Piloting for Acute Methadone Withdrawal and Sociability Consequences of NOWS

          Opioids such as morphine and methadone have been actively prescribed for years to treat chronic pain; however, the overprescribing of synthetic opioids in the last two decades has led to what is now known as the “opioid epidemic.” While these medications are often successful for treating pain, they also produce euphoric and reward-seeking behaviors that increase the possibility of misuse and abuse, now termed opioid use disorder (OUD). While the cause of developing dependence and decreasing tolerance is not yet fully understood, it has been hypothesized that down-regulation of the µ-opioid receptors is a factor. Acute withdrawal occurs during the detoxification process, characterized by somatic symptoms such as body tremors, flu-like symptoms, muscle aches, and insomnia. Other signs of negative affect include irritability and anxiety. The use of rodent models for opioid withdrawal has shown many similarities to the symptoms exhibited in humans. Further use of rodent models in opioid withdrawal may give more insight into the human physical and psychological dependence on opioids and the prevention of relapses. Methadone, in particular, is often regarded as safe to use during pregnancy. Analyzing the effects in rats recovering from in utero and early postnatal methadone withdrawal gives us a better indication of the adverse effects due to the mother taking methadone during pregnancy and the outcome of neonatal opioid withdrawal syndrome (NOWS). Our experiment aims to assess the baseline behavior of neonatal rats through a small series of designed experiments and determine any sex differences before the implementation of methadone. Such experiments include ultrasonic vocalization (USV) detection, determination of a withdrawal score, and an assessment of behavior within an enriched environment.

• Dams had ad libitum access to food and water 

• From postnatal days (PNDs) 14-18, 24 rats were recorded vocally and visually for 10 minutes using an ultasound vocalization (USV) detector (Med-Associates, Inc.) and a Sony digital camera. Each day, a subset of the rats were tested such that each rat was only tested once.

• The total number of wall climbs and jumps were scored. 

• The total number of USVs emitted in the 20-50kHz and 50-70kHz ranges were recorded.  

• On PND 21, litters were placed into enriched housing (n=4-6 per cage). 

• Cages were refreshed every three days with novel objects to maintain enrichment. Visual recordings were conducted after every other cage change for 30 minute periods during both the day and night cycle.

• Behavioral observations were then scored for:

  • Time spent in social contact (e.g., sniffing, grooming)

  • Interaction with the environment (e.g., running in a wheel)

  • Time spent in non-social contact (e.g., self-grooming)

  • Frequency of pins and nape attacks

The enrichment behavioral analysis is yet to be completed; however, from the USV data we saw an age-related main effect when assessing USV emissions, suggesting younger pups are more vocal when separated from mom. 

Future studies include replicating the experiments conducted with the administration of methadone. 

This would quantify the impairments associated with NOWS in animal models, and evaluate if enriched and stimulating environments can mitigate the social deficits observed in affected individuals.

Dunn, A. D., Robinson, S. A., Nwokafor, C., Estill, M., Ferrante, J., Shen, L., Lemchi, C. O., Creus-Muncunill, J., Ramirez, A., Mengaziol, J., Brynildsen, J. K., Leggas, M., Horn, J., Ehrlich, M. E., & Blendy, J. A. (2023).  Molecular and long-term behavioral consequences of neonatal opioid exposure and withdrawal in mice. Frontiers in Behavioral Neuroscience, 17, 1202099. https://doi.org/10.3389/fnbeh.2023.1202099

Ozdemir, D., Allain, F., Kieffer, B. L., & Darcq, E. (2023). Advances in the characterization of negative affect caused by acute and protracted opioid withdrawal using animal models. Neuropharmacology, 232, 109524. https://doi.org/10.1016/j.neuropharm.2023.109524 

The rats were bred at Miami University and housed in the LAR facility, in compliance with The Guide for the Care and Use of Laboratory Animals (8th Edition, National Academies Press, Washington, D.C., 2011) and approved by Miami University’s Institutional Animal Care and Use Committee.