Hypertonic saline (HS) holds promise as a novel resuscitation fluid for the treatment of trauma patients because HS inhibits neutrophil (PMN) activation and thereby prevents host tissue damage and associated post-traumatic complications. However, depending on conditions of cell activation, HS can increase PMN degranulation, which could exacerbate tissue damage in trauma victims. The cellular mechanism by which HS increases degranulation is unknown. Here we test if HS-induced ATP release from PMN and feedback via P1 and/or P2 receptors may be involved in the enhancement of degranulation by HS. We found that HS enhances elastase release and ERK and p38 MAPK activation when HS is added after activation of PMN with formyl peptide (fMLP) or phorbol ester (PMA). Agonists of P2 nucleotide and A3 adenosine receptors mimicked these enhancing effects of HS, while antagonists of A3 receptors or removal of extracellular ATP with apyrase diminished the response to HS. A1 adenosine receptor antagonists increased the enhancing effect of HS while A1 receptor agonists inhibited elastase release. These data suggest that HS up-regulates degranulation via ATP release and positive feedback through P2 and A3 receptors. We proposed that these feedback mechanisms could serve as potential pharmacological targets to fine-tune the clinical effectiveness of HS resuscitation.