A buprenorphine-validated rat model of opioid use disorder optimized to study sex differences in vulnerability to relapse

Abstract

Rationale: Opioid use disorder (OUD) is a major epidemic in the USA. Despite evidence indicating that OUD may be particularly severe for women, preclinical models have yet to establish sex as a major factor in OUD.

Objectives: Here, we examined sex differences in vulnerability to relapse following intermittent access fentanyl self-administration and protracted abstinence and used buprenorphine, the FDA-approved treatment for OUD, to test the validity of our model.

Methods: Following acquisition of fentanyl self-administration under one of two training conditions, male and female rats were given extended, 24-h/day access to fentanyl (0.25 μg/kg/infusion, 10 days) using an intermittent access procedure. Vulnerability to relapse was assessed using an extinction/cue-induced reinstatement procedure following 14 days of abstinence; buprenorphine (0 or 3 mg/kg/day) was administered throughout abstinence.

Results: Levels of drug-seeking were high following extended-access fentanyl self-administration and abstinence; buprenorphine markedly decreased drug-seeking supporting the validity of our relapse model. Females self-administered more fentanyl and responded at higher levels during subsequent extinction testing. Buprenorphine was effective in both sexes and eliminated sex and estrous phase differences in drug-seeking. Interestingly, the inclusion of a time-out during training had a major impact on later fentanyl self-administration in females, but not males, indicating that the initial exposure conditions can persistently impact vulnerability in females.

Conclusions: These findings demonstrate the utility of this rat model for determining sex and hormonal influences on the development and treatment of OUD.

Keywords: Addiction-like phenotype; Buprenorphine; Fentanyl; Intermittent access; Opioid; Self-administration; Sex differences; Treatment.

Fig. 1

Summary of experimental events. Male and female rats were trained to self-administer (SA) fentanyl without (noTO) or with a time-out (TO) after each infusion. Following acquisition of fentanyl SA (5 consecutive sessions wherein all 40 infusions available were obtained), rats were given extended 24-h/day, intermittent access (IntA) to fentanyl (0.25 μg/kg/infusion) for 10 sessions. No time-outs occurred after infusions during this phase of the study. Following the last IntA fentanyl SA session, rats were implanted with a subcutaneous minipump to deliver either buprenorphine (3 mg/kg/day) or control (vehicle or sham implant). Following a 14-day abstinence period, vulnerability to relapse was assessed in a single session using an extinction/cue-induced reinstatement procedure. The following morning, blood was collected to assess buprenorphine levels in plasma and estradiol/progesterone levels (females) in serum

Fig. 2

Effect of sex and timeout training condition on rates of acquisition and active and inactive lever responding during the 5-day acquisition criterion period. Mean (± SEM) number of days to acquire fentanyl self-administration (a) and the mean (± SEM) number of active (b) and inactive (c) lever responses for each of the five acquisition criterion sessions for males (M) and females (F) in the no time-out (noTO, n = 21 and 24, respectively) and time-out (TO, n = 20 and 30, respectively) training conditions. *Significant effect of time-out training condition over the 5 acquisition sessions (b) and during the last 2 sessions (c); #Significant effect of sex over the 5 sessions (b)

Fig. 3

Effect of sex and time-out training condition on subsequent fentanyl intake under extended, intermittent access conditions. Mean (± SEM) number of fentanyl infusions obtained during each of the 10 intermittent access sessions for males (M) and females (F) in the no time-out (noTO, n = 21 and 24, respectively) and time-out (TO, n = 20 and 30, respectively) training groups. *Significant difference between females in the noTO group and all other groups

Fig. 4

Effects of sex and time-out training condition on patterns of fentanyl self-administration under extended, intermittent access conditions. (a) Mean (± SEM) percent of total fentanyl intake that occurred in the dark phase for each of the 10 intermittent access sessions and the mean (± SEM) number of fentanyl infusions obtained during each hour of access averaged access sessions 1–2 (b) and 6–10 (c) of the intermittent access period for males (M) and females (F) in the no time-out (noTO, n = 21 and 24, respectively) and time-out (TO, n = 20 and 30, respectively) training groups. *Significant effect of light-dark phase: greater fentanyl intake in the dark versus light for sessions 1 and 2 (a, b) and lower intake in the dark versus light for sessions 6–10 (a, c). +Significant decrease in average percent intake in the dark phase from sessions 1–2 versus 6–10 (a). #Significantly lower fentanyl intake in males compared to the females in the dark phase for sessions 6–10 (a, c)

Fig. 5

Effect of sex, time-out training condition, and buprenorphine treatment on fentanyl-seeking during extinction testing. Mean (± SEM) number of responses made on the lever formerly associated with fentanyl during the first six 1-h extinction sessions (a, b) and across all extinction sessions run (c, d) for control treated (a, c) male (M) and female (F) rats in the no time-out (noTO, n = 11 and 12, respectively) and time-out (TO, n = 10 and 15, respectively) training groups and for buprenorphine-treated (b, d) male and female rats in the no time-out (n = 10 and 12, respectively) and time-out (n = 10 and 15, respectively) training groups. *Significantly higher responses during the first extinction session versus later ones (2–6) across the control and buprenorphine groups (a). #Significant overall effect of sex (a–d). +Significant overall effect of buprenorphine (c, d) and within the first 3 sessions (a, b)

Fig. 6

Effect of sex, time-out training condition, and buprenorphine treatment on fentanyl-seeking during cue-induced reinstatement testing. Mean (± SEM) number of responses made on the lever formerly associated with fentanyl during the last extinction session versus the reinstatement session for control treated (a) male (M) and female (F) rats in the no time-out (noTO, n = 11 and 12, respectively) and time-out (TO, n = 10 and 15, respectively) training groups and for buprenorphine-treated (b) male and female rats in the no time-out (n = 10 and 12, respectively) and time-out (n = 10 and 15, respectively) training groups. *Significant difference from the last extinction session (a, b). +Significant effect of buprenorphine (a, b)

Fig. 7

Effect of sex, time-out training condition, and buprenorphine treatment on body weight (g) during the abstinence/treatment and relapse testing phases of the study. Mean (± SEM) body weight (g) during early (Early, days 1–3), intermediate (Inter, days 6–8), and late (Late, days 12–14) abstinence as well as on the day of the relapse test (Relapse) for control treated (a) male (M) and female (F) rats in the no time-out (noTO, n = 11 and 12, respectively) and time-out (noTO, n = 10 and 15, respectively) training groups and for buprenorphine-treated (b) male and female rats in the no time-out (n = 10 and 12, respectively) and time-out (n = 10 and 15, respectively) training groups. #Significant effect of sex overall and at each of the time-points during abstinence/treatment and relapse testing (a, b). *Significant difference from Early (a). +Significant effect of buprenorphine (a, b)

Fig. 8

Effect of estrous cycle phase and buprenorphine treatment on fentanyl-seeking during extinction and reinstatement testing. a Mean total (± SEM) responses made on the lever formerly associated with fentanyl across all extinction sessions run (6–8) and b during the reinstatement test session for control (left panels) and buprenorphine-treated (BUP; right panels) females tested during estrus (Estrus; n = 11 and 12, respectively) versus non-estrus (Non-E; n = 16 and 15, respectively) phases. #Significant overall effect of estrous cycle phase (a, b). +Significant overall effect of buprenorphine (a, b) and for both control estrus and non-estrus rats as compared to buprenorphine rats (collapsed across estrous cycle phase; b)