Nicotine vapor inhalation escalates nicotine self-administration

Abstract

Humans escalate their cigarette smoking over time, and a major obstacle in the field of pre-clinical nicotine addiction research has been the inability to produce escalated nicotine self-administration in rats. In experiment 1, male Wistar rats were trained to respond for nicotine in 2-hour operant sessions, then exposed to chronic intermittent (12 hours/day) nicotine vapor and repeatedly tested for nicotine self-administration at 8-12 hours of withdrawal. Rats were tested intermittently on days 1, 3 and 5 of the vapor exposure procedure, then tested with nicotine vapor exposure on 6-15 consecutive days. Rats exhibited transient increases in operant nicotine responding during intermittent testing, regardless of vapor condition, and this responding returned to baseline levels upon resumption of consecutive-days testing (i.e. nicotine deprivation effect). Nicotine vapor-exposed rats then escalated nicotine self-administration relative to both their own baseline (∼200% increase) and non-dependent controls (∼3× higher). In experiment 2, rats were exposed or not exposed to chronic intermittent nicotine vapor, then tested for spontaneous and precipitated somatic signs of nicotine withdrawal. Eight hours following removal from nicotine vapor, rats exhibited robust mecamylamine-precipitated somatic signs of withdrawal. There was a strong correlation between nicotine flow rate and air-nicotine concentration, and the air-nicotine concentrations used in experiments 1 and 2 resemble concentrations experienced by human smokers. Collectively, these results suggest that chronic intermittent nicotine vapor inhalation produces somatic and motivational signs of nicotine dependence, the latter of which is evidenced by escalation of nicotine self-administration.

Keywords: Escalation; mecamylamine; nicotine dependence; nicotine self-administration; withdrawal.

Figure 1

[A] Mean ± SEM nicotine infusions (FR-1 schedule, 20 s timeout) by nicotine vapor-exposed (black circles) and air-exposed (white circles) rats during 2-hr operant sessions prior to (baseline = 3-day mean) and during chronic intermittent (12 hrs/day) nicotine vapor exposure. Rats were tested intermittently during the first 5 days of vapor exposure (72 hrs between end of baseline & day 1 test, 48 hrs between day 1 & 3 tests, and 48 hrs between day 3 & 5 tests). Rats were then tested daily during days 6-15 of nicotine vapor exposure. All tests occurred at 8-12 hrs following removal from nicotine vapor (i.e., withdrawal). Data suggest a tendency toward a nicotine deprivation effect on days 1-5 regardless of nicotine vapor condition, # p=0.057 main effect of day. Also, nicotine vapor-exposed rats exhibited significant elevations in nicotine responding on days 13, 14, and 15 of vapor exposure, * p<0.02 relative to non-dependent controls. Also shown are [B] active and [C] inactive levers by nicotine vapor-exposed (black circles) and air-exposed (white circles) rats over days, expressed as percent of baseline (3-day mean) prior to the start of vapor exposure. Change from baseline active lever responding by individual rats confirms raw data and statistical analyses. Nicotine vapor-exposed rats exhibited no change from baseline inactive lever responding, nor did non- dependent controls with the exception of a slight increase on days 6 & 7 of the protocol, confirming that the difference between groups in inactive lever responding was due to baseline differences after rats were split into groups matched for active lever responding. * p<0.05 relative to non-dependent controls.

Figure 2

Mean ± SEM somatic withdrawal scores by nicotine vapor-exposed (black bars) and air-exposed (white bars) rats injected with saline (vapor day 13) or mecamylamine (1.5 mg/kg, vapor day 14) at ^~8 hrs withdrawal from nicotine vapor. Scores from the 10-min observation period represent a summation of counts for behavioral signs that include blinks, gasps, writhes, head shakes, ptosis, teeth chattering, and yawns (Malin et al., 1992). Nicotine vapor-exposed rats exhibited higher scores overall than controls (p<0.05), and nicotine vapor-exposed rats injected with mecamylamine exhibited robust withdrawal relative to both saline injections and nicotine-naïve rats. * p<0.01 relative to non-dependent controls, # p<0.01 relative to saline injection.

Figure 3

Air-nicotine concentrations expressed as parts per million (ppm; left y-axis) and mg/m³ (right y-axis). Air-Nicotine concentrations are shown as a function of nicotine flow rate, expressed in liter per minute (LPM). There was a strong positive correlation between nicotine flow rate and air-nicotine concentrations (r²=0.99). Rats in Experiments 1 and 2 were exposed to a nicotine flow rate of 5 LPM, which produces air- nicotine concentrations similar to those experienced by human smokers.