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Repetitive neonatal pain increases spinal cord DNA methylation of the µ-opioid receptor

Pediatric Research volume 98, pages 1149–1154 (2025)Cite this article

Abstract

Background

Repetitive neonatal painful procedures experienced in the neonatal intensive care unit (NICU) are known to alter the development of the nociceptive system and have long-lasting consequences. Recent evidence indicates that NICU stay affects the methylation of the opioid receptor mu 1 encoding gene (Mor-1). Additionally, a preclinical model of neonatal procedural pain established lower adult post-operative MOR-1 levels in the spinal cord. Thus, we hypothesized that neonatal procedural pain increases the DNA methylation status of Mor-1 in the spinal cord and dorsal root ganglia (DRGs).

Methods

To this end, repetitive neonatal procedural pain was induced in animals, during the first postnatal week, a period equivalent to preterm human brain development. On postnatal day 10 methylation of Mor-1 promotor was assessed in the spinal cord and the DRG using bisulfite pyrosequencing.

Results

Our findings demonstrated that neonatal procedural pain increased spinal cord Mor-1 promotor DNA methylation in the ipsilateral side as compared to the contralateral side, an effect that was not observed in the control animals, nor in the DRG.

Conclusion

This study is the first to highlight a localized and noxious-stimuli-dependent effect of repetitive neonatal procedural pain on Mor-1 promotor methylation and emphasizes the need to explore the effects of repetitive neonatal procedural pain on the epigenome.

Impact

  • This study reveals that repetitive neonatal procedural pain is associated with increased DNA methylation of the Mor-1 promoter in the spinal cord of neonatal rats.

  • This is the first study to identify an effect of neonatal procedural pain on DNA methylation, emphasizing the critical need for further investigation into the epigenetic consequences of neonatal procedural pain.

  • These insights could lead to better management and treatment strategies to mitigate the long-term impacts of early pain exposure on neurodevelopment and behavior.

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Fig. 1: Experimental design and timeline.
Fig. 2: Mechanical sensitivity after needle prick or handling during the neonatal week.
Fig. 3: Spinal cord methylation levels at P10 of the Mor-1 promotor.

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Data availability

The data supporting this study’s findings are available from the authors upon reasonable request.

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Acknowledgements

The completion of this research project would not have been possible without the scientific discussions with colleagues L. Heijmans (Ph.D.), T.J. de Geus (MSc), I. Rudnick-Jansen (Ph.D.), M. Mons (Ph.D.), A. Tiane (Ph.D.), S. Chenine, P. Koulousakis (Ph.D.), and members of the Neuroepigenetics group, Maastricht University, Maastricht, the Netherlands. Optimization of the bisulfite pyrosequencing assays would not have been possible without the help of T. Doeswijk and M. Naldi. We also would like to thank D. Hermes, S. Claessen, and members of the animal facility for their technical support and expertise.

Funding

Funding was provided by internal research support based on grants from Erasmus-Sophia Children’s Hospital Rotterdam, the Netherlands (to S. H.P. Simons) and Maastricht University, Maastricht, the Netherlands (to E.A.J. Joosten and D.L.A. van den Hove).

Author information

Authors and Affiliations

  1. Department of Anesthesiology and Pain Management, Maastricht University Medical Centre+, Maastricht, the Netherlands

    Mathilde Baudat & Elbert A. J. Joosten

  2. Department of Translational Neuroscience, Mental Health and Neuroscience Research Institute, Maastricht University, Maastricht, the Netherlands

    Mathilde Baudat & Elbert A. J. Joosten

  3. Department of Neonatal and Pediatric Intensive Care, Division of Neonatology, Erasmus University Medical Centre Rotterdam- Sophia Children Hospital, Rotterdam, the Netherlands

    Sinno H. P. Simons

  4. Department of Psychiatry and Neuropsychology, Mental Health and Neuroscience Research Institute, Maastricht University, Maastricht, the Netherlands

    Daniël L. A. van den Hove & Renzo J. M. Riemens

Authors
  1. Mathilde Baudat
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Contributions

All authors substantially contributed to the conception and design of the experiment, interpreted the data and critically revised the manuscript. Acquisition of data and drafting of the article was performed by M. Baudat. The final manuscript was approved by all authors.

Corresponding author

Correspondence to Mathilde Baudat.

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The authors declare no competing interests.

Ethical approvals

All animal experiments were performed in accordance with the European Directive for Protection of Vertebrate Animal Use for Experimental and Other Scientific Purposes (86/609/EEC) and were approved by the Committee for Experiments on Animals, Maastricht, The Netherlands (DEC 2017-017).

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Baudat, M., Joosten, E.A.J., Simons, S.H.P. et al. Repetitive neonatal pain increases spinal cord DNA methylation of the µ-opioid receptor. Pediatr Res 98, 1149–1154 (2025). https://doi.org/10.1038/s41390-025-03892-7

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