Contingent Amygdala Inputs Trigger Heterosynaptic LTP at Hippocampus-To-Accumbens Synapses

Abstract

The nucleus accumbens shell (NAcSh) is a key brain region where environmental cues acquire incentive salience to reinforce motivated behaviors. Principal medium spiny neurons (MSNs) in the NAcSh receive extensive glutamatergic projections from limbic regions, among which, the ventral hippocampus (vH) transmits information enriched in contextual cues, and the basolateral amygdala (BLA) encodes real-time arousing states. The vH and BLA project convergently to NAcSh MSNs, both activated in a time-locked manner on a cue-conditioned motivational action. In brain slices prepared from male and female mice, we show that co-activation of the two projections induces long-term potentiation (LTP) at vH-to-NAcSh synapses without affecting BLA-to-NAcSh synapses, revealing a heterosynaptic mechanism through which BLA signals persistently increase the temporally contingent vH-to-NAcSh transmission. Furthermore, this LTP is more prominent in dopamine D1 receptor-expressing (D1) MSNs than D2 MSNs and can be prevented by inhibition of either D1 receptors or dopaminergic terminals in NAcSh. This heterosynaptic LTP may provide a dopamine-guided mechanism through which vH-encoded cue inputs that are contingent to BLA activation acquire increased circuit representation to reinforce behavior.SIGNIFICANCE STATEMENT In motivated behaviors, environmental cues associated with arousing stimuli acquire increased incentive salience, processes mediated in part by the nucleus accumbens (NAc). NAc principal neurons receive glutamatergic projections from the ventral hippocampus (vH) and basolateral amygdala (BLA), which transmit information encoding contextual cues and affective states, respectively. Our results show that co-activation of the two projections induces long-term potentiation (LTP) at vH-to-NAc synapses without affecting BLA-to-NAc synapses, revealing a heterosynaptic mechanism through which BLA signals potentiate the temporally contingent vH-to-NAc transmission. Furthermore, this LTP is prevented by inhibition of either D1 receptors or dopaminergic axons. This heterosynaptic LTP may provide a dopamine-guided mechanism through which vH-encoded cue inputs that are contingent to BLA activation acquire increased circuit representation to reinforce behavior.

Keywords: LTP; amygdala; dopamine; heterosynaptic; hippocampus; nucleus accumbens.

Figure 1.

Electrophysiological separation of vH-to-NAcSh and BLA-to-NAcSh projections with the Dual-rhodopsin system. A, Diagram showing injection of a single AAV, either AAV-ChR2-EYFP or AAV-Chrimson-tdTomato, in a single NAcSh-projecting brain region. B, Diagram showing the experimental setup, in which EPSCs were recorded from NAcSh MSNs in response to optogenetic stimulation of single rhodopsin-expressing presynaptic fibers by sequential application of lasers of three different wavelengths. C, Example EPSCs evoked in the same MSN by optogenetic stimulation of ChR2-expressing presynaptic fibers with 445-, 473-, or 635-nm laser. D, Summary of the amplitudes of ChR2-evoked EPSCs by 445-, 473-, and 635-nm lasers at different powers. E, Summary of the same results in D after normalizing the amplitudes of EPSCs to those evoked by the 445-nm laser at 2 mW. F, Example EPSCs evoked by optogenetic stimulation of Chrimson-expressing presynaptic fibers with 445-, 473-, or 635-nm laser. G, Summary of the amplitudes of Chrimson-evoked EPSCs by 445-, 473-, and 635-nm lasers at different powers. H, Summary of the same results in G after normalizing the amplitudes of EPSCs to those evoked by the 635-nm laser at 2 mW. I, Diagram showing recording of the same MSN in response to optogenetic stimulation of vH-to-NAcSh versus BLA-to-NAcSh projections. J, Schematics of the alternating stimulation protocol, in which the ChR2-expressing vH projection and Chrimson-expressing BLA projection were alternatingly stimulated with a two-pulse train: interpulse interval, 50 ms; train interval within each projection, 15 s; train interval between the two projections, 7.5 s. K, EPSCs induced by the alternating stimulation protocol in an example MSN (colored traces depict the average of individual gray traces). L, Summary showing a higher mean PPR at BLA-to-NAcSh synapses compared with vH-to-NAcSh synapses. M, Example EPSCs from vH-to-NAcSh and BLA-to-NAcSh synapses scaled and aligned at their peaks. N, Summary showing similar rise and decay kinetics of EPSCs from vH-to-NAcSh versus BLA-to-NAcSh synapses. O, Schematics of Sync protocol, in which the application of 445- and 635-nm lasers was synchronized over 2 min (15 s apart) after the alternating stimulation procedure. P, Example EPSCs evoked by alternating stimulation of vH-to-NAcSh and BLA-to-NAcSh projections and by Sync stimulation. Q, Summary showing that summed amplitudes of vH-to-NAcSh and BLA-to-NAcSh EPSCs in individual MSNs were similar to the amplitudes of Sync-evoked EPSCs. **p < 0.01

Figure 2.

Sync-induced heterosynaptic LTP at vH-to-NAcSh synapses. A, Experimental schematics for inducing Sync-induced LTP. B, F, Trials of amplitudes of EPSCs from vH-to-NAcSh (B) and BLA-to-NAcSh (F) synapses in an example MSN before and after Sync. C, G, Summaries showing that Sync stimulation of vH-to-NAcSh and BLA-to-NAcSh projections selectively induced LTP at vH-to-NAcSh (C), but not BLA-to-NAcSh, synapses (G). D, H, Summaries showing that Sync did not change the mean PPR at vH-to-NAcSh (D) or BLA-to-NAcSh (H) synapses. E, I, Summaries showing that the normalized PPRs are negatively correlated to the normalized amplitudes of EPSCs from vH-to-NAcSh (E), but not BLA-to-NAcSh (I), synapses after Sync. **p < 0.01

Figure 3.

Sync LTP differed in D1 and D2 MSNs. A, Diagram showing the dual-recording, dual-rhodopsin experimental setup, in which a D1 and a putative D2 MSN were simultaneously recorded in response to optogenetic stimulation of vH-to-NAcSh and BLA-to-NAcSh projections. Inset, Example image showing dual recording of tdTomato-positive (D1) and tdTomato-negative (D2) MSNs. B, Example EPSCs in dual-recorded D1 and D2 MSNs evoked by optogenetic stimulation of vH-to-NAcSh and BLA-to-NAcSh projections. C, Summaries showing similar amplitudes of EPSCs in dual-recorded D1 and D2 MSNs evoked by stimulation of either vH-to-NAcSh or BLA-to-NAcSh projections. D, Example EPSCs in dual-recorded D1 and D2 MSNs evoked before and during Sync (stimulation schematics shown in upper panel). E, G, Summaries showing that Sync stimulation of vH and BLA projections induced LTP at both vH-to-D1 and vH-to-D2 synapses (E) without changing the amplitudes of EPSCs from BLA-to-D1 or D2 synapses (G). F, H, Summaries showing that, after Sync, the relative amplitudes of EPSCs from vH-to-D1 synapses were higher than vH-to-D2 synapses (F), while the amplitudes of EPSCs from BLA-to-D1 and BLA-to-D2 synapses were similar after Sync (H). I–N, Summaries showing in two parallel experiments performed in tandem that EPSCs evoked from vH- (I) and BLA-to-D1 and D2 synapses (L) remained constant over the 25-min recording without Sync, while Sync stimulation induced LTP at both vH-to-D1 and vH-to-D2 synapses (J) with higher magnitude at vH-to-D1 synapses (K), and Sync stimulation did not affect BLA-to-D1 or BLA-to-D2 synaptic transmission (M) or relative strengths between these synapses (N). *p < 0.05; **P < 0.01

Figure 4.

Inhibition of NMDA or GABA_A receptors did not prevent Sync-induced heterosynaptic LTP. A, B, Summaries showing that, in the presence of D-AP5, Sync-induced LTP was intact at both vH-to-D1 and vH-to-D2 synapses (A) and remained absent at BLA-to-D1 and BLA-to-D2 synapses (B). C, D, Summaries showing that, in the presence of picrotoxin, Sync-induced LTP was intact at vH-to-D1 and vH-to-D2 synapses (C) and remained absent at BLA-to-D1 and BLA-to-D2 synapses (D). E, Summary of the LTP magnitude in each experimental condition. The LTP magnitude was represented as the ratio of the amplitude of EPSCs during 11–20 min after Sync over the amplitude of EPSCs during the 5-min recording before Sync. Data statistical results are combined from all related recordings in this and previous figures. *P < 0.05; **p < 0.01.

Figure 5.

DA D1 receptor-coupled signaling was essential for Sync-induced LTP. A, B, Summaries showing that, in the presence of SCH23390, Sync-induced LTP was prevented at vH-to-D1 and vH-to-D2 synapses (A) and remained absent at BLA-to-D1 and BLA-to-D2 synapses (B). C, D, Summaries showing that, in the presence of eticlopride, Sync-induced LTP was intact at vH-to-D1 and vH-to-D2 synapses (C) and remained absent at BLA-to-D1 and BLA-to-D2 synapses (D). E, F, Summaries showing that, with GDP-β-S perfused in MSNs, Sync-induced LTP was intact at vH-to-D1 and vH-to-D2 synapses (E) and remained absent at BLA-to-D1 and BLA-to-D2 synapses (F). G, Summary of the LTP magnitude in each experimental condition. The LTP magnitude was represented as the ratio of the amplitude of EPSCs after the Sync induction over the amplitude of EPSCs before Sync. Data and statistical results are combined from all related recordings in this and previous figures. *p < 0.05; **p < 0.01.

Figure 6.

Chemogenetic inhibition of DA presynaptic terminals in the NAcSh prevented Sync-induced LTP. A, Perfusion of CNO prevented the induction of Sync-induced LTP at vH-to-NAcSh synapses in slices in which presynaptic DA terminals selectively expressed hM4Di, but not in control slices. B, Perfusion of CNO did not affect BLA-to-NAcSh synaptic transmission before or after Sync stimulation in slices with or without hM4Di expression in presynaptic DA terminals. C, D, Diagrams illustrating the hypothesis that during cue-affect conditioning, co-activation of the cue-encoding vH and affect-encoding BLA projections together with a DA-mediated “teaching signal” create a heterosynaptic condition (C) to induced LTP at vH-to-NAcSh synapses. Through this LTP, the conditioned cue encoded by the vH projection gains increased power to excite NAcSh MSNs (D).