Background Hibernating American black bears have significantly different clotting parameters than their summer active counterparts, affording them protection against venous thromboembolism during prolonged periods of immobility. We sought to evaluate if significant differences exist between the expression of microRNAs in the plasma of hibernating black bears compared with their summer active counterparts, potentially contributing to differences in hemostasis during hibernation. Materials and methods MicroRNA sequencing was assessed in plasma from 21 American black bears in summer active (n = 11) and hibernating states (n = 10), and microRNA signatures during hibernating and active state were established using both bear and human genome. MicroRNA targets were predicted using messenger RNA (mRNA) transcripts from black bear kidney cells. In vitro studies were performed to confirm the relationship between identified microRNAs and mRNA expression, using artificial microRNA and human liver cells. Results Using the bear genome, we identified 15 microRNAs differentially expressed in the plasma of hibernating black bears. Of these microRNAs, three were significantly downregulated (miR-141-3p, miR-200a-3p, and miR-200c-3p), were predicted to target SERPINC1, the gene for antithrombin, and demonstrated regulatory control of the gene mRNA expression in cell studies. Conclusions Our findings suggest that the hibernating black bears’ ability to maintain hemostasis and achieve protection from venous thromboembolism during prolonged periods of immobility may be due to changes in microRNA signatures and possible upregulation of antithrombin expression.