Induction and Blockade of Adolescent Cocaine-Induced Habits

Abstract

Background: Cocaine use during adolescence increases vulnerability to drug dependence and decreases the likelihood that individuals will seek treatment as adults. Understanding how early-life cocaine exposure influences decision-making processes in adulthood is thus critically important.

Methods: Adolescent or adult mice were exposed to subchronic cocaine, then behavioral sensitivity to changes in the predictive relationship between actions and their consequences was tested. Dendritic spines on the principal pyramidal neurons of the orbitofrontal prefrontal cortex (oPFC) were also imaged and enumerated. To determine whether cytoskeletal regulatory systems in the oPFC influenced decision-making strategies, we then inhibited the activity of Abl family and Rho kinases as well as NR2B-containing N-methyl-D-aspartate receptors. We also attempted to block the reinstatement of cocaine seeking in cocaine self-administering mice.

Results: Adult mice with a history of subchronic cocaine exposure in adolescence engaged habit-based response strategies at the expense of goal-directed decision-making strategies and had fewer dendritic spines in the oPFC. Inhibition of the cytoskeletal regulatory Abl family kinases in the oPFC recapitulated these neurobehavioral deficiencies, whereas Rho kinase inhibition corrected response strategies. Additionally, the NR2B-selective N-methyl-D-aspartate receptor antagonists ifenprodil and CP-101,606 blocked cocaine-induced habits; this was dependent on Abl family signaling in the oPFC. Ifenprodil also mitigated cue-induced reinstatement of cocaine seeking in mice self-administering cocaine.

Conclusions: We suggest that adolescent cocaine exposure confers a bias toward habit-based behavior in adulthood via long-term cellular structural modifications in the oPFC. Treatments aimed at mitigating the durable consequences of early-life cocaine use may benefit from targeting cytoskeletal regulatory systems.

Keywords: Abl2; Arg kinase; Ifenprodil; Incubation; OFC; Orbital; Response-outcome.

Figure 1. Mice exposed to cocaine as adolescents are insensitive to changes in response-outcome contingencies as adults, and oPFC dendritic spines are modified

(a) We exposed mice to cocaine from P24-28 (preadolescence) or P31-35 (early adolescence). We then trained mice to generate 2 nose poke responses for food reinforcers as adults. Response acquisition curves represent both responses/min, and there were no side biases throughout. (b) Mice with a history of early-adolescent cocaine exposure were later unable to differentiate between responses that were likely, vs. unlikely, to be reinforced (“non-degraded” vs. “degraded”). Instead, these mice engaged two familiar responses equally. (c) Amphetamine administered from P31-35 did not induce this behavioral inflexibility. (d) Separate adolescent mice were exposed to cocaine, then tested without a drug washout period. A main effect of the response choice indicated that these mice could differentiate between the responses that were more, vs. less, likely to be reinforced in a goal-directed manner. (e) Subchronic cocaine exposure in adult mice also did not impact instrumental response selection, including (f) when mice were given a drug washout period (as in adolescent mice). (g) Dendritic spines on pyramidal neurons in deep-layer oPFC were imaged in adult mice previously exposed to cocaine from P24-28 or P31-35. Density was reduced following P31-35 exposure. (h) Cocaine shifted dendritic spine head diameter cumulative density curves leftward, indicating that dendritic spine heads were also generally smaller in adult mice with a history of cocaine exposure, regardless of age of exposure. Average diameters are inset. (i) Average dendritic spine length increases in the oPFC during adolescent development, however (j) dendritic spines in mature mice with a history of adolescent (P31-35) cocaine exposure were shorter. Representative dendrites are below. Scale bar=10 μm. Means+SEMs,*p<0.05,**p<0.001. Group sizes are indicated.

Figure 2. Bidirectional regulation of goal-directed action

(a) We trained mice to respond for food reinforcers (fig.S1), then infused into the oPFC STI-571 following instrumental contingency degradation. Infusion sites are represented on images from the Mouse Brain Library (67). (b) STI-571 caused an inability to select between actions that were likely, vs. unlikely, to be reinforced (“non-degraded” vs. “degraded”). (c) Dendritic spine densities were reduced within 150 μm of the infusion sites. Representative dendrites are adjacent. (d) Abl2 activity is enriched by inhibiting NR2B-containing NMDARs, blocking cortactin from being trafficked from the spine. The inhibition of Rho-kinase also amplifies Abl2 influence (see 28,29). These events can be induced pharmacologically by ifenprodil and fasudil, respectively, ultimately stabilizing neural structure. (e) Both ifenprodil and fasudil blocked cocaine-induced behavioral inflexibility, indicated by a preference for the response that was more, vs. less, likely to be reinforced. (f) We focused the rest of our report on ifenprodil, revealing that ifenprodil mitigated cocaine-induced reductions in dendritic spine density in the oPFC (0 represents no change from control). Inset: An oPFC neuron with distal branches highlighted and representative spines. (g-h) Further, STI-571 infusions occluded the effects of ifenprodil in enriching goal-directed action selection following adolescent cocaine exposure. Infusion sites are represented on images from the Mouse Brain Library (67). (i) Separate, drug-naïve mice were trained extensively to nose poke for food reinforcers (fig.S1). Subsequently, control mice developed habit-based response strategies, as expected. Ifenprodil restored goal-directed response selection. (j) When ifenprodil treatment was delayed 4 hours following instrumental contingency degradation, it had no effect. Means+SEMs, *p<0.05,**p<0.001. Group sizes are indicated; numbers within parentheses refer to the number of dendrites sampled.

Figure 3. The NMDAR NR2B-selective antagonist CP-101,606 occludes cocaine-induced behavioral inflexibility

(a) Mice were trained to respond for food reinforcers, with no differences between groups (fig.S1). Mice with a history of cocaine exposure subsequently failed to differentiate between responses that were likely, vs. unlikely, to be reinforced, as expected. CP-101,606 (“CP”) blocked these deficiencies. (b) With additional testing, cocaine-exposed mice were ultimately able to differentiate between responses, indicating that cocaine delays, but does not fully block, the development and expression of goal-directed response strategies. Means+SEMs, *p<0.05 following interactions. Group sizes are indicated.

Figure 4. Ifenprodil reduces the cue-induced reinstatement of cocaine seeking

(a) Experimental timeline. (b) Cocaine-reinforced response rates during the response acquisition phase did not differ between mice that would ultimately be treated with vehicle vs. ifenprodil. (c) Mice required the same amount of training to develop stable response rates, regardless of whether they would or would not be ultimately treated with ifenprodil. (d) Further, ifenprodil treatment immediately following each extinction training session (arrows) had no effects on response extinction. (e) Nonetheless, ifenprodil reduced cocaine-seeking behaviors when mice were primed with cocaine-associated cues in a reinstatement test. Ifenprodil-treated mice responded less on the previously active aperture than vehicle-treated mice, and additionally, their responding on the active aperture did not significantly differ from responding on the inactive apertures (p=0.086 between apertures in ifenprodil mice; p<0.001 for the same comparison in vehicle-treated mice). Ifenprodil was not on-board at this time. Means+SEMs, *p<0.05. Group sizes are indicated.