Hypertonic saline (HS) resuscitation is a promising new approach in the prevention of tissue damage in trauma patients, because of its capacity to block neutrophil (PMN) activation. We have studied the molecular mechanisms through which HS controls PMN function and have found that HS triggers the release of ATP. ATP can augment PMN responses via P2 receptors. ATP converted to adenosine inhibits PMN activation via P1 receptors. We hypothesize that the balance between ATP and adenosine determines whether HS increases or dampens PMN function. In the following three Specific Aims, we propose to define the components that control this ATP/adenosine balance and determine pharmaceutical approaches to shift it toward adenosine, and thus toward inhibition of PMN function.
Specific Aim 1 : ATP release and P2 receptor activation: The mechanisms of ATP release will be studied, a novel method for real-time monitoring of ATP release will be established, and P2 receptor subtype expression and colocalization with sites of ATP release and hydrolysis will be assessed.
Specific Aim 2) Adenosine formation and P1 receptor activation: The dynamics of expression and colocalization of ecto-enzymes generating and hydrolyzing adenosine, specifically alkaline phosphatase, adenosine deaminase, and of the P1 receptors that are activated by adenosine will be studied.
Specific Aim 3) Modulation of the effect ofHS: The effect of ATP released from other cells surrounding PMN will be studied and pharmacological approaches to improve the efficacy of HS resuscitation by modulating adenosine accumulation will be tested with human whole blood and a mouse model of hemorrhagic shock. The proposed experiments will elucidate general principles of HS-induced ATP release and the control of PMN function by extracellular ATP and its products. This knowledge will allow us to better understand how HS resuscitation can regulate PMN function and how the clinical effectiveness of HS can be optimized to attenuate inflammation and PMN-induced organ damage in trauma victims.
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