Membrane excitability of nucleus accumbens neurons gates the incubation of cocaine craving

Abstract

After drug withdrawal, a key factor triggering relapse is progressively intensified cue-associated drug craving, termed incubation of drug craving. After withdrawal from cocaine self-administration, incubation of cocaine craving develops more reliably in rats compared to mice. This species difference provides an opportunity to determine rat-specific cellular adaptations, which may constitute the critical mechanisms that contribute to incubated cocaine craving in humans. Expression of incubated cocaine seeking is mediated, in part, by cocaine-induced cellular adaptations in medium spiny neurons (MSNs) within the nucleus accumbens (NAc). In rats, decreased membrane excitability in NAc MSNs is a prominent cellular adaptation, which is induced after cocaine self-administration and lasts throughout prolonged drug withdrawal. Here, we show that, similar to rats, mice exhibit decreased membrane excitability of dopamine D₁ receptor (D1)-, but not D₂ (D2)-, expressing MSNs within the NAc shell (NAcSh) after 1 d withdrawal from cocaine self-administration. However, in contrast to rats, this membrane adaptation does not persist in mice, diminishing after 45-d withdrawal. We also find that restoring the membrane excitability of NAcSh MSNs after cocaine withdrawal decreases cocaine seeking in rats. This suggests that drug-induced membrane adaptations are essential for behavioral expression of incubated cocaine craving. In mice, however, experimentally inducing hypoactivity of D1 NAcSh MSNs after cocaine withdrawal does not alter cocaine seeking, suggesting that MSN hypo-excitability alone is insufficient to increase cocaine seeking. Together, our results demonstrate an overall permissive role of cocaine-induced hypoactivity of NAcSh MSNs in gating increased cocaine seeking after prolonged cocaine withdrawal.

Fig. 1. Cocaine-induced membrane adaptations in mice.

A Schematic showing experimental procedure of self-administration training and electrophysiological recording on withdrawal d1. B Diagram showing the four striatal subregions where MSNs were sampled. Summaries showing the infusion (C) and operant responding (D) in mice used for the withdrawal d1 recording. Examples and summaries of numbers of evoked action potentials in MSNs from the NAcSh (E), NAcCo (F), DMS (G), and DLS (H) in mice 1 d after self-administration training. Insets, example action potentials evoked by 300 pA current steps. I Schematic showing that electrophysiological recording was performed on d 45 after self-administration training. Summaries showing the infusion (J) and operant responding (K) results in mice used for the withdrawal d 45 recording. Examples and summaries of numbers of evoked action potentials in MSNs from the NAcSh (L), NAcCo (M), DMS (N), and DLS (O) in mice on d 45 after self-administration training. All comparisons were performed with animal-based 2-way mixed ANOVA. *p < 0.05.

Fig. 2. Membrane excitability of D1 and D2 MSNs in mice after 1d withdrawal from cocaine self-administration.

A Schematic showing electrophysiological recording performed on d 1 after self-administration training. Summarized self-administration results showing the infusion (B) and operant responding (C) in mice used for the withdrawal d1 recording. Examples and summaries of numbers of evoked action potentials in D1 or D2 MSNs from the NAcSh (D, E), NAcCo (F, G), DMS (H, I), and DLS (J, K) in mice on d 1 after self-administration training. All comparisons were performed using animal-based 2-way mixed ANOVA. *p < 0.05.

Fig. 3. Membrane excitability of D1 and D2 MSNs in mice after 45d withdrawal from cocaine self-administration.

A Schematic showing electrophysiological recording on d 45 after self-administration training. Summaries showing self-administration results, including saline/cocaine infusions (B) and operant responding (C), in mice used for the withdrawal d45 recording. Examples and summaries of numbers of evoked action potentials in D1 or D2 MSNs from the NAcSh (D, E), NAcCo (F, G), DMS (H, I), and DLS (J, K) in mice on d 45 after self-administration training. All comparisons were performed using animal-based 2-way mixed ANOVA.

Fig. 4. Permissive role of cocaine-induced membrane adaptation in cue-induced cocaine seeking in rats.

A Schematic showing electrophysiological recording in rats on d 45 after self-administration training. B Summarized infusion results of rats with saline or cocaine self-administration. Examples (C) and summary (D) of numbers of action potentials evoked in NAcSh MSNs from saline- and cocaine-trained rats. E Example AHPs in NAcSh MSNs from a saline- and a cocaine-trained rat after 45 d of withdrawal from self-administration training. F Summaries showing increased amplitudes of mAHP in NAcSh MSNs in cocaine-trained rats compared to saline-trained rats. G Schematic showing intra-NAcSh administration of apamin, followed by cue-induced cocaine seeking test on withdrawal d 45 after self-administration training. H Summaries of self-administration results (infusions) of rats that later received intra-NAcSh injection of aCSF vehicle or apamin. I Schematic illustration showing the postmortem-identified locations of the cannula that delivered apamin within the NAcSh. J Summaries showing that intra-NAcSh injection of apamin on withdrawal d 45 after cocaine self-administration decreased cue-induced cocaine seeking. *p < 0.05; **p < 0.01.

Fig. 5. Induction of NAcSh D1 MSN hypoactivity fails to induce incubated cocaine seeking in mice.

A Schematic showing experimental procedures in D1-Cre mice, including intra-NAc expression of hM4D in D1 MSNs, 5-d self-administration training, cue-induced cocaine seeking tests on withdrawal d1 and d45, and C21 administration on withdrawal d45 before operant testing. B Diagram showing intra-NAcSh injection of AAV9-hSyn-DIO-hM4Di-mCherry. C Example image showing viral-mediated expression of hM4Di-mCherry in NAcSh D1 MSNs. D Summaries of cocaine self-administration results in mice used for vehicle injection. E Summaries showing cue-induced cocaine seeking (lever presses) on withdrawal d 1 and d45 in mice that received vehicle injection on withdrawal d 45. F Summaries of cocaine self-administration results in mice used for C21 injection. G Summaries showing cue-induced cocaine seeking on withdrawal d1 and d45 in mice that received C21 injection on withdrawal d45. H Summaries showing that chemogenetic suppression of D1 NAc MSNs in mice on withdrawal d 45 after cocaine self-administration did not affect cue-induced cocaine seeking.