Effects of Highly Palatable Diet on motivation for food and resistance to punishment in rats: Role of sex and age of exposure.

Exposure to highly palatable food is believed to induce behavioral and neurobiological changes that may produce addiction-like behavior and increase the risks of obesity and overweight. Studies in rodents have led to conflicting results suggesting that several factors such as sex and age of exposure contribute to the development of maladaptive behaviors towards food. In addition, it is not clear whether effects of exposure to highly palatable diets (HPD) persist after their discontinuation, which would indicate long-term risks to develop addiction-like behavior. In this study, we investigated the persistent effects of an intermittent 8-week exposure to HPD in male and female rats as a function of age of exposure (adult and adolescent). We found that intermittent exposure to HPD did not alter body weight, but it affected consumption of standard food during the time of exposure in all groups. In addition, in adults, HPD produced a decrease in the initial baseline responding in FR1 schedules, an effect that persisted for 4 weeks in males but not in female rats. However, we found that exposure to HPD did not affect resistance to punishment measured by progressive shock strength break points or motivation for food as measured by progressive-ratio break points regardless of sex or age of exposure. Altogether, these results do not provide support for the hypothesis that intermittent exposure to HPD produce persistent increases in the vulnerability to develop addiction-like behaviors towards palatable food.

Delay of punishment highlights differential vulnerability to developing addiction-like behavior toward sweet food.

Resistance to punishment is commonly used to measure the difficulty in refraining from rewarding activities when negative consequences ensue, which is a hallmark of addictive behavior. We recently developed a progressive shock strength (PSS) procedure in which individual rats can titrate the amount of punishment that they are willing to tolerate to obtain food rewards. Here, we investigated the effects of a range of delays (0-12 s) on resistance to punishment measured by PSS break points. As expected from delay discounting principles, we found that delayed shock was less effective as a punisher, as revealed by higher PSS breakpoints. However, this discounting effect was not equally distributed in the population of rats, and the introduction of a delay highlighted the existence of two populations: rats that were sensitive to immediate punishment were also sensitive to delayed shock, whereas rats that were resistant to immediate punishment showed strong temporal discounting of delayed punishment. Importantly, shock-sensitive rats suppressed responding even in subsequent non-punishment sessions, and they differed from shock-resistant rats in anxiety-like behavior, but not in sensitivity to pain. These results show that manipulation of temporal contingencies of punishment in the PSS procedure provides a valuable tool to identify individuals with a double vulnerability to addiction: low sensitivity to aversion and excessive discounting of negative future consequences. Conversely, the shock-sensitive population may provide a model of humans who are vulnerable to opportunity loss due to excessive anxiety.

A self-adjusting, progressive shock strength procedure to investigate resistance to punishment: Characterization in male and female rats.

Indifference to harmful consequences is one of the main characteristics of compulsive behaviors and addiction. Animal models that provide a rapid and effective measure of resistance to punishment could be critical for the investigation of mechanisms underlying these maladaptive behaviors. Here, analogous to the progressive ratio (PR) procedure widely used to evaluate appetitive motivation as the response requirement is increased, we developed a self-adjusting, progressive shock strength (PSS) procedure. The PSS provides, within a single session, a break point that quantifies the propensity to work for a reward in spite of receiving electric footshock that progressively increases in duration. In both male and female rats, the PSS break point was sensitive to 1) hunger; and 2) changes in the qualitative, but not quantitative, incentive value of the reward. In systematic comparisons between PSS and PR procedures in the same rats, we found that both measures are sensitive to manipulations of motivational states, but they are not intercorrelated, suggesting that they measure overlapping but partially distinct processes. Importantly, the PSS procedure represents a refinement in the 3Rs principles of animal research because animals can control the strength of shock that they are willing to tolerate. This self-adjusting PSS procedure may represent a useful tool to investigate mechanisms underlying maladaptive behavior that persists in certain individuals despite harmful consequences.

Prevention of relapse to methamphetamine self-administration by environmental enrichment: involvement of glucocorticoid receptors.

RATIONALE: In rodents, environmental enrichment (EE) produces both preventive and curative effects on drug addiction, and this effect is believed to depend at least in part on EE's actions on the stress system. OBJECTIVES: This study investigated whether exposure to EE during abstinence reduces methamphetamine seeking after extended self-administration. In addition, we investigated whether these effects are associated with alterations in the levels of glucocorticoid receptors (GR) in the brain and whether administration of GR antagonists blocks methamphetamine relapse. METHODS: We allowed rats to self-administer methamphetamine for twenty 14-h sessions. After 3 weeks of abstinence either in standard (SE) or EE conditions, we measured methamphetamine seeking in a single 3-h session. Then, we used western blot techniques to measure GR levels in several brain areas. Finally, in an independent group of rats, after methamphetamine self-administration and abstinence in SE, we administered the GR antagonist mifepristone, and we investigated methamphetamine seeking. RESULTS: Exposure to EE reduced methamphetamine seeking and reversed methamphetamine-induced increases in GR levels in the ventral and dorsal hippocampus. In addition, EE decreased GR levels in the amygdala in drug-naive animals, but this effect was prevented by previous exposure to methamphetamine. Administration of mifepristone significantly decreased methamphetamine seeking. CONCLUSIONS: The anti-craving effects of EE are paralleled by restoration of methamphetamine-induced dysregulation of GR in the hippocampus. These results provide support for the hypothesis that the effect of EE on methamphetamine relapse is at least in part mediated by EE's action on the brain stress system.

Environmental enrichment-inspired pharmacological tools for the treatment of addiction.

Environmental enrichment (EE) has been shown to produce powerful beneficial effects in animal models of addiction. In particular, the ability of EE to promote abstinence and prevent relapse may allow for the identification of brain mechanisms responsible for the recovery from addiction. Indeed, the effects of EE on specific brain mechanisms could be mimicked by old or new molecules, which may become novel medications, called enviromimetics. Here, we review the best known enviromimetics for the treatment of addiction and suggest that, whereas these compounds may be relatively ineffective by themselves, they may be useful complements for existing therapeutic approaches to manage addiction which includes behavioural, environmental and pharmacological interventions.

Dopamine and addiction: what have we learned from 40 years of research.

Among the neurotransmitters involved in addiction, dopamine (DA) is clearly the best known. The critical role of DA in addiction is supported by converging evidence that has been accumulated in the last 40 years. In the present review, first we describe the dopaminergic system in terms of connectivity, functioning and involvement in reward processes. Second, we describe the functional, structural, and molecular changes induced by drugs within the DA system in terms of neuronal activity, synaptic plasticity and transcriptional and molecular adaptations. Third, we describe how genetic mouse models have helped characterizing the role of DA in addiction. Fourth, we describe the involvement of the DA system in the vulnerability to addiction and the interesting case of addiction DA replacement therapy in Parkinson's disease. Finally, we describe how the DA system has been targeted to treat patients suffering from addiction and the result obtained in clinical settings and we discuss how these different lines of evidence have been instrumental in shaping our understanding of the physiopathology of drug addiction.

Persistent Neuroadaptations in the Expression of Genes Involved in Cholesterol Homeostasis Induced by Chronic, Voluntary Alcohol Intake in Rats.

Alcohol use disorder (AUD) is associated with persistent adaptations in the brain that are believed to participate in the long-lasting vulnerability to relapse after abstinence. Cholesterol, the major sterol compound found in the central nervous system (CNS), plays a major role in maintenance of neuronal morphology, synaptogenesis and synaptic communication and may be involved in alcohol-induced neuroadaptations. In this study, we investigated whether alcohol consumption in a two-bottle choice paradigm followed by 3 weeks of abstinence could alter the expression of genes encoding proteins involved in cholesterol homeostasis in brain regions involved in addiction and relapse, namely the prefrontal cortex (PFC), the nucleus accumbens (NAc), the mesencephalon and the amygdala. We found that voluntary alcohol intake followed by 3 weeks of forced abstinence produces changes in the transcription of several genes encoding proteins directly involved in cholesterol synthesis such as 3-hydroxyl-3-methylglutaryl-coenzyme A (HMGCoA) reductase, farnesyl-diphosphate farnesyltransferase 1 (FDFT1) and farnesyl diphosphate synthase (FDPS) and in its regulation such as sterol regulatory element-binding factor-2 (SREBF2), in cholesterol transport such as ATP-binding cassette subfamily A member 1 (ABCA1) and in cholesterol degradation such as CYP46A1. Interestingly, these changes appeared to be region-specific and suggest that previous chronic exposure to alcohol might durably increase cholesterol metabolism in the PFC, the NAc and the mesencephalon and decrease cholesterol metabolism in the amygdala. Altogether, these results suggest that alcohol consumption leads to durable deregulations in cholesterol metabolism in key areas involved in loss of control over drug use and addiction. These long-term neuroadaptations may participate in the changes in brain structure and functioning that are responsible for the long-lasting risks of relapse to alcohol.

Generalization of effects of environmental enrichment on seeking for different classes of drugs of abuse.

BACKGROUND: Addiction is a chronic disease characterized by persistent vulnerability to relapse during abstinence. In animal models of addiction, accumulating evidence suggests that exposure to environmental enrichment (EE) during periods of abstinence can have curative effects on addiction and reduce the risks of relapse. However, until present most studies have mainly focused on cocaine. In this study, we investigated whether EE could have beneficial effects on cue-induced seeking for several psychoactive drugs belonging to different pharmacological classes such as methamphetamine (METH), heroin (HER) and nicotine (NIC). METHODS: After self-administration training of METH, HER and NIC, rats were housed in enriched (EE) or standard environments (SE) for 21-28 days of forced abstinence and then drug-seeking behavior was assessed in the absence of the drug. RESULTS: We found that, compared to SE housing, exposure to EE reduced drug seeking behavior for all drugs tested. CONCLUSIONS: These findings suggest that the anti-craving effects of EE are general for a wide variety of drugs and support the hypothesis that environmental stimulation may be a general intervention for attenuating relapse in humans.

Behavioral flexibility predicts increased ability to resist excessive methamphetamine self-administration.

Drug addiction is often associated with cognitive deficits and behavioral inflexibility that may contribute to the development and maintenance of addictive behaviors by reducing addicts' ability to control their behavior toward the drug. In this study, we investigated the relationships between pre-drug levels of behavioral flexibility and the risk to develop uncontrolled methamphetamine (METH) self-administration. First, we measured individual performance in an inter-dimensional set-shifting procedure in which animals have to switch between an external visual rule and an internal side rule in order to obtain food pellets. Then we allowed rats to self-administer METH for twenty long 14-hour sessions, and we investigated the relationships between behavioral flexibility and measures of control over drug intake. Rats rapidly acquired to self-administer high levels of METH which resulted in moderate weight loss. After several sessions of self-administration, whereas some rats progressively increased their METH intake, other rats showed very long voluntary pauses between drug injections and showed no escalation in METH self-administration. Interestingly, we found that behavioral flexibility is correlated with METH self-administration and that more flexible rats take less METH and do not escalate drug taking. These results suggest that traits of behavioral flexibility may protect against the development of excessive and dysregulated drug taking. Conversely, the inability to adapt behavioral responses as a function of the environmental contingencies may contribute to the risks to develop addiction to METH.

Reduction of Cocaine-Induced Locomotor Effects by Enriched Environment Is Associated with Cell-Specific Accumulation of ΔFosB in Striatal and Cortical Subregions.

Background: Early exposure to enriched environments has been shown to decrease the locomotor effects induced by repeated injections of cocaine and modify basal and cocaine-induced total protein levels of the transcription factor ΔFosB in the whole striatum of mice. In this study, we aimed at characterizing whether the profile of ΔFosB accumulation induced by enriched environments and cocaine would be similar or different in terms of brain areas and cell type. Methods: We used mice expressing the eGFP protein in D1 receptor positive (D1R(+)) neurons to determine whether Δ FosB induced by enriched environment or cocaine injections (5×15 mg/kg) would occur in selective subpopulations of neurons in several subregions of the striatum and prefrontal cortex. Results: We found that: (1) exposure to enriched environment reduces cocaine-induced locomotor activation, confirming our previous findings; (2) exposure to enriched environment by itself increases the accumulation of Δ FosB mostly in D1R(-) cells in the shell part of the nucleus accumbens and dorsal striatum, whereas in the nucleus accumbens core, Δ FosB accumulates in both D1R(+) and D1R(-) neurons; (3) in standard environment mice, cocaine induces accumulation of Δ FosB selectively in D1R(+) cells in the nucleus accumbens, dorsal striatum, and infralimbic cortex; and (4) the effects of enriched environments and cocaine on accumulation of Δ FosB were reciprocally blocked by their combination. Conclusions: Altogether, these results suggest that the enriched environment-induced reduction in behavioral effects of cocaine might result from 2 distinct effects on ΔFosB in striatal medium-sized spiny neurons belonging to the direct and indirect pathways.

Functional interaction between Lypd6 and nicotinic acetylcholine receptors.

Nicotinic acetylcholine receptors (nAChRs) affect multiple physiological functions in the brain and their functions are modulated by regulatory proteins of the Lynx family. Here, we report for the first time a direct interaction of the Lynx protein LY6/PLAUR domain-containing 6 (Lypd6) with nAChRs in human brain extracts, identifying Lypd6 as a novel regulator of nAChR function. Using protein cross-linking and affinity purification from human temporal cortical extracts, we demonstrate that Lypd6 is a synaptically enriched membrane-bound protein that binds to multiple nAChR subtypes in the human brain. Additionally, soluble recombinant Lypd6 protein attenuates nicotine-induced hippocampal inward currents in rat brain slices and decreases nicotine-induced extracellular signal-regulated kinase phosphorylation in PC12 cells, suggesting that binding of Lypd6 is sufficient to inhibit nAChR-mediated intracellular signaling. We further show that perinatal nicotine exposure in rats (4 mg/kg/day through minipumps to dams from embryonic day 7 to post-natal day 21) significantly increases Lypd6 protein levels in the hippocampus in adulthood, which did not occur after exposure to nicotine in adulthood only. Our findings suggest that Lypd6 is a versatile inhibitor of cholinergic signaling in the brain, and that Lypd6 is dysregulated by nicotine exposure during early development. Regulatory proteins of the Lynx family modulate the function of nicotinic receptors (nAChRs). We report for the first time that the Lynx protein Lypd6 binds to nAChRs in human brain extracts, and that recombinant Lypd6 decreases nicotine-induced ERK phosphorylation and attenuates nicotine-induced hippocampal inward currents. Our findings suggest that Lypd6 is a versatile inhibitor of cholinergic signaling in the brain.

Statins Reduce the Risks of Relapse to Addiction in Rats.

Statins are drugs that have been used for decades in humans for the treatment of hypercholesterolemia. More recently, several lines of evidence demonstrate that statins, in addition to their peripheral effects, produce a wide variety of effects in the brain and may be beneficial in neurological and psychiatric conditions. In this study, we allowed rats to self-administer cocaine for several weeks and, at the end of self-administration training, we treated them with low doses of statins daily for a 21-day period of abstinence. Chronic administration of brain-penetrating statins, simvastatin (1 mg/kg) and atorvastatin (1 mg/kg), reduced cocaine seeking compared with vehicle, whereas administration of pravastatin (2 mg/kg), a statin with low brain penetrability, did not. Importantly, the effects of brain-penetrating statins persisted even after discontinuation of the treatment and were specific for drug seeking because drug taking was not altered by simvastatin treatment. Finally, the effects of simvastatin were found to generalize to another drug of abuse such as nicotine, but not to food reward, and to reinstatement of cocaine seeking induced by stress. These results demonstrate that brain-penetrating statins can reduce risks of relapse to addiction. Given their well-known safety profile in humans, statins could be a novel effective treatment for relapse to cocaine and nicotine addiction and their use could be implemented in clinical settings without major health risks.

Chronic stimulation of the tone of endogenous anandamide reduces cue- and stress-induced relapse in rats.

BACKGROUND: The endogenous cannabinoid system plays an important role in motivation, stress, and drug abuse. Pharmacologically, the endocannabinoid system can be stimulated by either agonists of CB1 receptors or inhibition of metabolic degradation of endogenous cannabinoids and consequent increases in their brain levels. METHODS: Here, we investigated whether chronic administration during a period of withdrawal of the fatty acid amide hydrolase inhibitor URB597, which increases anandamide levels, would decrease the risks of relapse to cocaine seeking. Rats were allowed to self-administer cocaine and then they underwent forced withdrawal for 28 days, during which they were treated with URB597 or vehicle. One day after the last injection, we investigated cocaine seeking in one 6h extinction session and relapse triggered by re-exposure to drug-associated cues or a pharmacological stressor. RESULTS: We found that administration of URB597 significantly decreases cocaine-seeking behavior and cue- and stress-induced relapse. CONCLUSION: These results suggest that stimulation of the endocannabinoid system could be helpful to prevent relapse to cocaine addiction.

Prior stimulation of the endocannabinoid system prevents methamphetamine-induced dopaminergic neurotoxicity in the striatum through activation of CB2 receptors.

Methamphetamine toxicity is associated with cell death and loss of dopamine neuron terminals in the striatum similar to what is found in some neurodegenerative diseases. Conversely, the endocannabinoid system (ECS) has been suggested to be neuroprotective in the brain, and new pharmacological tools have been developed to increase their endogenous tone. In this study, we evaluated whether ECS stimulation could reduce the neurotoxicity of high doses of methamphetamine on the dopamine system. We found that methamphetamine alters the levels of the major endocannabinoids, anandamide (AEA) and 2-arachidonoyl glycerol (2-AG) in the striatum, suggesting that the ECS participates in the brain responses to methamphetamine. Δ(9)-tetrahydrocannabinol (THC), a cannabis-derived agonist of both CB1 and CB2 cannabinoid receptors, or inhibitors of the main enzymes responsible for the degradation of AEA and 2-AG (URB597 and JZL184, respectively), blunted the decrease in striatal protein levels of tyrosine hydroxylase induced by methamphetamine. In addition, antagonists of CB2, but not of CB1, blocked the preventive effects of URB597 and JZL184, suggesting that only the former receptor subtype is engaged in neuroprotection exerted by ECS stimulation. Finally, we found that methamphetamine increases striatal levels of the cytokine tumor necrosis factor alpha, an effect that was blocked by ECS stimulation. Altogether, our results indicate that stimulation of ECS prior to the administration of an overdose of methamphetamine considerably reduces the neurotoxicity of the drug through CB2 receptor activation and highlight a protective function for the ECS against the toxicity induced by drugs and other external insults to the brain. This article is part of the Special Issue entitled 'CNS Stimulants'.

Loss of environmental enrichment increases vulnerability to cocaine addiction.

Life experiences, especially during critical periods of maturation, such as adolescence, can dramatically affect vulnerability to diseases at adulthood. Early exposure to positive environmental conditions such as environmental enrichment (EE) has been shown to reduce the occurrence and the intensity of neurological and psychiatric disorders including drug addiction. However, whether or not exposure to EE during early stages of life would protect from addiction when, at adulthood, individuals may find themselves in non-enriched conditions has not been investigated. Here we show that switching mice from EE to non-enriched standard environments not only results in the loss of the preventive effects of EE but also increases the rewarding effects of cocaine. This enhanced vulnerability is associated with emotional distress and with increased levels in the mRNA levels of corticotropin releasing factor (CRF) in the bed nucleus of the stria terminalis (BNST), as well as with increases in CREB phosphorylation in the BNST and in the shell of the nucleus accumbens. The increased sensitivity to the rewarding effects of cocaine is completely blocked by the CRF antagonist antalarmin, confirming a major role of the CRF system in the negative consequences of this environmental switch. These results indicate that positive life conditions during early stages of life, if they are not maintained at adulthood, may have negative emotional consequences and increase the risks to develop drug addiction.

Neuroprotection by neuropeptide Y in cell and animal models of Parkinson's disease.

This study was aimed to investigate the potential neuroprotective effect of neuropeptide Y (NPY) on the survival of dopaminergic cells in both in vitro and in animal models of Parkinson's disease (PD). NPY protected human SH-SY5Y dopaminergic neuroblastoma cells from 6-hydroxydopamine-induced toxicity. In rat and mice models of PD, striatal injection of NPY preserved the nigrostriatal dopamine pathway from degeneration as evidenced by quantification of (1) tyrosine hydroxylase (TH)-positive cells in the substantia nigra pars compacta, levels of (2) striatal tyrosine hydroxylase and dopamine transporter, (3) dopamine and 3,4-dihydroxyphenylacetic acid (DOPAC) as well as (4) rotational behavior. NPY had no neuroprotective effects in mice treated with Y(2) receptor antagonist or in transgenic mice deficient for Y(2) receptor suggesting that NPY effects are mediated through this receptor. Stimulation of Y(2) receptor by NPY triggered the activation of both the ERK1/2 and Akt pathways but did not modify levels of brain derived neurotrophic factor (BDNF) or glial cell line-derived neurotrophic factor. These results open new perspectives in neuroprotective therapies using NPY and suggest potential beneficial effects in PD.

Early exposure to environmental enrichment alters the expression of genes of the endocannabinoid system.

Early environmental enrichment (EE) produces several changes in gene expression in the brain and confers protection against the behavioral, neurochemical and molecular effects of repeated administration of drugs of abuse. Because the endogenous cannabinoid system (ECS) is known to play an important role in the rewarding effects of drugs, we investigated whether the positive effects of early exposure to EE are associated with changes in the expression of genes encoding for proteins that belong to the ECS in C57 mice. Using in situ hybridization, we compared the expression of the cannabinoid receptor CB1, fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MGL) enzymes in brain regions involved in drug addiction in mice reared in either EE or standard environments (SE) from weaning until adulthood. We found that EE increases CB1 mRNA levels in the hypothalamus and in the basolateral amygdala but decreased them in the basomedial amygdala. Similarly, we found that FAAH mRNA levels are higher in the hypothalamus and the basolateral amygdala of EE mice compared to SE mice, with no change in the basomedial amygdala. In contrast, MGL mRNA levels were not affected by EE in any of the areas analyzed. The regional selectivity of EE-induced changes may indicate that early exposure to EE induces changes in the ECS that could result in reduced responses to stress, as confirmed in EE mice in a novelty-induced suppression of feeding test, and, ultimately, in resistance to addiction.

Environmental enrichment does not reduce the rewarding and neurotoxic effects of methamphetamine.

Abuse of amphetamine analogues, such as methamphetamine (METH), represents an important health problem because of their powerful addictive and neurotoxic effects. Abuse of METH induces dopamine neuron terminals loss and cell death in the striatum similar to what is found in other neurodegenerative processes. Exposing mice and rats to enriched environments (EE) has been shown to produce significant protective effects against drug-induced reward as well as against neurodegenerative processes. Here, we investigated whether exposure to EE could reduce the METH-induced reward and neurotoxicity. For this, we reared mice for 2 months during early stages of life in standard environments or EE and then, at adulthood, we tested the ability of METH to induce conditioned place preference and neurotoxicity. We found that, contrary to what we found with other drugs such as cocaine and heroin, EE was unable to reduce the rewarding effects of METH. In addition, contrary to what we found with other toxins such as MPTP, EE did not diminish the striatal neurotoxicity induced by METH (4 x 10 mg/kg) as measured by dopamine content, tyrosine hydroxylase protein levels and apoptosis. Our results demonstrate that the rewarding and neurotoxic effects of METH are not reduced by EE and highlight the great risks associated with the increased popularity of this drug amongst the young population.

NPY promotes chemokinesis and neurogenesis in the rat subventricular zone.

The subventricular zone (SVZ) is a major reservoir for stem cells in the adult mammalian brain. Neural stem cells supply the olfactory bulb with new interneurons and provide cells that migrate towards lesioned brain areas. Neuropeptide Y (NPY), one of the most abundant neuropeptides in the brain, was previously shown to induce neuroproliferation on mice SVZ cells. In the present study, performed in rats, we demonstrate the endogenous synthesis of NPY by cells in the SVZ that suggests that NPY could act as an autocrine/paracrine factor within the SVZ area. We observed that NPY promotes SVZ cell proliferation as previously reported in mice, but does not affect self-renewal of SVZ stem cells. Additionally, this study provides the first direct evidence of a chemokinetic activity of NPY on SVZ cells. Using pharmacological approaches, we demonstrate that both the mitogenic and chemokinetic properties of NPY involve Y1 receptor-mediated activation of the ERK1/2 MAP kinase pathway. Altogether, our data establish that NPY through Y1 receptors activation controls chemokinetic activity and, as for mice, is a major neuroproliferative regulator of rat SVZ cells.