Activation of the adrenal gland stress response is of utmost importance to survive sepsis. Experimental and clinical evidence exists demonstrating that adrenal gland often develops functional and structural damage due to sepsis with mechanisms remaining largely unknown. In the present study, we have used RNA Sequencing (RNA-Seq) technology to analyze changes in adrenal transcriptome elucidated by bacterial LPS. We aimed to find particularly alterations in genes that were previously not reported to be involved in the adrenal gland dysregulation in contexts of sepsis. Our results demonstrate that systemic administration of LPS significantly altered expression of 8458 genes as compared to saline injected animals. The subsequent quality and functional analysis of these gene signatures revealed that LPS-induced highly homogenous transcriptional response in total upregulating 4312 and downregulating 4146 genes. Furthermore, functional annotation analysis together with gene enrichment set analysis (GSEA) clearly demonstrated that adrenal response to LPS involved alterations in multiple pathways related to the inflammatory response along with previously unexplored activation of the hypoxia pathway. In addition, LPS strongly downregulated genes involved in the adrenal homeostasis, development, and regeneration. Those alterations were subsequently verified in clinically relevant cecal ligation and puncture (CLP)-induced sepsis model. Collectively, our study demonstrates that RNA-seq is a very useful method that can be applied to search for new unexplored pathways potentially involved in adrenal gland dysregulation during sepsis.