Affective Signs Of Nicotine Withdrawal Research Articles

N-oleoyl alanine attenuates nicotine reward and spontaneous nicotine withdrawal in mice

BackgroundAs nicotine dependence represents a longstanding major public health issue, new nicotine cessation pharmacotherapies are needed. Administration of N-oleoyl glycine (OlGly), an endogenous lipid signaling molecule, prevents nicotine-induced conditioned place preference (CPP) through a peroxisome proliferator-activated receptor-alpha (PPARα) dependent mechanism, and also ameliorated withdrawal signs in nicotine-dependent mice. Pharmacological evidence suggests that the methylated analog of OlGly, N-oleoyl alanine (OlAla), has an increased duration of action and may offer translational benefit. Accordingly, OlAla was assessed in nicotine CPP and dependence assays as well as its pharmacokinetics compared to OlGly. MethodsICR female and male mice were tested in nicotine-induced CPP with and without the PPARα antagonist GW6471. OlAla was also assessed in nicotine-dependent mice following removal of nicotine minipumps: somatic withdrawal signs, thermal hyper-nociception and altered affective behavior (i.e., light/dark box). Finally, plasma and brain were collected after administration of OlGly or OlAla and analyzed by high-performance liquid chromatography tandem mass spectrometry. ResultsOlAla prevented nicotine-induced CPP, but this effect was not blocked by GW6471. OlAla attenuated somatic and affective nicotine withdrawal signs, but not thermal hyper-nociception in nicotine-dependent mice undergoing withdrawal. OlAla and OlGly showed similar time-courses in plasma and brain. ConclusionsThe observation that both molecules showed similar pharmacokinetics argues against the notion that OlAla offers increased metabolic stability. Moreover, while these structurally similar lipids show efficacy in mouse models of reward and dependence, they reduce nicotine reward through distinct mechanisms.

Differential roles of diacylglycerol lipase (DAGL) enzymes in nicotine withdrawal

Nicotine and tobacco-related deaths remains a leading cause of preventable death and disease in the United States. Several studies indicate that modulation of the endocannabinoid system, primarily of the endocannabinoid 2-Arachidonoylglycerol (2-AG), alters nicotinic dependence behaviors in rodents. This study, using transgenic knock-out (KO) mice, evaluated the role of the two 2-AG biosynthesis enzymes, (Diacylglycerol lipase-α) DAGL-α and DAGL-β in spontaneous nicotine withdrawal. DAGL-α deletion prevents somatic and affective signs of nicotine withdrawal, while DAGL-β deletion plays a role in hyperalgesia due to nicotine withdrawal. These results suggest a differential role of these enzymes in the various signs of nicotine withdrawal. Our behavioral findings relate to the distribution of these enzymes with DAGL-β being highly expressed in macrophages and DAGL-α in neurons. This study offers new potential targets for smoking cessation therapies.

The role of pituitary adenylyl cyclase activating polypeptide in affective signs of nicotine withdrawal.

Recent evidence implicates endogenous pituitary adenylyl cyclase activating polypeptide (PACAP) in the aversive effect of nicotine. In the present study, we assessed if nicotine-induced conditioned place preference (CPP) or affective signs of nicotine withdrawal would be altered in the absence of PACAP and if there were any sex-related differences in these responses. Male and female mice lacking PACAP and their wild-type controls were tested for baseline place preference on day 1, received conditioning with saline or nicotine (1mg/kg) on alternate days for 6days and were then tested for CPP the next day. Mice were then exposed to four additional conditioning and were tested again for nicotine-induced CPP 24hr later. Controls were conditioned with saline in both chambers and tested similarly. All mice were then, 96hr later, challenged with mecamylamine (3mg/kg), and tested for anxiety-like behaviors 30min later. Mice were then, 2hr later, forced to swim for 15min and then tested for depression-like behaviors 24hr later. Our results showed that male but not female mice lacking PACAP expressed a significant CPP that was comparable to their wild-type controls. In contrast, male but not female mice lacking PACAP exhibited reduced anxiety- and depression-like behaviors compared to their wild-type controls following the mecamylamine challenge. These results suggest that endogenous PACAP is involved in affective signs of nicotine withdrawal, but there is a sex-related difference in this response.

Activation of PPARγ Attenuates the Expression of Physical and Affective Nicotine Withdrawal Symptoms through Mechanisms Involving Amygdala and Hippocampus Neurotransmission.

An isoform of peroxisome proliferator-activated receptors (PPARs), PPARγ, is the receptor for the thiazolidinedione class of anti-diabetic medications including pioglitazone. Neuroanatomical data indicate PPARγ localization in brain areas involved in drug addiction. Preclinical and clinical data have shown that pioglitazone reduces alcohol and opioid self-administration, relapse to drug seeking, and plays a role in emotional responses. Here, we investigated the behavioral effect of PPARγ manipulation on nicotine withdrawal in male Wistar rats and in male mice with neuron-specific PPARγ deletion (PPARγ(-/-)) and their littermate wild-type (PPARγ(+/+)) controls. Real-time quantitative RT-PCR and RNAscope in situ hybridization assays were used for assessing the levels of expression and cell-type localization of PPARγ during nicotine withdrawal. Brain site-specific microinjections of the PPARγ agonist pioglitazone were performed to explore the role of this system on nicotine withdrawal at a neurocircuitry level. Results showed that activation of PPARγ by pioglitazone abolished the expression of somatic and affective nicotine withdrawal signs in rats and in (PPARγ(+/+)) mice. This effect was blocked by the PPARγ antagonist GW9662. During early withdrawal and protracted abstinence, the expression of PPARγ increased in GABAergic and glutamatergic cells of the amygdala and hippocampus, respectively. Hippocampal microinjections of pioglitazone reduced the expression of the physical signs of withdrawal, whereas excessive anxiety associated with protracted abstinence was prevented by pioglitazone microinjection into the amygdala. Our results demonstrate the implication of the neuronal PPARγ in nicotine withdrawal and indicates that activation of PPARγ may offer an interesting strategy for smoking cessation.SIGNIFICANCE STATEMENT Smoking cessation leads the occurrence of physical and affective withdrawal symptoms representing a major burden to quit tobacco use. Here, we show that activation of PPARγ prevents the expression of both somatic and affective signs of nicotine withdrawal. At molecular levels results show that PPARγ expression increases in GABAergic cells in the hippocampus and in GABA- and glutamate-positive cells in the basolateral amygdala. Hippocampal microinjections of pioglitazone reduce the insurgence of the physical withdrawal signs, whereas anxiety linked to protracted abstinence is attenuated by pioglitazone injected into the amygdala. Our results demonstrate the implication of neuronal PPARγ in nicotine withdrawal and suggest that PPARγ agonism may represent a promising treatment to aid smoking cessation.

The β3 subunit of the nicotinic acetylcholine receptor is required for nicotine withdrawal-induced affective but not physical signs or nicotine reward in mice

Nicotinic acetylcholine receptors (nAChRs) are the primary target for nicotine, the addictive component in tobacco products. These pentameric receptors are made up of various subunits which contribute to the diverse functions of nAChR subtypes. The β3 subunit of the nAChR has been understudied in nicotine dependence, even though it is expressed in brain regions important for drug reward. Therefore, we assessed nicotine dependence behaviors in β3 wildtype (WT) and knockout (KO) male and female mice. We evaluated nicotine reward in the conditioned place preference (CPP) test and then measured nicotine withdrawal signs after chronic exposure to the drug. For the withdrawal studies, mice were continuously infused with 24 mg/kg/day of nicotine using surgically implanted osmotic mini-pumps for 14 days. Mini-pumps were removed at day 15, and withdrawal signs (somatic signs, hyperalgesia, anhedonia-like measure using the sucrose preference test and anxiety-like behaviors using the light dark boxes) were collected at 24 h intervals for three days following spontaneous withdrawal of nicotine. Nicotine-induced CPP did not differ between β3 KO and WT mice. β3 KO mice displayed similar somatic symptoms and hyperalgesia compared to WT mice but showed significant absence in affective (anhedonia and anxiety-like behaviors) withdrawal signs in nicotine-dependent mice. These observations suggest that the β3 nicotinic subunits do not seem to influence nicotine reward but plays an important role in affective nicotine withdrawal signs. Given the health burden of tobacco use disorder and the modest effect of smoking cessation aids, it is important to understand underlying factor contributing to nicotine dependence. The results of this study will further our knowledge of the role of the β3 nAChR subunit in nicotine reward and withdrawal behaviors in hopes of finding new molecular targets for smoking cessation aids.

Recent Updates in Animal Models of Nicotine Withdrawal: Intracranial Self-Stimulation and Somatic Signs.

Tobacco use is one of the leading causes of disease, disability, and death in the world. Despite the negative health consequences of smoking and the fact that most people would like to quit, very few people are able to maintain abstinence. Nicotine, the main psychoactive component of tobacco, is mildly rewarding and plays a role in the initiation and maintenance of smoking. Cessation of nicotine intake induces mild somatic withdrawal symptoms that may contribute to the maintenance of smoking. However, it has been hypothesized that negative affective nicotine withdrawal signs are of greater motivational significance in contributing to relapse and continued smoking (Markou and Koob (eds) Intracranial self-stimulation thresholds as a measure of reward, vol 2. Oxford University Press, New York, 1993; Koob et al., Semin Neurosci 5:351-358, 1993). Intracranial self-stimulation (ICSS) has been established as a method to assess the acute rewarding properties of nicotine and the dysphoria associated with nicotine withdrawal. The ICSS method provides a means to measure the negative affective aspects of nicotine withdrawal in animal models and may contribute to the understanding of the neurobiological bases of nicotine dependence.

Assessment of nicotine withdrawal-induced changes in sucrose preference in mice

Anhedonia, induced by nicotine withdrawal, may serve as an important affective sign that reinforces tobacco use and smoking relapse rates in humans. Animal models provide a way to investigate the underlying neurobiological factors involved in the decrease in responding for positive affective stimuli during nicotine withdrawal and may aid in drug development for nicotine dependence. Thus, we explored the use of the sucrose preference test to measure nicotine withdrawal-induced reduction in response for positive affective stimuli in mice. C57BL/6J and knockout (KO) mice were chronically exposed to different doses of nicotine through surgically implanted subcutaneous osmotic minipumps for 14days and underwent spontaneous nicotine withdrawal on day 15. A sucrose preference time course was performed and the results were compared to another well-established affective sign of nicotine withdrawal, the reduction in time spent in light side, using the Light Dark Box test. Subsequently, our results demonstrated a time-dependent and dose-related reduction in sucrose preference in nicotine withdrawn male C57BL/6J mice, indicative of a decrease in responding for positive affective stimuli. Furthermore, the sucrose preference reduction during nicotine withdrawal was consistent with decrease in time spent in the light side of the Light Dark Box test. We also found the reduction for positive affective stimuli and time spent in the light side was not present in nicotine withdrawn β2 and α6 KO mice, suggesting that these nicotinic subunits are involved in the affective signs of nicotine withdrawal. Thus, this report highlights the potential utility of the sucrose preference test as a useful measure of the decrease in responding for positive affective stimuli during spontaneous nicotine withdrawal.

Effects of orally-bioavailable short-acting kappa opioid receptor-selective antagonist LY2456302 on nicotine withdrawal in mice

Kappa opioid receptor (KOR) signaling has been implicated in mediating behavioral and biochemical effects associated with drug dependence. The most commonly used KOR antagonists, norbinaltorphimine (norBNI) and (3R)-7-Hydroxy-N{(1S)-1-{[(3R,4R)-4-(3-hydroxyphenyl)-3,4-dimethyl-1-piperidinyl]methyl}-2-methylpropyl}-1,2,3,4-tetrahydro-3-isoquinoline-carboxamide (JDTic), have provided a wealth of information in this area; however, the delayed onset and long-lasting effects of these antagonists complicate experimental design and interpretation of results, and make them less than ideal for clinical studies. Initial studies with the recently developed KOR antagonist, LY2456302, show that the compound is a short acting, high-affinity, selective KOR antagonist with therapeutic potential for mood disorders and ethanol use in animal models, and is well tolerated in humans. The goal of the current study was to evaluate the effectiveness of LY2456302 in alleviating the nicotine withdrawal syndrome in mice. Mice were chronically treated with nicotine for 14 days and physical and affective nicotine withdrawal signs were measured using a spontaneous nicotine withdrawal model and conditioned place aversion (CPA) following pre-treatment with LY2456302, administered orally. Vehicle treated nicotine withdrawn mice displayed significant anxiety-related behavior, somatic signs, hyperalgesia, and CPA. Similar to previous studies with norBNI and JDTic, LY2456302 alleviated the nicotine withdrawal syndrome, as evidenced by decreased expression of nicotine withdrawal induced anxiety-related behavior, somatic signs, and CPA, and increased hotplate latency in nicotine withdrawn mice following pre-treatment. Given the current results, and with its favorable pharmacokinetic and pharmacodynamic profile, LY2456302 may be a useful therapeutic agent for treatment of multiple aspects of the nicotine withdrawal syndrome.

The α3β4* nicotinic acetylcholine receptor subtype mediates nicotine reward and physical nicotine withdrawal signs independently of the α5 subunit in the mouse

The 15q25 gene cluster contains genes that code for the α5, α3, and β4 nicotinic acetylcholine receptor (nAChRs) subunits, and in human genetic studies, has shown the most robust association with smoking behavior and nicotine dependence to date. The limited available animal studies implicate a role for the α5 and β4 nAChR subunits in nicotine dependence and withdrawal; however studies focusing on the behavioral role of the α3β4* nAChR receptor subtype in nicotine dependence are lacking. Because of the apparent role of the α3β4* nAChR subtype in nicotine dependence, the goal of the current study was to better evaluate the involvement of this subtype in nicotine mediated behavioral responses. Using the selective α3β4* nAChR antagonist, α-conotoxin AuIB, we assessed the role of α3β4* nAChRs in acute nicotine, nicotine reward, and physical and affective nicotine withdrawal. Because α5 has also been implicated in nicotine dependence behaviors in mice and can form functional receptors with α3β4*, we also evaluated the role of the α3β4α5* nAChR subtype in nicotine reward and somatic nicotine withdrawal signs by blocking the α3β4* nAChR subtype in α5 nAChR knockout mice with AuIB. AuIB had no significant effect on acute nicotine behaviors, but dose-dependently attenuated nicotine reward and physical withdrawal signs, with no significant effect in affective withdrawal measures. Interestingly, AuIB also attenuated nicotine reward and somatic signs in α5 nAChR knockout mice. This study shows that α3β4* nAChRs mediate nicotine reward and physical nicotine withdrawal, but not acute nicotine behaviors or affective nicotine withdrawal signs in mice. The α5 subunit is not required in the receptor assembly to mediate these effects. Our findings suggest an important role for the α3β4* nAChR subtype in nicotine reward and physical aspects of the nicotine withdrawal syndrome.

Nicotine reward and affective nicotine withdrawal signs are attenuated in calcium/calmodulin-dependent protein kinase IV knockout mice.

The influx of Ca2+ through calcium-permeable nicotinic acetylcholine receptors (nAChRs) leads to activation of various downstream processes that may be relevant to nicotine-mediated behaviors. The calcium activated protein, calcium/calmodulin-dependent protein kinase IV (CaMKIV) phosphorylates the downstream transcription factor cyclic AMP response element binding protein (CREB), which mediates nicotine responses; however the role of CaMKIV in nicotine dependence is unknown. Given the proposed role of CaMKIV in CREB activation, we hypothesized that CaMKIV might be a crucial molecular component in the development of nicotine dependence. Using male CaMKIV genetically modified mice, we found that nicotine reward is attenuated in CaMKIV knockout (−/−) mice, but cocaine reward is enhanced in these mice. CaMKIV protein levels were also increased in the nucleus accumbens of C57Bl/6 mice after nicotine reward. In a nicotine withdrawal assessment, anxiety-related behavior, but not somatic signs or the hyperalgesia response are attenuated in CaMKIV −/− mice. To complement our animal studies, we also conducted a human genetic association analysis and found that variants in the CaMKIV gene are associated with a protective effect against nicotine dependence. Taken together, our results support an important role for CaMKIV in nicotine reward, and suggest that CaMKIV has opposing roles in nicotine and cocaine reward. Further, CaMKIV mediates affective, but not physical nicotine withdrawal signs, and has a protective effect against nicotine dependence in human genetic association studies. These findings further indicate the importance of calcium-dependent mechanisms in mediating behaviors associated with drugs of abuse.

Brain activity and emotional processing in smokers treated with varenicline

Prior evidence suggests that varenicline, an effective smoking cessation treatment, may relieve negative affective signs of nicotine withdrawal. We examined varenicline effects on emotional processing in 25 abstinent smokers after 13 days of varenicline and placebo using a within-subject cross-over design. Blood oxygen level dependent (BOLD) functional magnetic resonance imaging was acquired while subjects completed a face emotion identification task. Results showed a significant drug effect, characterized by decreased BOLD signal in dorsal anterior cingulate/medial frontal cortex, occipital cortex and thalamus. Increased BOLD signal was observed in the middle temporal gyrus. Varenicline improved correct response time; however, neither BOLD signal nor performance effects were moderated by emotion type. An exploratory region of interest analysis suggests that varenicline reduced amygdala activity independent of emotional valence. Taken together, these results suggest that observed drug effects on brain activity do not reflect affective changes but rather enhanced early processing of perceptual features of facial stimuli.

Differential Role of Nicotinic Acetylcholine Receptor Subunits in Physical and Affective Nicotine Withdrawal Signs

It has been suggested that the negative effects associated with nicotine withdrawal promote continued tobacco use and contribute to the high relapse rate of smoking behaviors. Thus, it is important to understand the receptor-mediated mechanisms underlying nicotine withdrawal to aid in the development of more successful smoking cessation therapies. The effects of nicotine withdrawal are mediated through nicotinic acetylcholine receptors (nAChRs); however, the role of nAChRs in nicotine withdrawal remains unclear. Therefore, we used mecamylamine-precipitated, spontaneous, and conditioned place aversion (CPA) withdrawal models to measure physical and affective signs of nicotine withdrawal in various nAChR knockout (KO) mice. beta2, alpha7, and alpha5 nAChR KO mice were chronically exposed to nicotine through surgically implanted osmotic minipumps. Our results show a loss of anxiety-related behavior and a loss of aversion in the CPA model in beta2 KO mice, whereas alpha7 and alpha5 KO mice displayed a loss of nicotine withdrawal-induced hyperalgesia and a reduction in somatic signs, respectively. These results suggest that beta2-containing nAChRs are involved in the affective signs of nicotine withdrawal, whereas non-beta2-containing nAChRs are more closely associated with physical signs of nicotine withdrawal; thus, the nAChR subtype composition may play an important role in the involvement of specific subtypes in nicotine withdrawal.

Adaptations in cholinergic transmission in the ventral tegmental area associated with the affective signs of nicotine withdrawal in rats

Chronic administration of nicotine induces adaptations in the brain reward circuit to counteract the acute drug effects; when nicotine administration ceases, these adaptations remain unopposed and lead to drug withdrawal. The present studies were conducted to assess the effects of chronic nicotine administration on nicotinic acetylcholine receptor (nAChR) activity in the ventral tegmental area (VTA) and the nucleus accumbens (Nacc) shell. A discrete-trial intracranial self-stimulation procedure that provides current-intensity thresholds as measures of brain reward function was used in rats. Previous studies have shown that withdrawal from nicotine-induced elevations in brain reward thresholds that are indicative of a decrease in brain reward function. We show here that injections of the nAChR antagonist dihydro-β-erythroidine (DHβE; 0.6–20 μg total bilateral dose) into the VTA, but not outside the VTA, resulted in significant elevations in brain reward thresholds in nicotine dependent rats (9 mg/kg/day nicotine hydrogen tartrate) while having no effect in saline-treated controls. By contrast, DHβE (0.6–20 μg total bilateral dose) injected into the Nacc shell had no effect on brain reward thresholds of nicotine- or saline-treated rats. The adaptations in cholinergic transmission in the VTA are likely to mediate, at least partly, the affective signs of nicotine withdrawal in humans.