A RAT MODEL OF HUMAN FENTANYL ABUSE

Abstract Background Fentanyl is a synthetic opioid that is widely used in anesthesiology, but its illicit use is rapidly increasing in the United States, contributing to >70,000 overdose deaths in 2021. In addition to dose-dependent respiratory depression, at high doses fentanyl induces unconsciousness and muscle rigidity, the mechanisms of which are poorly understood. Animal models are urgently needed to study these effects, especially in the context of repeated exposures that are common in individuals with opioid use disorder (OUD). Aims & Objectives The aim of this study was to investigate the effects of repeated high-dose fentanyl injections on loss of righting reflex, heart rate, muscle activity, and respiratory gas exchange to establish a rat model of human fentanyl abuse, and to assess how tolerance develops for these different endpoints. Methods Male and female Sprague-Dawley rats were studied (n=42). We first assessed the dose- response relationship for fentanyl-induced unconsciousness by rapidly injecting 12.5, 25, 50, or 100 & micro;g/kg IV, and recording the time to return of righting reflex (RORR). We administered 100 & micro;g/kg IV as the standard dose for the remainder of the experiments, and injected this dose twice a week, with at least 72 hours in between exposures, for five consecutive weeks. We evaluated time to RORR using video recording. In a separate cohort of rats, we implanted electrocardiogram (ECG) and electromyogram (EMG) electrodes. Finally, we conducted experiments with a cohort of rats with preimplanted arterial catheters for arterial blood gas (ABG) analysis at 5, 15, and 60 minutes after fentanyl injection. We used mixed-effect models, effect-size measures, and Friedman’ s test for statistic analysis. Data are presented as median [interquartile range]. Results In total, 39 animals received 100 & micro;g/kg IV as the first dose, of which five died. The dose- response experiments demonstrated a significant prolongation of time to RORR with increasing dose (median time to RORR after 100 & micro;g/kg was 49.5 [7.8] min, n=4). Repeated fentanyl injections resulted in a significant decrease in time to RORR when comparing the first (45.6 [41.3] min] and fifth injections (15.0 [6.4] min, p=0.0012), but not the first and tenth injections (34.8 [34.5] min, n=6). Fentanyl induced significant bradycardia, and recovery of the heart rate was significantly faster after the fifth and tenth injection when compared to the first injection (n=7-9). EMG amplitude increased significantly after fentanyl injection compared to awake/resting activity. Repeated fentanyl exposure resulted in significantly increased rigidity when comparing the first and tenth injections. ABG analysis revealed that arterial pH and pO2 were significantly decreased, and pCO2 was significantly increased at 5 and 15 min after the first, fifth, and tenth injections compared to baseline. Discussion & Conclusion In this study, we established a robust rodent model of human fentanyl abuse, with repeated, rapid administration of high doses that recapitulate the life-threatening effects of fentanyl overdose including unconsciousness, muscle rigidity, and respiratory depression. Ten injections of high-dose fentanyl (100 & micro;g/kg) do not induce tolerance to most these effects. Some side effects may be even more pronounced after repeated injections.