RNA Sequencing Reveals Transcriptional Changes in Muscle Tissue in Response to Muscle Pain in Exercised and Sedentary Mice

Abstract

Exercise produces analgesia through multiple peripheral, spinal, and supraspinal pathways. However, uncovering these pathways one by one is time consuming, biased based on prior literature, and requires significant number animals to be tested. RNA sequencing allows for discovery of numerous transcriptional differences and can lead to the discovery of novel nociceptive pathways. The purpose of this study was to measure transcriptional differences in response to muscle pain between sedentary and exercised animals. Male and female C57/Black6 mice were grouped as pain free, sedentary with muscle pain model, and exercised with muscle pain model. The muscle pain model was induced with 2 intramuscular injections of acidic saline (20μl, pH 5.0±0.1) combined with 6 minutes of fatiguing muscle contractions delivered to the left gastrocnemius muscle. For exercise, animals were individually housed with 24hr access to a running wheel for 8 weeks prior to induction of the pain model. The left gastrocnemius muscle was harvested 24hrs after induction of the pain model and RNA was extracted. Bulk RNA sequencing was performed on an Illumina NovaSeq 6000. Differentially expressed genes (DEGs) were generated based on raw counts at the gene and transcript level using DESeq2. Within each sex, DEGs were determined between the pain free and sedentary groups and between exercised and sedentary with pain model groups. DEGs were then imported into iPathwayGuide commercial software and results were compared between sexes. Between pain free and sedentary in pain animals 160 DEGs were discovered in males and 661 were discovered in females. Between exercised and sedentary groups with the muscle pain model, 13 DEGs were discovered in males and 195 in females. iPathwayGuide analysis for all data sets revealed activation of immune system pathways including chemokine signaling, cytokine-cytokine receptor interaction, and antigen processing and presentation. Future work will confirm results with qPCR and behavioral pharmacology. Grant support from NIH AR073187.