Role of skeletal muscle vasodilatory pathway in adaptive thermogenesis in Sunit Mongolian sheep

The Sunite Mongolian sheep is one of the superior livestock breeds in Inner Mongolia, capable of surviving and reproducing on the Xilingol Grassland at temperatures below -30 °C, demonstrating remarkable cold adaptation. However, research on adaptive thermogenesis in its muscle tissue remains limited. To investigate skeletal muscle's role in adaptive thermogenesis, shoulder muscle tissues were collected from three healthy castrated male sheep each in winter (January) and summer (July) for RNA sequencing (RNA-seq) and differential expression analysis. Differentially expressed genes (DEGs) were primarily enriched in pathways associated with starvation response, hormonal regulation, and energy metabolism. Gene expression analysis revealed significant upregulation of nitric oxide synthase 3 (NOS3) and adrenergic receptor-β2 (ADRB2) in the adrenergic-cyclic guanosine monophosphate (cGMP)- protein kinase G(PKG)vasodilatory signaling pathway during winter (P < 0.01). Further validation showed that ADRB2 was extensively expressed in skeletal muscle vascular endothelial cells, with significantly increased winter expression. Subsequent in vivo experiments confirmed these results, intravenous injection of isoproterenol (ISO) dose-dependently increased cGMP concentration and NOS3 expression (P < 0.05). In summary, we speculate that epinephrine binds to ADRB2 in the skeletal muscle of Sunite Mongolian sheep during winter, activating intracellular NOS3 to synthesize nitric oxide (NO). Through the cGMP-PKG signaling pathway, this induces vasodilation, thereby transporting more nutrients and enhancing metabolic energy production and heat generation, primarily via glucose metabolism, to adapt to cold environments. This study provides new insights into cold adaptation in sheep, suggesting ADRB2/NOS3 as potential molecular markers for breeding cold-resistant breeds to enhance farming efficiency.