There are two main novel findings in the current study. Although varenicline did not increase P50 across all subjects and conditions, data indicate that it acts electrophysiologically as a functional agonist to replace nicotine during periods of smoking cessation. Second, acute nicotine has the same effects on the mouse P20 as smoking does on the P50 in humans. This is a crucial translational link for studies that make such an assumption without direct data. Similarly, varenicline has the same effect on the mouse P20 as it does on the human P50. There has been much debate regarding how ERP components in mouse and human align. This study provides very strong support that the mouse P20 is the appropriate correlate of the human P50.
We found that nicotine in mice and smoking (vs. abstinence) in humans enhanced habituation of the P20 and P50, respectively. In both species, enhancement of habituation involved an increased response to S1 without a change in the response to S2. This finding is consistent with previous reports that S1 amplitude changes, in the absence of S2 amplitude changes, can be observed following cholinergic modulation of auditory habituation (Crawford et al., 2002
; Metzger et al., 2007
; Rudnick, Koehler, Picciotto, & Siegel, 2009
). Varenicline increased P20 amplitude in mice without an effect on habituation. In the human crossover study, subjects receiving placebo during Phase 1 exhibited decreased P50 amplitude during abstinence, which was attenuated by varenicline during Phase 2. Subjects receiving varenicline during Phase 1 followed by placebo during Phase 2 exhibited no change in P50 amplitude during either treatment phase. Although we cannot offer a definitive explanation for the effect of treatment order, a pharmacologic carryover effect cannot be ruled out. In summary, these findings support the hypothesis that varenicline can modulate amplitude of the P20 and P50 components ().
Consolidated results from mouse and human experiments demonstrate translational validity of event-related potentials technique
We show an acute effect of smoking status on P50 habituation in healthy current smokers. While previous studies employed brief periods of abstinence (6–15 hr), we employed a longer period of abstinence (3 days), which was confirmed by exhaled CO (Adler et al., 1993
; Crawford et al., 2002
; Domino, 2003
; Domino & Kishimoto, 2002
). Therefore, our paradigm may have been more sensitive to effects of smoking and abstinence compared with studies with a shorter duration of abstinence. In vivo radiotracer experiments suggest that nicotine can take several days to clear high-affinity binding sites (i.e., α4β2 nAChRs) in humans and nonhuman primates (Staley et al., 2006
). Therefore, the effects of abstinence may follow a protracted time course. One potential drawback of our approach is that each subject started from a different level of baseline smoking. We controlled for this possibility by including self-reported baseline cigarette consumption as a covariate in our analyses.
Auditory habituation depends on the responses to S1 and S2, and frequently, these amplitudes are condensed into a single ratio. This approach can obscure the mechanism of habituation enhancements, which may depend on an increased response to S1, decreased response to S2, or both. It has been proposed that nicotine inhibits the response to S2 by activating α7 nAChRs interneurons in the CA3 region of the hippocampus (Adler et al., 1998
; Stevens et al., 1998
). However, this mechanism alone is not sufficient to explain our data because nicotine and smoking increased the amplitude of the S1 response without significantly affecting the amplitude of the S2 response. Previous studies in rodents and humans show similar changes in the response to S1 but not S2 (Crawford et al., 2002
; Cromwell & Woodward, 2007
; Metzger et al., 2007
; Phillips et al., 2007
). Varenicline, a relatively selective α4β2 nAChRs partial agonist (Mihalak et al., 2006
), increased amplitude but not habituation of auditory ERPs. Therefore, it is likely that brain regions rich in α4β2 nAChRs, the target of varenicline, contribute to the amplifying effect of nicotine on the S1 response. Consistent with this hypothesis, nicotine enhances action potential propagation and synaptic release in thalamocortical circuits via DHβE-sensitive nAChRs (Kawai et al., 2007
; Lambe, Picciotto, & Aghajanian, 2003
). These circuits are an obligate stage of auditory processing and participate in generation of the midlatency ERP components (Hinman & Buchwald, 1983
; McGee, Kraus, Comperatore, & Nicol, 1991
). Even if nicotine increases the response to S1 by enhancing thalamic transmission, it must also activate inhibitory networks in order to prevent the response to S2 from increasing in amplitude as well. Therefore, it is likely that both α7 nAChRs- and α4β2 nAChRs-expressing brain regions are involved in auditory habituation. Our initial hypothesis was that varenicline would attenuate the effects of nicotine in the presence of nicotine but mimic the effects of a full agonist when given alone, consistent with activity as a partial agonist. Data support that varenicline acted as a functional agonist when given alone. The combination of varenicline and smoking were similar to either smoking or varenicline alone.
Varenicline increased P20 amplitude in mice and P50 amplitude during abstinence in humans. Changes in EEG power and ERP amplitude are thought to reflect changes in arousal (Kishimoto & Domino, 1998
; Pickworth, Herning, & Henningfield, 1989
). Because decreased arousal is a symptom of nicotine withdrawal, varenicline’s effects on sensory habituation may contribute to its therapeutic efficacy. However, nonspecific increases in arousal may also contribute to sleep disturbances observed in some clinical studies (Gonzales et al., 2006
; Oncken et al., 2006
). Nonetheless, the aforementioned connection between ERP amplitude and arousal remains speculative because stimulants such as amphetamine can decrease amplitude (Maxwell, Kanes, et al., 2004
). Interestingly, the smoking cessation medication bupropion reduces the amplitude of ERPs in mice, similar to amphetamine (Siegel et al., 2005
). To the best of our knowledge, the effects of bupropion on human ERPs are not known. Although alpha-7 nAChRs agonists such as 3-(2,4)-dimethoxybenzylidine anabaseine and tropisetron reverse the effects of cocaine or amphetamine on ERPs, their effects on P50 or smoking status in humans are not known (Hashimoto, Iyo, Freedman, & Stevens, 2005
; Stevens et al., 1999
). The most direct interpretation of increased ERP amplitude may simply be that it reflects a greater degree of phase synchrony in ongoing EEG rhythms (Jansen, Agarwal, Hegde, & Boutros, 2003
; Makeig et al., 2002
Our human data reveal that treatment order significantly modulated the effects of abstinence and varenicline on P50 amplitude. It is possible that subjects receiving varenicline prior to placebo failed to undergo a reduction in P50 amplitude during the placebo phase because varenicline had long-lasting effects in the brain, despite the 5- to 7-day washout period and 17-hr half-life of varenicline (Obach et al., 2006
). An alternative explanation for the order effect may be that abstinence failed to reduce P50 amplitude because of a floor effect. Overall P50 amplitudes also differed by treatment order, suggesting that there may have been baseline asymmetries in sensory processing between subjects randomized to different treatment orders; however, in the absence of baseline ERP data, we cannot examine this directly. To simplify interpretation of results, future within-subject designs may benefit from extending the washout period to 2 weeks.
Previous studies indicate that nicotine in rodents produces similar biological and behavioral effects as smoking in humans, and this study further supports the face validity of mouse models (Blendy et al., 2005
; Corrigall, 1999
; Lerman et al., 2007
; Liu et al., 2003
; Slawecki, Gilder, Roth, & Ehlers, 2003
). However, there are limitations to this approach. Besides nicotine, cigarettes contain psychoactive compounds such as monoamine oxidase inhibitors that may play a role in dependence and auditory habituation (Crawford et al., 2002
; Guillem et al., 2005
; Siegel et al., 2005
). Furthermore, our mouse study used acute doses of nicotine, which likely fail to produce the types of nAChRs upregulation associated with chronic smoking. Although previous studies in mice have shown biological effects at the dose of varenicline used in the present study, the lack of a dose–response relationship for ERPs is a potential limitation since we cannot determine the effects of higher doses, which may have increased P50 amplitude in the VP group (Raybuck et al., 2008
; Rollema et al., 2009
). Despite these caveats, the extensive interspecies overlap in our results suggests that acute doses of cholinergic agents approximate chronic exposure in humans. Thus, EEG in mice allows for rapid screening of novel treatments for ND that may ameliorate sensory deficits associated with abstinence (Lerman et al.).