Validation of a nicotine vapor self-administration model in rats with relevance to electronic cigarette use

Abstract

The debate about electronic cigarettes is dividing healthcare professionals, policymakers, manufacturers, and communities. A key limitation in our understanding of the cause and consequences of vaping is the lack of animal models of nicotine vapor self-administration. Here, we developed a novel model of voluntary electronic cigarette use in rats using operant behavior. We found that rats voluntarily exposed themselves to nicotine vapor to the point of reaching blood nicotine levels that are similar to humans. The level of responding on the active (nicotine) lever was similar to the inactive (air) lever and lower than the active lever that was associated with vehicle (polypropylene glycol/glycerol) vapor, suggesting low positive reinforcing effects and low nicotine vapor discrimination. Lever pressing behavior with nicotine vapor was pharmacologically prevented by the α4β2 nicotinic acetylcholine receptor partial agonist and α7 receptor full agonist varenicline in rats that self-administered nicotine but not vehicle vapor. Moreover, 3 weeks of daily (1 h) nicotine vapor self-administration produced addiction-like behaviors, including somatic signs of withdrawal, allodynia, anxiety-like behavior, and relapse-like behavior after 3 weeks of abstinence. Finally, 3 weeks of daily (1 h) nicotine vapor self-administration produced cardiopulmonary abnormalities and changes in α4, α3, and β2 nicotinic acetylcholine receptor subunit mRNA levels in the nucleus accumbens and medial prefrontal cortex. These findings validate a novel animal model of nicotine vapor self-administration in rodents with relevance to electronic cigarette use in humans and highlight the potential addictive properties and harmful effects of chronic nicotine vapor self-administration.

Fig. 1. Method development and experimental design.

a Schematic diagram of the operant vapor self-administration chambers. b Comparison of blood nicotine levels between rats that self-administered nicotine vapor (0.5 mg/ml) and rats that self-administered nicotine intravenously (0.03 mg/kg/infusion) in a 1-h session. Error bars represent the SEM of blood nicotine concentration (n = 3–4 rats). c Experimental timeline of self-administration in the nicotine vapor group and vehicle vapor group. d Self-administration of vehicle vapor (0 mg/ml) and nicotine vapor (0.05, 0.5, 5, and 50 mg/ml, 1 session/dose) in rats. The data are expressed as the mean ± SEM of 16–24 rats. e Blood nicotine levels in rats that self-administered vapor (0, 0.05, 0.5, 5, and 50 mg/ml nicotine, 1 session/dose). No nicotine was detected in blood from rats that self-administered vehicle vapor or vapor that contained 0.05 mg/ml nicotine. The data are expressed as the mean ± SEM of 4–5 rats.

Fig. 2. Varenicline decreased nicotine vapor self-administration.

a Experimental timeline of self-administration in the nicotine vapor group and vehicle vapor group. b Acquisition and maintenance of nicotine vapor self-administration during 2 weeks of daily short access (1 h) to nicotine vapor (0.5 mg/ml, n = 24 [12 females, 12 males]) or vehicle vapor (1:1, propylene glycol:glycerol, n = 24 [13 females, 11 males]). Error bars represent the SEM of lever presses. c Nicotine vapor and vehicle vapor self-administration following pretreatment with varenicline (1.5 mg/ml, s.c.). Error bars represent the SEM of lever presses.

Fig. 3. Rats that self-administered nicotine vapor exhibited dependence-like behaviors during mecamylamine-precipitated withdrawal.

a Experimental timeline of the behavioral analysis in the nicotine vapor group and vehicle vapor group. b Withdrawal scores in rats that self-administered nicotine vapor (0.5 mg/ml) increased following subcutaneous mecamylamine administration (0.5 or 1.5 mg/ml). Error bars represent the SEM of withdrawal score (n = 24 rats [12 females, 12 males]). c Withdrawal scores in rats that self-administered vehicle vapor (1:1, propylene glycol:glycerol) following subcutaneous mecamylamine administration did not change (0.5 mg/ml). Error bars represent the SEM of withdrawal scores (n = 24 rats [13 females, 11 males]). d The percent change in pain thresholds relative to baseline (BSL) in rats that self-administered nicotine vapor (0.5 mg/ml) decreased following subcutaneous mecamylamine administration (0.5 mg/ml). Error bars represent the SEM of the percent change in pain thresholds relative to baseline (n = 24 rats [12 males, 12 females]). e Mecamylamine had no effect on pain thresholds in rats that self-administered vehicle vapor (0.5 mg/ml). Error bars represent the SEM of the percent change in pain thresholds relative to baseline (n = 24 rats [13 males, 11 females]).

Fig. 4. Rats that self-administered nicotine vapor exhibited addiction-like behaviors after 3 weeks of protracted abstinence.

a Experimental timeline of the behavioral analysis in the nicotine vapor group and vehicle vapor group. b The time spent in the open arms of the elevated plus maze decreased in rats that self-administered nicotine vapor compared with vehicle vapor. Error bars represent the SEM of the time spent on the open arms (n = 24 rats [12–13 females, 11–12 males]). c The percent change in pain thresholds relative to baseline in rats that self-administered nicotine vapor decreased compared with their own baseline (BSL) and with vehicle vapor during protracted abstinence following a 3-week incubation period. Error bars represent the SEM of the percent change in pain thresholds relative to baseline (n = 24 rats [12–13 females, 11–12 males]). d Number of lever presses in the nicotine vapor group (n = 24 rats [12 females, 12 males]) and vehicle vapor group (n = 24 rats [13 females, 11 males]) in a 1-h session before (self-administration [SA]) and after (reexposure [R]) a 3-week incubation period. Error bars represent the SEM of the number of lever presses.

Fig. 5. Three weeks of daily (1 h) nicotine vapor self-administration produced alveolar simplification.

a Experimental timeline of organ harvest in the nicotine vapor group and vehicle vapor group. Representative images of hematoxylin and eosin staining of the lungs in rats that self-administered (b) chronic vehicle vapor, c acute nicotine vapor (0.5 mg/ml, one session), d chronic nicotine vapor (0.5 mg/ml, 23 sessions), and e chronic nicotine vapor (0.5 mg/ml, 23 sessions) after 3 weeks of abstinence. f The mean linear intercept of alveolar airspace distance increased in rats that chronically self-administered nicotine vapor compared with vehicle vapor and acute self-administration. Error bars represent the SEM of six tissue sample replicates per rat (n = 4 rats [2 females, 2 males]).

Fig. 6. Receptor dysregulation after protracted abstinence from nicotine vaping.

a Experimental timeline of brain harvest in the nicotine vapor group and vehicle vapor group. b Change in α4 nicotinic acetylcholine receptor subunit gene expression in the nucleus accumbens in rats that self-administered vehicle vapor and nicotine vapor after 3 weeks of protracted abstinence. c Change in β2 nicotinic acetylcholine receptor subunit gene expression in the nucleus accumbens in rats that self-administered vehicle vapor and nicotine vapor after 3 weeks of protracted abstinence. d Change in α3 nicotinic acetylcholine receptor subunit gene expression in the nucleus accumbens in rats that self-administered vehicle vapor and nicotine vapor after 3 weeks of protracted abstinence. e Change in α4 nicotinic acetylcholine receptor subunit gene expression in the medial prefrontal cortex in rats that self-administered vehicle vapor and nicotine vapor after 3 weeks of protracted abstinence. f Change in β2 nicotinic acetylcholine receptor subunit gene expression in the medial prefrontal cortex in rats that self-administered vehicle vapor and nicotine vapor after 3 weeks of protracted abstinence. g Change in α3 nicotinic acetylcholine receptor subunit gene expression in the medial prefrontal cortex in rats that self-administered vehicle vapor and nicotine vapor after 3 weeks of protracted abstinence. Error bars represent the mean ± SEM of the fold change in gene expression, normalized to the vapor group (n = 7–8 rats).