Fentanyl exposure during preconception and gestation permanently dysregulates endogenous opioid peptides and sympathoadrenal-medullary axis in the offspring

Abstract

In the United States, the alarming increase in opioid use disorder diagnoses during pregnancy in the last decade has increased the incidence of neonatal opioid withdrawal syndrome (NOWS). Although 8 per 1,000 newborns are diagnosed with NOWS each year, the lack of prospective studies is a roadblock in the development of approaches to reduce adverse health outcomes in this vulnerable population. This study used a preclinical model to assess short- and long-term effects of preconceptional and gestational fentanyl (FEN) exposure on morphometrics, hormonal plasma profile, and sensitivity to opioid re-exposure in the offspring. Sprague Dawley female rats self-administered FEN citrate [fixed-ratio 1 (FR1), 2.5 μg/kg] or vehicle (NaCl 0.9%) during preconception and until gestational day 21. In utero fentanyl exposure (IUFE) did not influence neonatal weight and morphometrics; however, IUFE pups exhibited a higher frequency of behaviors indicative of somatic withdrawal compared with controls (CTLs). In male and female weanlings, IUFE induced the dysregulation of endogenous opioid peptides (EOPs) and increased metanephrine levels compared with CTL counterparts. However, only adult females with IUFE showed increased EOPs and metanephrine levels, FEN-induced hyperalgesia, and greater FEN-induced hypotensive and bradycardic effects compared with CTL counterparts. This preclinical model suggests a long-lasting association between IUFE-induced neuroendocrine dysregulation and adverse effects of opioid re-exposure in female offspring.

Keywords: endogenous opioid peptides; opioid withdrawal; preconception; prenatal fentanyl exposure; self-administration.

Figure 1. Fentanyl self-administration in pregnant rats

(A) timeline of events involving dams and their offspring, (B) fentanyl consumption in female rats self-administering VEH or FEN during preconception and gestation, (C) discrimination percentage of active versus inactive lever presses in female rats self-administering vehicle or fentanyl; the horizontal dotted line represents 66.6% active lever press discrimination passing criteria during preconception and gestation, and (D) weight gain projection during preconception and gestation. Data were collected, analyzed, averaged per group, and reported as mean ± SEM; n=3 dams per group.µg=microgram; kg=kilogram; g=gram. *P<0.05 versus control. FEN, fentanyl; SA, self-administration; VEH, vehicle; D, gestational day.

Figure 2. Pregnancy outcomes in dams self-administering fentanyl or vehicle during pre-conception and pregnancy

(A) birth weight and morphometric analysis in male offspring, (B) birth weight and morphometric analysis in female offspring, (C) cephalization index and body weight correlation in male offspring, (D) cephalization index and body weight correlation in female offspring, (E) body weight gain trajectory and tibia length in adult male offspring, and (F) body weight gain trajectory and tibia length in adult female offspring. Data were collected, analyzed, averaged per group, and reported as mean ± SEM; n=22 males and 18 females per group. BL, body length; BW, body weight; CTL, control, offspring from dams self-administering saline; FL, full body length; GD, anogenital distance; HC, head circumference; IUFE, in utero fentanyl exposure, offspring from dams self-administering fentanyl; TL, tail length; WC, waist circumference.

Figure 3. Somatic signs of withdrawal in IUFE and CTL offspring at PD0, 1, and 3

(A) GWS in male offspring, (B) GWS in female offspring, (C) tremors in male offspring, (D) tremors in female offspring, (E) SMJ-like movements in male offspring, (F) SMJ-like movements in female offspring. Data were collected, analyzed, averaged per group, and reported as mean ± SEM; n=22 males and 18 females per group. *P<0.05 versus CTL. CTL, control, offspring from dams self-administering saline; GWS, global withdrawal score; IUFE, in utero fentanyl exposure, offspring from dams self-administering fentanyl; PD, postnatal day; SMJ, spontaneous myotonic jerks. *P<0.05 versus control.

Figure 4. Plasma endogenous opioid peptides and adrenal-derived hormones in adult male and female CTL and IUFE offspring

Plasma levels of (A) met-enkephalin, (B) β-endorphin, (C) dynorphin A, (D) ACTH, (E) corticosterone, (F) aldosterone, (G) metanephrine, and (H) norepinephrine. Data were collected, analyzed, averaged per group, and reported as mean ± SEM; n=6 females per group. *P<0.05 versus CTL. µg=microgram; kg=kilogram; g=gram. CTL, control, offspring from dams self-administering saline; FEN, fentanyl; IUFE, in utero fentanyl exposure, offspring from dams self-administering fentanyl; SA, self-administration; VEH, vehicle.

Figure 5. Nociception and plasma fentanyl exposure in adult CTL and IUFE offspring

(A) hot plate test in weanling and adult female offspring, (B) progressive hot plate test in response to a 20 χg/kg dose of fentanyl, (C) pharmacokinetics of fentanyl in response to a 20 µg/kg dose, i.p. Data were collected, analyzed, averaged per group, and reported as mean ± SEM; n=6 females per group. *P<0.05 versus CTL. CTL, control, offspring from dams self-administering saline; HP, hot plate test; IUFE, in utero fentanyl exposure, offspring from dams self-administering fentanyl.

Figure 6. Cardiovascular response of adult female IUFE and CTL offspring to different fentanyl doses

(A) MAP in response to 20 μg/kg of fentanyl, (B) HR in response to 20 μg/kg of fentanyl, (C) MAP in response to 200 μg/kg of fentanyl, (D) HR in response to 200 μg/kg of fentanyl. Data were collected, analyzed, averaged per group, and reported as mean ± SEM; n=6 females per group. *P<0.05 versus CTL. CTL, control, offspring from dams self-administering saline; HR, heart rate; IUFE, in utero fentanyl exposure, offspring from dams self-administering fentanyl; MAP, mean arterial pressure.