Receptor for activated protein kinase C 1 (RACK1) is a multifaceted scaffolding protein known to be involved in the regulation of signaling events required for neuronal protection. In the present study, we investigated the role of RACK1 in secondary brain injury in a rat traumatic brain injury (TBI) model. A weight-drop TBI model was established in Sprague Dawley rats, and RACK1 in vivo knockdown and overexpression were performed 24h before TBI insult. The IRE1 inhibitor 3,5-dibromosalicylaldehyde (DBSA) was administered by intracerebroventricular injection 1h after TBI insult. Real-time PCR, Western blotting, immunofluorescence, neuronal apoptosis, brain water content, and neurological scores were evaluated. Our results revealed that TBI induced increased expression of endogenous RACK1, phosphorylated inositol-requiring enzyme 1 (p-IRE1), X-box binding protein-1 (XBP1) and glucose-regulated protein 78 (GRP78) in neurons. RACK1 overexpression significantly ameliorated neuronal apoptosis, blood-brain barrier disruption, brain edema and neurological deficits at 48h after TBI, which was concomitant with upregulation of p-IRE1, XBP1 and GRP78 expression, while its knockdown induced the opposite effects. Furthermore, DBSA administration reversed the protective effects of RACK1 overexpression against brain injury and decreased the expression of p-IRE1, XBP1 and GRP78. In summary, the upregulation of RACK1 following brain contusion exerted neuroprotective effects against secondary brain injury, which were probably mediated by activation of the IRE1-XBP1 pathway.