Therapeutic hypothermia promotes protection after traumatic brain injury (TBI). The mechanisms underlying hypothermic protection are multifactorial and may include the modulation of microRNA (miRNA) expression after trauma. We utilized microarrays to examine the effects of posttraumatic hypothermia on the expression of 388 rat miRNAs. Animals were subjected to sham or moderate fluid percussion brain injury, followed by 4 hours of hypothermia (33°C) or normothermia (37°C) and euthanized at 7 or 24 hours. At 7 hours, 47 miRNAs were significantly different (P<0.05) between TBI and sham (15 higher in TBI and 31 lower). After 24 hours, 15 miRNAs differed by P<0.05 (7 higher and 9 lower). The expression of miRNAs was altered by posttraumatic hypothermia. At 7 hours, seven were higher in hypothermia than normothermia and five were lower. Some miRNAs (e.g., miR-874 and miR-451) showed the most difference with hypothermia, with changes verified by quantitative reverse transcriptase-PCR. Regionally specific miRNAs also showed responses to TBI and hypothermia treatments by in situ hybridization. In addition, in vitro neuronal stretch injury studies showed similar temperature-sensitive responses to specific miRNAs. These novel data indicate that the reported beneficial effects of early hypothermia on traumatic outcome may include temperature-sensitive miRNAs involved in basic cell-processing events.