Cue-induced Reinstatement Of Cocaine Seeking Research Articles

A multivariate regressor of patterned dopamine release predicts relapse to cocaine

Understanding mesolimbic dopamine adaptations underlying vulnerability to drug relapse is essential to inform prognostic tools for effective treatment strategies. However, technical limitations have hindered the direct measurement of sub-second dopamine release invivo for prolonged periods of time, making it difficult to gauge the weight that these dopamine abnormalities have in determining future relapse incidence. Here, we use the fluorescent sensor GrabDA to record, with millisecond resolution, every single cocaine-evoked dopamine transient in the nucleus accumbens (NAc) of freely moving mice during self-administration. We reveal low-dimensional features of patterned dopamine release that are strong predictors of cue-induced reinstatement of cocaine seeking. Additionally, we report sex-specific differences in cocaine-related dopamine responses related to a greater resistance to extinction in males compared with females. These findings provide important insights into the sufficiency of NAc dopamine signaling dynamics-in interaction with sex-for recapitulating persistent cocaine seeking and future relapse vulnerability.

Therapeutic potential of PIMSR, a novel CB1 receptor neutral antagonist, for cocaine use disorder: evidence from preclinical research

Cannabinoid CB1 receptors (CB1Rs) have been major targets in medication development for the treatment of substance use disorders. However, clinical trials with rimonabant, a CB1R antagonist/inverse agonist, failed due to severe side effects. Here, we evaluated the therapeutic potential of PIMSR, a neutral CB1R antagonist lacking an inverse agonist profile, against cocaine’s behavioral effects in experimental animals. We found that systemic administration of PIMSR dose-dependently inhibited cocaine self-administration under fixed-ratio (FR5), but not FR1, reinforcement, shifted the cocaine self-administration dose-response curve downward, decreased incentive motivation to seek cocaine under progressive-ratio reinforcement, and reduced cue-induced reinstatement of cocaine seeking. PIMSR also inhibited oral sucrose self-administration. Importantly, PIMSR alone is neither rewarding nor aversive as assessed by place conditioning. We then used intracranial self-stimulation (ICSS) to explore the possible involvement of the mesolimbic dopamine system in PIMSR’s action. We found that PIMSR dose-dependently attenuated cocaine-enhanced ICSS maintained by electrical stimulation of the medial forebrain bundle in rats. PIMSR itself failed to alter electrical ICSS, but dose-dependently inhibited ICSS maintained by optical stimulation of midbrain dopamine neurons in transgenic DAT-Cre mice, suggesting the involvement of dopamine-dependent mechanisms. Lastly, we examined the CB1R mechanisms underlying PIMSR’s action. We found that PIMSR pretreatment attenuated Δ9-tetrahydrocannabinol (Δ9-THC)- or ACEA (a selective CB1R agonist)-induced reduction in optical ICSS. Together, our findings suggest that the neutral CB1R antagonist PIMSR deserves further research as a promising pharmacotherapeutic for cocaine use disorder.

PPL-103: A mixed opioid partial agonist with desirable anti-cocaine properties

Cocaine use disorder (CUD) is a persistent public health problem for which no effective medications are available. PPL-103 is an opioid receptor ligand with partial agonist activity at mu, kappa and delta opioid receptors, with a greater efficacy for kappa and low efficacy at mu receptors. Because chronic cocaine use induces changes in the kappa opioid receptor/dynorphin system, we hypothesized that a kappa partial agonist, such as PPL-103, would attenuate the aversive properties of the upregulated kappa system, resulting in effective treatment approach for CUD. We tested the effects of PPL-103 on cocaine self-administration models that recapitulate core aspects of CUD in humans. We found that PPL-103 reduced both long and short access cocaine self-administration, motivation to respond for cocaine, and binge-like cocaine taking, in rats. Operant responding for food, fentanyl and locomotor behavior were not altered at doses that decreased cocaine infusions. Repeated PPL-103 treatment did not lead to tolerance development. PPL-103 also reduced both priming- and cue-induced reinstatement of cocaine seeking, being more effective in the former. Surprisingly, PPL-103 reduced self-administration parameters and reinstatement in rats previously treated with the long-acting kappa receptor antagonist JDTic more potently than in non-JDTic treated animals, whereas naltrexone injected to rats subsequent to JDTic administration increased self-administration, suggesting that the partial mu agonist activity, rather than kappa agonism is important for reduction in cocaine taking and seeking. However, partial kappa activation seems to increase safety by limiting dysphoria, tolerance and addiction development. PPL-103 displays a desirable profile as a possible CUD pharmacotherapy.

Increased density and antagonistic allosteric interactions in A2AR-D2R heterocomplexes in extinction from cocaine use, lost in cue induced reinstatement of cocaine seeking

Neurochemical studies were previously performed on the effects of a 10 day extinction learning from cocaine self-administration on D2R and A2AR recognition and D2R Gi/o coupling in the ventral striatum. In the present study biochemical receptor binding and proximity ligation assay were used to study possible changes in the allosteric receptor-receptor interactions and the density of the A2AR-D2R heterocomplexes in the ventral striatum (nucleus accumbens shell) in extinction from cocaine self-administration including cue induced reinstatement of cocaine seeking. A significant and clear-cut reduction of active lever pressing was observed in extinction on day 10 from cocaine use. In cue induced reinstatement of cocaine self-administration a significant return in active lever presses developed. In extinction, significant increases in the density of A2AR-D2R and D2R-Sigma1R heterocomplexes were observed in nucleus accumbens shell. In contrast, cue-induced reinstatement of cocaine seeking produced no significant changes in these heteroreceptor complexes of the nucleus accumbens shell. In the 3H raclopride/quinpirole competition binding experiments, the extinction led to a significant increase in the D2R Ki, High dissociation constant in the ventral striatum upon ex vivo exposure to CGS 21680 (100 nM), compared to the same exposure performed in membrane preparations from yoked saline rats. No significant changes in D2R Ki, High values were observed in membrane preparations from rats after cue-induced reinstatement of cocaine-seeking undergoing the same exposure ex vivo to CGS 21680 when compared with membrane preparations from yoked saline rats undergoing the same procedures. It seems likely that increased formation of A2AR-D2R and putative A2AR-D2R-Sigma1R heterocomplexes in the nucleus accumbens shell is part of the mechanism for the enhanced antagonistic allosteric A2AR-D2R interactions developed in extinction learning from cocaine. It reduces cocaine reward through reduced D2R function, and these inhibitory mechanisms are no longer in operation in cue induced reinstatement of cocaine seeking.

Activation of mesocorticolimbic dopamine projections initiates cue-induced reinstatement of reward seeking in mice.

Drug addiction is characterized by relapse when addicts are re-exposed to drug-associated environmental cues, but the neural mechanisms underlying cue-induced relapse are unclear. In the present study we investigated the role of a specific dopaminergic (DA) pathway from ventral tegmental area (VTA) to nucleus accumbens core (NAcore) in mouse cue-induced relapse. Optical intracranial self-stimulation (oICSS) was established in DAT-Cre transgenic mice. We showed that optogenetic excitation of DA neurons in the VTA or their projection terminals in NAcore, NAshell or infralimbic prefrontal cortex (PFC-IL) was rewarding. Furthermore, activation of the VTA-NAcore pathway alone was sufficient and necessary to induce reinstatement of oICSS. In cocaine self-administration model, cocaine-associated cues activated VTA DA neurons as assessed by intracellular GCaMP signals. Cue-induced reinstatement of cocaine-seeking was triggered by optogenetic stimulation of the VTA-NAcore pathway, and inhibited by chemogenetic inhibition of this pathway. Together, these results demonstrate that cue-induced reinstatement of reward seeking is in part mediated by activation of the VTA-NAcore DA pathway.

CB1 receptor antagonist AM4113 reverts the effects of cannabidiol on cue and stress-induced reinstatement of cocaine-seeking behaviour in mice

No pharmacological treatments are yet approved for patients with cocaine use disorders. Cannabidiol, a constituent of the C. sativa plant has shown promising results in rodent models of drug addiction. However, the specific effects and mechanisms of action of cannabidiol in rodent operant models of extinction-based abstinence and drug-seeking relapse remain unclear. Cannabidiol (10 and 20 mg/kg, i.p.) was injected during extinction training to male CD-1 mice previously trained to self-administer cocaine (0.75 mg/kg/infusion). Then, we evaluated the reinstatement of cocaine seeking induced by cues and stressful stimuli (footshock). We found that cannabidiol (10 and 20 mg/kg) did not modulate extinction learning. After cannabidiol 20 mg/kg treatment, increased levels of CB1 receptor protein were found in the prelimbic and orbitofrontal regions of the prefrontal cortex, and in the ventral striatum; an effect paralleled by a reduction of striatal ∆FosB accumulation and an increment of GluR2 AMPA receptor subunits. Furthermore, cue-induced reinstatement of cocaine seeking was attenuated by cannabidiol. Unexpectedly, cannabidiol 20 mg/kg facilitated stress-induced restoration of cocaine-seeking behaviour. To ascertain the participation of CB1 receptors in these behavioural changes, we administered the CB1 antagonist AM4113 (5 mg/kg) before each reinstatement session. Both, the attenuation of cue-induced reinstatement and the facilitation of stress-induced reestablishment were abolished by AM4113 in cannabidiol 20 mg/kg-treated mice. Our results reveal a series of complex CB1-related changes induced by cannabidiol with a varying impact on the reinstatement of cocaine-seeking behaviour that could limit its therapeutic applications.

Reduced cue-induced reinstatement of cocaine-seeking behavior in Plcb1\u2009+/\u2212\u2009mice

Cocaine addiction causes serious health problems, and no effective treatment is available yet. We previously identified a genetic risk variant for cocaine addiction in the PLCB1 gene and found this gene upregulated in postmortem brains of cocaine abusers and in human dopaminergic neuron-like cells after an acute cocaine exposure. Here, we functionally tested the contribution of the PLCB1 gene to cocaine addictive properties using Plcb1+/− mice. First, we performed a general phenotypic characterization and found that Plcb1+/− mice showed normal behavior, although they had increased anxiety and impaired short-term memory. Subsequently, mice were trained for operant conditioning, self-administered cocaine for 10 days, and were tested for cocaine motivation. After extinction, we found a reduction in the cue-induced reinstatement of cocaine-seeking behavior in Plcb1+/− mice. After reinstatement, we identified transcriptomic alterations in the medial prefrontal cortex of Plcb1+/− mice, mostly related to pathways relevant to addiction like the dopaminergic synapse and long-term potentiation. To conclude, we found that heterozygous deletion of the Plcb1 gene decreases cue-induced reinstatement of cocaine-seeking, pointing at PLCB1 as a possible therapeutic target for preventing relapse and treating cocaine addiction.

Ibudilast attenuates cocaine self-administration and prime- and cue-induced reinstatement of cocaine seeking in rats

Ibudilast is a non-selective phosphodiesterase (PDE) inhibitor and glial cell modulator which has shown great promise for the treatment of drug and alcohol use disorders in recent clinical studies. However, it is unknown whether and how ibudilast affects cocaine seeking behavior. Here we show that systemic administration of ibudilast dose-dependently reduced cocaine self-administration under fixed- and progressive-ratio reinforcement schedules in rats and shifted cocaine dose-response curves downward. In addition, ibudilast decreased cocaine prime- and cue-induced reinstatement of cocaine seeking. These results indicate that ibudilast was effective in reducing the reinforcing effects of cocaine and relapse to cocaine seeking. Chronic cocaine exposure induces cAMP-related neuroadaptations in the reward circuitry of the brain. To investigate potential mechanisms for ibudilast-induced attenuation of cocaine self-administration, we recorded from ventral tegmental area (VTA) dopamine neurons in ex vivo midbrain slices prepared from rats that had undergone saline and cocaine self-administration. We found cocaine self-administration led to a decrease in inhibitory postsynaptic currents (IPSCs), an increase in the AMPAR/NMDAR ratio, and an increase in the excitation to inhibition (E/I) ratio. Ibudilast pretreatments enhanced GABAergic inhibition and did not further change cocaine-induced potentiation of excitation, leading to normalization of the E/I ratio. Restoration of the balance between excitation and inhibition in VTA dopamine neurons may contribute to the attenuation of cocaine self-administration by ibudilast.

Sex-specific role for prefrontal cortical protein interacting with C kinase 1 in cue-induced cocaine seeking.

Disruption of prefrontal glutamate receptor interacting protein (GRIP), which anchors GluA2-containing AMPA receptors (AMPARs) into the synaptic membrane, potentiates cue-induced cocaine seeking in both males and females. Protein interacting with C kinase 1 (PICK1) plays an opposing role to that of GRIP, removing AMPARs from the synapse. Consistent with our hypothesis that disruption of PICK1 in the mPFC would lead to a decrease in addiction-like behaviour, we found that conditional deletion of PICK1 in the mPFC attenuates cue-induced cocaine seeking in male mice. However, prefrontal PICK1 deletion had the opposite effect in females, leading to an increase in cue-induced reinstatement of cocaine seeking. We did not see any effects of PICK1 knockdown on sucrose taking or seeking, suggesting the sex-specific effects do not generalise to natural reinforcers. These findings suggest the role of PICK1 in the prefrontal cortex of females may not be consistent with its accepted role in males. To determine whether these sex differences were influenced by gonadal hormones, we gonadectomised a cohort of males and found that removal of circulating androgens eliminated the effect of prefrontal PICK1 knockdown. As there was no effect of gonadectomy on its own on any of the behavioural measures collected, our results suggest that androgens may be involved in compensatory downstream effects of PICK1 knockdown. Taken together, these results highlight the need for consideration of sex as a biological variable when examining mechanisms underlying all behaviours, as convergent sex differences can reveal different mechanisms where behavioural sex differences do not exist.

Cell Adhesion Factors in the Orbitofrontal Cortex Control Cue-Induced Reinstatement of Cocaine Seeking and Amygdala-Dependent Goal Seeking.

Repeated cocaine exposure causes dendritic spine loss in the orbitofrontal cortex, which might contribute to poor orbitofrontal cortical function following drug exposure. One challenge, however, has been verifying links between neuronal structural plasticity and behavior, if any. Here we report that cocaine self-administration triggers the loss of dendritic spines on excitatory neurons in the orbitofrontal cortex of male and female mice (as has been reported in rats). To understand functional consequences, we locally ablated neuronal β1-integrins, cell adhesion receptors that adhere cells to the extracellular matrix and thus support dendritic spine stability. Degradation of β1-integrin tone: (1) caused dendritic spine loss, (2) exaggerated cocaine-seeking responses in a cue-induced reinstatement test, and (3) impaired the ability of mice to integrate new learning into familiar routines, a key function of the orbitofrontal cortex. Stimulating Abl-related gene kinase, overexpressing Proline-rich tyrosine kinase, and inhibiting Rho-associated coiled-coil containing kinase corrected response strategies, uncovering a β1-integrin-mediated signaling axis that controls orbitofrontal cortical function. Finally, use of a combinatorial gene silencing/chemogenetic strategy revealed that β1-integrins support the ability of mice to integrate new information into established behaviors by sustaining orbitofrontal cortical connections with the basolateral amygdala.SIGNIFICANCE STATEMENT Cocaine degenerates dendritic spines in the orbitofrontal cortex, a region of the brain involved in interlacing new information into established behaviors. One challenge has been verifying links between cellular structural stability and behavior, if any. In this second of two related investigations, we study integrin family receptors, which adhere cells to the extracellular matrix and thereby stabilize dendritic spines (see also DePoy et al., 2019). We reveal that β1-integrins in the orbitofrontal cortex control food- and cocaine-seeking behaviors. For instance, β1-integrin loss amplifies cocaine-seeking behavior and impairs the ability of mice to integrate new learning into familiar routines. We identify likely intracellular signaling partners by which β1-integrins support orbitofrontal cortical function and connectivity with the basolateral amygdala.

Restoring glutamate homeostasis in the nucleus accumbens via endocannabinoid-mimetic drug prevents relapse to cocaine seeking behavior in rats

Impaired glutamate homeostasis is a key characteristic of the neurobiology of drug addiction in rodent models and contributes to the vulnerability to relapse to drug seeking. Although disrupted astrocytic and presynaptic regulation of glutamate release has been considered to constitute with impaired glutamate homeostasis in rodent model of drug relapse, the involvement of endocannabinoids (eCBs) in this neurobiological process has remained largely unknown. Here, using cocaine self-administration in rats, we investigated the role of endocannabinoids in impaired glutamate homeostasis in the core of nucleus accumbens (NAcore), which was indicated by augmentation of spontaneous synaptic glutamate release, downregulation of metabotropic glutamate receptor 2/3 (mGluR2/3), and mGluR5-mediated astrocytic glutamate release. We found that the endocannabinoid, anandamide (AEA), rather than 2-arachidonoylglycerol elicited glutamate release through presynaptic transient receptor potential vanilloid 1 (TRPV1) and astrocytic cannabinoid type-1 receptors (CB1Rs) in the NAcore of saline-yoked rats. In rats with a history of cocaine self-administration and extinction training, AEA failed to alter synaptic glutamate release in the NAcore, whereas CB1R-mediated astrocytic glutamate release by AEA remained functional. In order to induce increased astrocytic glutamate release via exogenous AEA, (R)-methanandamide (methAEA, a metabolically stable form of AEA) was chronically infused in the NAcore via osmotic pumps during extinction training. Restoration of mGluR2/3 function and mGluR5-mediated astrocytic glutamate release was observed after chronic methAEA infusion. Additionally, priming or cue-induced reinstatement of cocaine seeking was inhibited in methAEA-infused rats. These results demonstrate that enhancing endocannabinoid signaling is a potential pathway to restore glutamate homeostasis and may represent a promising therapeutic strategy for preventing cocaine relapse.

The PI3-Kinase p110β Isoform Controls Severity of Cocaine-Induced Sequelae and Alters the Striatal Transcriptome

BackgroundThe PI3-kinase (PI3K) complex is a well-validated target for mitigating cocaine-elicited sequelae, but pan-PI3K inhibitors are not viable long-term treatment options. The PI3K complex is composed of p110 catalytic and regulatory subunits, which can be individually manipulated for therapeutic purposes. However, this possibility has largely not been explored in behavioral contexts. MethodsHere, we inhibited PI3K p110β in the medial prefrontal cortex (mPFC) of cocaine-exposed mice. Behavioral models for studying relapse, sensitization, and decision-making biases were paired with protein quantification, RNA sequencing, and cell type–specific chemogenetic manipulation and RNA quantification to determine whether and how inhibiting PI3K p110β confers resilience to cocaine. ResultsViral-mediated PI3K p110β silencing reduced cue-induced reinstatement of cocaine seeking by half, blocked locomotor sensitization, and restored mPFC synaptic marker content after exposure to cocaine. Cocaine blocked the ability of mice to select actions based on their consequences, and p110β inhibition restored this ability. Silencing dopamine D2 receptor–expressing excitatory mPFC neurons mimicked cocaine, impairing goal-seeking behavior, and again, p110β inhibition restored goal-oriented action. We verified the presence of p110β in mPFC neurons projecting to the dorsal striatum and orbitofrontal cortex and found that inhibiting p110β in the mPFC altered the expression of functionally defined gene clusters within the dorsal striatum and not orbitofrontal cortex. ConclusionsSubunit-selective PI3K silencing potently mitigates drug seeking, sensitization, and decision-making biases after exposure to cocaine. We suggest that inhibiting PI3K p110β provides neuroprotection against cocaine by triggering coordinated corticostriatal adaptations.

Prelimbic cortical projections to rostromedial tegmental nucleus play a suppressive role in cue-induced reinstatement of cocaine seeking

The prelimbic (PL) region of prefrontal cortex has been implicated in both driving and suppressing cocaine seeking in animal models of addiction. We hypothesized that these opposing roles for PL may be supported by distinct efferent projections. While PL projections to nucleus accumbens core have been shown to be involved in driving reinstatement of cocaine seeking, PL projections to the rostromedial tegmental nucleus (RMTg) may instead suppress reinstatement of cocaine seeking, due to the role of RMTg in behavioral inhibition. Here, we used a functional disconnection approach to temporarily disrupt the PL-RMTg pathway during cue- or cocaine-induced reinstatement. Male Sprague Dawley rats self-administered cocaine during daily 2-h sessions for ≥10 days and then underwent extinction training. Reinstatement of extinguished cocaine seeking was elicited by cocaine-associated cues or cocaine prime. Prior to reinstatement, rats received microinjections of the GABA agonists baclofen/muscimol (1/0.1 mM) into unilateral PL and the AMPA receptor antagonist NBQX (1 mM) into contralateral or ipsilateral RMTg. Functional disconnection of PL-RMTg via contralateral inactivation markedly increased cue-induced reinstatement, but did not increase cocaine-induced reinstatement or drive reinstatement of extinguished cocaine seeking in the absence of cues or cocaine. Enhanced cue-induced reinstatement was also observed with ipsilateral inactivation of PL and RMTg, but not with unilateral inactivation of PL or RMTg alone, indicating that both ipsilateral and contralateral projections from PL to RMTg have an inhibitory influence on behavior. These data further support a suppressive role for PL in cocaine seeking by implicating PL efferent projections to RMTg in inhibiting cue-induced reinstatement.

Cocaine Triggers Astrocyte-Mediated Synaptogenesis

BackgroundSynaptogenesis is essential in forming new neurocircuits during development, and this is mediated in part by astrocyte-released thrombospondins (TSPs) and activation of their neuronal receptor, α2δ-1. Here, we show that this developmental synaptogenic mechanism is utilized during cocaine experience to induce spinogenesis and the generation of AMPA receptor–silent glutamatergic synapses in the adult nucleus accumbens shell (NAcSh). MethodsUsing multidisciplinary approaches including astrocyte Ca2+ imaging, genetic mouse lines, viral-mediated gene transfer, and operant behavioral procedures, we monitor the response of NAcSh astrocytes to cocaine administration and examine the role of astrocytic TSP–α2δ-1 signaling in cocaine-induced silent synapse generation as well as the behavioral impact of astrocyte-mediated synaptogenesis and silent synapse generation. ResultsCocaine administration acutely increases Ca2+ events in NAcSh astrocytes, while decreasing astrocytic Ca2+ blocks cocaine-induced generation of silent synapses. Furthermore, knockout of TSP2, or pharmacological inhibition or viral-mediated knockdown of α2δ-1, prevents cocaine-induced generation of silent synapses. Moreover, disrupting TSP2–α2δ-1–mediated spinogenesis and synapse generation in NAcSh decreases cue-induced cocaine seeking after withdrawal from cocaine self-administration and cue-induced reinstatement of cocaine seeking after drug extinction. ConclusionsThese results establish that silent synapses are generated by an astrocyte-mediated synaptogenic mechanism in response to cocaine experience and embed critical cue-associated memory traces that promote cocaine relapse.

Amperometric measurements of cocaine cue and novel context-evoked glutamate and nitric oxide release in the nucleus accumbens core.

Cue-induced reinstatement of cocaine seeking after self-administration (SA) and extinction relies on glutamate release in the nucleus accumbens core (NAcore), which activates neuronal nitric oxide synthase interneurons. Nitric oxide (NO) is required for structural plasticity in NAcore medium spiny neurons, as well as cued cocaine seeking. However, NO release in the NAcore during reinstatement has yet to be directly measured. Furthermore, the temporal relationship between glutamate release and the induction of an NO response also remains unknown. Using wireless amperometric recordings in awake behaving rats, we quantified the magnitude and temporal dynamics of novel context- and cue-induced reinstatement-evoked glutamate and NO release in the NAcore. We found that re-exposure to cocaine-conditioned stimuli following SA and extinction increased extracellular glutamate, leading to release of NO in the NAcore. In contrast, exposing drug-naïve rats to a novel context led to a lower magnitude rise in glutamate in the NAcore relative to cue-induced reinstatement. Interestingly, novel context exposure evoked a higher magnitude NO response relative to cue-induced reinstatement. Despite differences in magnitude, novel context evoked-NO release in the NAcore was also temporally delayed when compared to glutamate. These results demonstrate a dissociation between the magnitude of cocaine cue- and novel context-evoked glutamate and NO release in the NAcore, yet similarity in the temporal dynamics of their release. Together, these data contribute to a greater understanding of the relationship between glutamate and NO, two neurotransmitters implicated in encoding the valence of distinct contextual stimuli.

Cocaine Triggers Glial-Mediated Synaptogenesis

Synaptogenesis is essential in forming new neurocircuits during development, and this is mediated in part by astrocyte-released thrombospondins (TSPs) and activation of their neuronal receptor, α2δ-1. Here, we show that this developmental synaptogenic mechanism is utilized during cocaine experience to induce spinogenesis and the generation of AMPA receptor-silent glutamatergic synapses in the adult nucleus accumbens (NAc). Specifically, cocaine administration activates NAc astrocytes, and preventing this activation blocks cocaine-induced generation of silent synapses. Furthermore, knockout of TSP2, or pharmacological inhibition or viral-mediated knockdown of α2δ-1, prevents cocaine-induced generation of silent synapses. Moreover, disrupting TSP2-α2δ-1-mediated spinogenesis and silent synapse generation in the NAc occludes cue-induced cocaine seeking after withdrawal from cocaine self-administration and cue-induced reinstatement of cocaine seeking after drug extinction. These results establish that silent synapses are generated by an astrocyte-mediated synaptogenic mechanism in response to cocaine experience and embed critical cue-associated memory traces that promote cocaine relapse.

Dynamic CRMP2 Regulation of CaV2.2 in the Prefrontal Cortex Contributes to the Reinstatement of Cocaine Seeking.

Cocaine addiction remains a major health concern with limited effective treatment options. A better understanding of mechanisms underlying relapse may help inform the development of new pharmacotherapies. Emerging evidence suggests that collapsin response mediator protein 2 (CRMP2) regulates presynaptic excitatory neurotransmission and contributes to pathological changes during diseases, such as neuropathic pain and substance use disorders. We examined the role of CRMP2 and its interactions with a known binding partner, CaV2.2, in cocaine-seeking behavior. We employed the rodent self-administration model of relapse to drug seeking and focused on the prefrontal cortex (PFC) for its well-established role in reinstatement behaviors. Our results indicated that repeated cocaine self-administration resulted in a dynamic and persistent alteration in the PFC expression of CRMP2 and its binding partner, the CaV2.2 (N-type) voltage-gated calcium channel. Following cocaine self-administration and extinction training, the expression of both CRMP2 and CaV2.2 was reduced relative to yoked saline controls. By contrast, cued reinstatement potentiated CRMP2 expression and increased CaV2.2 expression above extinction levels. Lastly, we utilized the recently developed peptide myr-TAT-CBD3 to disrupt the interaction between CRMP2 and CaV2.2 in vivo. We assessed the reinstatement behavior after infusing this peptide directly into the medial PFC and found that it decreased cue-induced reinstatement of cocaine seeking. Taken together, these data suggest that neuroadaptations in the CRMP2/CaV2.2 signaling cascade in the PFC can facilitate drug-seeking behavior. Targeting such interactions has implications for the treatment of cocaine relapse behavior.

Glutamate receptor interacting protein acts within the prefrontal cortex to blunt cocaine seeking

Glutamate receptor interacting protein (GRIP) is a neuronal scaffolding protein that anchors GluA2-containing AMPA receptors to the cell membrane. GRIP plays a critical role in activity-dependent synaptic plasticity, including that which occurs after drug exposure. Given that cocaine administration alters glutamate receptor trafficking within the prefrontal cortex (PFC), a better understanding of the role of receptor trafficking proteins could lead to a more complete understanding of addictive phenotypes. AMPA receptor trafficking in general, and GRIP specifically, is known to play a role in cocaine seeking and conditioned reward in the nucleus accumbens, but its role in the PFC has not been characterized. The current study demonstrates that conditional deletion of GRIP1 in the medial prefrontal cortex increases the motivation for cocaine and potentiates cue-induced reinstatement of cocaine seeking in male and female mice. As no effects of PFC GRIP1 deletion were seen in reinstatement of food seeking, strategy set-shifting, or reversal learning the effects on cocaine seeking are not related to generalized alterations in cognitive function. While disrupting GRIP1 might be expected to lead to decreased AMPA transmission, our electrophysiological data indicate an increase in sEPSC amplitude in the prefrontal cortex and a corresponding decrease in paired pulse facilitation in the nucleus accumbens. Taken together this suggests a strengthening of the PFC to NAc input following prefrontal GRIP1 deletion that may mediate the enhanced drug seeking behavior.

Single prolonged stress decreases sign-tracking and cue-induced reinstatement of cocaine-seeking

Exposure to prolonged, uncontrollable stress reduces reward-seeking behavior, resulting in anhedonia in neuropsychiatric disorders, such as posttraumatic stress disorder. However, it is unclear to what degree stressed subjects lose interest in rewards themselves or in reward-related cues that instigate reward-seeking behavior. In the present study, we investigated the effects of single prolonged stress (SPS) on cue-directed behavior in two different procedures: Pavlovian conditioned approach (PCA) and cue-induced reinstatement of cocaine-seeking. In Experiment 1, rats were exposed to SPS and tested for the acquisition of sign-tracking (cue-directed) and goal-tracking (reward-directed) behaviors during a PCA procedure. In Experiment 2, rats were exposed to SPS and tested for the expression of sign- and goal-tracking as well as cue-induced reinstatement of cocaine-seeking. Because dopaminergic activity in the nucleus accumbens is known to play a central role in many cue-directed behaviors, including both sign-tracking and cue-induced reinstatement, Experiment 3 used in vivo microdialysis to measure the effect of SPS on baseline and evoked dopamine levels in the nucleus accumbens. SPS decreased sign-tracking and increased goal-tracking during the acquisition of PCA behavior without affecting reward consumption. In addition, SPS decreased cue-induced reinstatement without affecting cocaine self-administration. Finally, SPS decreased evoked but not baseline levels of dopamine in the nucleus accumbens. These results suggest that SPS decreases the motivational, but not consummatory, aspects of reward-seeking behavior, which may result from long-term, SPS-induced reductions in dopamine release in the nucleus accumbens.

PKMζ in the nucleus accumbens acts to dampen cocaine seeking.

The constitutively active, atypical protein kinase C, protein kinase M-ζ (PKMζ), is exclusively expressed in the brain and its expression increases following exposure to drugs of abuse. However, the limitations of currently available tools have made it difficult to examine the role of PKMζ in cocaine self-administration and relapse. The current study demonstrates that constitutive deletion of PKMζ potentiates cue-induced reinstatement of cocaine seeking and increases both food and cocaine self-administration, without affecting cue-driven food seeking in both male and female mice. Conditional deletion of PKMζ within the nucleus accumbens recapitulated the increase in cocaine taking and seeking seen in the constitutive knockout mice, but only in male animals. Site-specific knockdown of PKMζ in the nucleus accumbens had no effect on cocaine taking or seeking in female mice. Additionally, neither male nor female mice exhibited any alterations in food self-administration or cue-induced reinstatement of food seeking following accumbal deletion of PKMζ. Taken together these results indicate that PKMζ may act to dampen cocaine taking and seeking. Furthermore, these results indicate that PKMζ is playing divergent roles in reward seeking in males and females.