Administration of drugs of abuse results in the induction of interoceptive (i.e., internally perceived) states that are subject to basic learning processes related to substance use disorders (Thompson et al., 2019; Wise and Kiyatkin, 2011). For example, interoceptive stimuli induced by administration of cocaine and morphine can be respectively conditioned to modulate relapse-like behaviour (Wise et al., 2008; Keiflin et al., 2008) and tolerance and withdrawal (Kim et al., 1999; McDonald and Siegel, 2004). Interoceptive stimuli induced by drug administration are also amenable to hierarchical learning structures such as occasion setting (Bevins and Besheer, 2014; Bevins and Murray, 2011; Bevins and Palmatier, 2004). Occasion setters (OSs) are stimuli on which the relationship between a conditioned stimulus (CS) and associated unconditioned stimuli (US) are contingent; a positive-feature (FP) OS is predictive that a CS-US relationship is in effect, whereas a negative-feature (FN) OS is predictive that a CS-US relationship is not in effect (Fraser and Holland, 2019). Morphine-evoked interoceptive stimuli function as FP and FN OSs for the induction of goal tracking (i.e., conditioned approach to a location of reward delivery) (Andrade et al., 2021) and conditioned aversion (Huang et al., 2023; Martin et al., 1990; Mukherjee et al., 2023; Skinner et al., 1995). Given OSs gate the induction of reinstatement of drug seeking (Chaudhri et al., 2008) and the expression of extinction (Kearns and Weiss, 2007), sensitivity to behaviours evoked by interoceptive drug stimuli conditioned as OSs may be related to relapse. Therefore, an understanding of the factors underlying this property may aid in the development of relapse prevention strategies for OUD.

We previously reported that female and male rats were differentially sensitive to generalization of the FP morphine OS to other doses of morphine (Andrade et al., 2021), but the mechanisms underlying this difference have not been studied. In humans, the magnitude of morphine analgesia in women is smaller in the luteal phase compared with the follicular phase and compared with men (Ribeiro-Dasilva et al., 2011), suggesting that some sex differences in sensitivity to opioids are mediated by activational effects of gonadal hormones in females. Animal literature is consistent with this idea in that morphine analgesia is blunted during estrus (Stoffel et al., 2003) and proestrus (Berglund and Simpkins, 1988), enhanced by adult ovariectomy (OVX) (Krzanowska and Bodnar, 1999), and inhibited in adult OVX rats by acute treatment with 17ß-estradiol (E2) alone or in combination with progesterone (Ratka and Simpkins, 1991). These effects may be general to a variety of opioid-related behaviours because heroin self-administration is also inhibited by E2 treatment in adult OVX rats (Smith et al., 2022, Smith et al., 2021). One possible mechanism underlying our previous finding of blunted generalization of a FP morphine OS in female rats relative to males is thus inhibition of the pharmacological effects of morphine by gonadal hormones in females. Given this possibility, we trained adult male and female rats to discriminate a morphine stimulus as an interoceptive FP OS from saline and then compared generalization of this stimulus to a range of doses of morphine while monitoring estrous phase. We hypothesized that if gonadal hormones affect morphine generalization, then this behaviour would be less pronounced in proestrous and estrous females relative to metestrous and diestrous females and males.