Role of nucleus accumbens shell neuronal ensembles in context-induced reinstatement of cocaine-seeking

Abstract

Environmental contexts previously associated with drug use provoke relapse to drug use in humans and reinstatement of drug seeking in animal models of drug relapse. We examined whether context-induced reinstatement of cocaine seeking is mediated by activation of context-selected nucleus accumbens neurons. We trained rats to self-administer cocaine in Context A and extinguished their lever-pressing in a distinct Context B. On test day, reexposure to the cocaine-associated Context A reinstated cocaine seeking and increased expression of the neural activity marker Fos in 3.3% of accumbens shell and 1.6% of accumbens core neurons. To assess a causal role for these activated neurons, we used the Daun02 inactivation procedure to selectively inactivate these neurons. We trained c-fos-lacZ transgenic rats to self-administer cocaine in Context A and extinguished their lever-pressing in Context B. On induction day, we exposed rats to either Context A or a novel Context C for 30 min and injected Daun02 or vehicle into accumbens shell or core 60 min later. On test day, 3 d after induction day, the ability of Context A to reinstate cocaine seeking and increase neuronal activity in accumbens shell was attenuated when Daun02 was previously injected after exposure to Context A. Daun02 injections after exposure to the novel Context C had no effect on context-induced reinstatement of cocaine seeking despite much greater numbers of Fos-expressing neurons induced by Context C. Daun02 injections in accumbens core had no effect. Our data suggest that context-induced reinstatement of cocaine seeking is mediated by activation of context-selected accumbens shell but not core neuronal ensembles.

Keywords: Daun02 inactivation; Fos; conditioned cues; drug environment; extinction; self-administration.

Figure 1.

Cocaine self-administration training and extinction of drug-reinforced responding. A, Rats were trained to self-administer cocaine using 1 mg/kg/infusion for the first 6 d and 0.5 mg/kg/infusion for the next 6 d. Data are mean ± SEM number of infusions, and active and inactive lever-presses during cocaine self-administration training in Context A in Experiment 1 (n = 34), Experiment 2 (n = 42), and Experiment 3 (n = 21). B, Mean ± SEM number of presses on the active lever and inactive lever during the first 12 extinction sessions conducted in the absence of cocaine in a different (Extinction) Context B in Experiments 1–3.

Figure 2.

Context-induced reinstatement of cocaine seeking is associated with Fos induction in accumbens shell. A, Reinstatement test: Total number of active lever and inactive lever-presses in rats tested in the Extinction or Cocaine context. Data are mean ± SEM. *p < 0.05, different from the Extinction context. n = 10 or 11 per group. B, Number of Fos-IR nuclei per mm² in accumbens shell. Data are mean ± SEM. C, Number of Fos-IR nuclei per mm² in accumbens core. D, Top two panels, Fos-IR nuclei in accumbens core and shell captured at 400× magnification. Bottom three panels, 100× magnification of areas used for quantifying Fos-IR neurons in accumbens shell and core of rats that were exposed to Extinction, Cocaine, and Novel contexts. *p < 0.05, different from the Extinction context. ^#p < 0.05, different from the Cocaine context. n = 8–11 per group.

Figure 3.

Fos and NeuN immunohistochemistry in nucleus accumbens of rats after exposure to training, extinction, or novel contexts. A, Data are mean ± SEM of Fos + NeuN double-labeled neurons as a percentage of all NeuN-labeled neurons in accumbens shell. B, Data are mean ± SEM of Fos + NeuN double-labeled neurons as a percentage of all NeuN-labeled neurons in accumbens core. *Different from extinction context; #different from cocaine context, p < 0.05. C, Red-labeled nuclei indicate expression of the general neuronal nuclei marker NeuN. D, Green-labeled nuclei indicate Fos expression. E, Merged image indicating Fos + NeuN double-labeled nuclei in yellow. Scale bar, 50 μm. n = 4 per group.

Figure 4.

Daun02 inactivation of activated accumbens shell neurons decreased context-induced reinstatement of cocaine seeking. A, Timeline for Daun02 inactivation experiment. Cocaine self-administration training in Context A and extinction of cocaine seeking in Context B. On induction day, rats were exposed to the training Context A or a novel Context C, and injected bilaterally with Daun02 or vehicle into the accumbens 90 min after the beginning of exposure. Context-induced reinstatement was assessed in Context A 3 d later. B, Total active lever-presses during the second day of extinction after vehicle or Daun02 injections. C, Total active lever-presses during context-induced reinstatement of cocaine seeking on test day. D, Dots indicate approximate area of injector tip. E, Accumbens shell βgal expression after the context-induced reinstatement test. F, Representative images of βgal-labeled nuclei in accumbens shell. Data are mean ± SEM. *p < 0.05, different from vehicle-ABAA (Cocaine) group. n = 10 or 11 per group.

Figure 5.

Daun02 inactivation of activated accumbens core neurons had no effect on context-induced reinstatement of cocaine seeking. A, Total active lever-presses during the second day of extinction after vehicle or Daun02 injections. B, Total active lever-presses during context-induced reinstatement of cocaine seeking on test day. C, Dots indicate approximate area of injector tip. D, Accumbens core βgal expression after the context-induced reinstatement test. E, Representative images of βgal-labeled nuclei in accumbens core. Data are mean ± SEM. *p < 0.05, different from vehicle. n = 10 or 11 per group.