Electronic Nicotine Delivery Systems (ENDS) Flavorants Act As Non‐Competitive Antagonists of nAChRs and Contribute To Reinforcement‐Related Behavior

Abstract

Significance Approximately 42% of the American population who use ENDS are between the ages of 12 and 18. The dramatic rise in adolescent ENDS use is likely attributed to the continuously growing flavor market, with more than 15,000 options available. In an attempt to combat this rise in vaping, it is crucial we determine the pharmacological impact of ENDS flavorants on nicotinic acetylcholine receptors (nAChRs) in the addiction-related circuitry of the brain. Methods We utilized cultured neuroblastoma-2a cells transiently transfected to contain α4β2 and α6β2β3 nAChRs and Ca2+ influx assays to examine the pharmacology of ENDS flavorants on nAChR function. In parallel, flavorants were also examined for their impact on nAChR upregulation using confocal microscopy. Lastly, we utilized vapor self-administration assays with male and female mice to study how these flavorants alter reinforcement-related behaviors. To determine significance, we used one- and two-way ANOVAs with appropriate post-hoc analyses. Results Menthol and two green apple flavorants (farnesol and farnesene) exhibited pharmacology consistent with the designation of NCAs on α4β2 nAChRs (IC50 values of 50–100μM). Further, both menthol and farnesol induced nAChR upregulation, whereas farnesene uniquely induced a stoichiometric shift towards high-sensitivity nAChRs. In self-administration assays, we observed that male mice self-administered menthol + nicotine, green apple + nicotine, or green apple alone, and both green apple and menthol enhanced nicotine reinforcement-related behavior. However, only green apple flavorants contributed to reinforcement-related behavior in the absence of nicotine. Conclusions Although ENDS flavors have been deemed “generally recognized as safe” by the FDA, we have identified several chemical flavorants that act as NCAs of α4β2 nAChRs and exert an impact on reinforcement-related behavior in a mouse model of vaping-related behavior. Given the constant rise in flavored ENDS use, further studies on flavors, ENDS devices, and the underlying mechanisms that take place while using these products is necessary to contribute in the regulation of ENDS use.