).
Figure 3
Figure 3. Memory loss depends on paired CS–US presentations.
Htr1aDG or Htr1aKO mice were exposed (a,b) to tone–shock pairs on days 1–7 and 9–10, but left in the home cage on day 8, (c,d) to paired tone–shock presentations on days 1–7 and 9–10, but received tone only presentations on day 8, or (e,f) to tone–shock pairs on days 1–7 and 9–10, but received unpaired tone and shock presentations on day 8. Under these conditions treatment with 8-OH-DPAT on day 8 failed to cause a decrease in (a,c,e) conditioned responding or (b,d,f) fEPSP responses in either Htr1aDG or Htr1aKO mice (mean±s.e.m.; n=9–10; two-way analysis of variance followed by Holm–Sidak post hoc test).
Figure 4
Figure 4. Loss of plasticity depends on entorhinal cortex inputs and local adenosine signalling.
(ac) Mice locally infected with an (a) AAV expressing the Venus fluorescent protein and HA-tagged hM4D DREADD receptor (HA-hM4D) for pharmacogenetic neural inhibition under control of the Synapsin (Syn) gene promoter showed expression in (b) entorhinal cortex (EC) and PP projections to hippocampus (DG, CA3 and CA1). (c) Systemic administration of the selective hM4D agonist CNO (3 mg kg−1, intraperitoneal) to wild-type mice caused a significant decrease in fEPSP recorded in CA1 following stimulation of olfactory bulb-entorhinal cortex projections in animals infected with AAV-hM4D, but not in non-infected (control) animals. Significant suppression of fEPSPs was observed 1 h after agonist treatment and persisted for at least 2 h (mean±s.e.m.; n=4–7; *P<0.05; two-way analysis of variance followed by Holm–Sidak post hoc test). (d,e) Htr1aDG (Tg) mice (d) expressing hM4D in the entorhinal cortex and its projections to hippocampus (red arrows) and implanted with stimulating and recording electrodes in the SC and CA1 regions, respectively, were (e) subjected to trace eye-blink conditioning with tone–shock presentations on days 1–12. Systemic pre-treatment with the hM4D agonist CNO (3 mg kg−1, intraperitoneal) on day 8 reversed the loss of SC plasticity induced by 8-OH-DPAT treatment on that day (mean±s.e.m.; n=12; *P<0.05; two-way analysis of variance followed by Holm–Sidak post hoc test). (f,g) Htr1aDG mice (f) implanted with stimulating and recording electrodes in the SC and CA1 regions, respectively, and bilateral cannulae aimed at dorsal CA1 were (g) subjected to trace eye-blink conditioning as above. Pre-treatment on day 8 with the adenosine A1 receptor antagonist, DPCPX (82 nmol) caused a significant reversal of the SC depotentiation induced by 8-OH-DPAT treatment on that day when compared with vehicle pretreated mice. Notably, reversal of DG inhibition-induced plasticity by DPCPX persisted through day 12 (mean±s.e.m.; n=7; *P<0.05; two-way analysis of variance followed by Holm–Sidak post hoc test).
Figure 5
Figure 5. Inhibition of DG induces persistent memory loss during trace fear conditioning.
(a) Mice received six tone–shock pairings (tone: 20 s, 85 dB, 3 kHz; shock: 2 s, 0.4 mA; 20 s apart) in context A. Twenty-four hours later mice were placed into context B and presented with a tone (60 s) to assess recall (recall 1) after which they were briefly removed, the grid floor covering was taken out, and they were replaced into context B and subjected again to six tone–shock pairings (tone+shock) or six tones only (tone only). Twenty-four hours later mice were placed in context C and presented with a tone (60 s) to assess recall (recall 2). (b,c) Pre-treatment (0.3 mg kg−1, subcutaneous) with 8-OH-DPAT on the second day significantly reduced freezing in Htr1aDG (Tg), but not Htr1aKO (KO) control mice during recall 2 compared with recall 1 in the (b) tone–shock, but not (c) tone only conditions (t-test; n=9–10). (d,e) Pre-treatment (20 nmoles per mouse, i.c.v.) of wild-type mice on the second day of trace fear conditioning (a) with [Pro30,Nle31,Bpa32,Leu34]NPY(28–36), but not vehicle significantly reduced freezing during recall 2 compared with recall 1 in the (d) tone–shock, but not (e) tone only conditions (mean±s.e.m.; n=8–14; *P<0.05; t-test).
Figure 6
Figure 6. Model for function of PP-CA1 inputs to the hippocampus.
Area CA1 of the hippocampus receives information directly from the entorhinal cortex (direct PP-CA1 pathway) and also indirectly via the tri-synaptic circuit. (a) Presentation of paired CS–US promotes potentiation of SC synapses (+) via the indirect pathway depotentiation of SC synapses (–) via the PP-CA1 pathway. In an animal having successfully undergone learning, potentiation and depotentiation are balanced, SC synaptic strength is stable and memories can be retrieved. (b) Inhibition of DG during CS–US presentation suppresses potentiation via the indirect pathway, unmasking depotentiation of SC synapses and promoting memory loss.

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