Static contraction of skeletal muscle evokes reflex increases in blood pressure and heart rate. Previous studies showed that P2X receptors located at the dorsal horn of the spinal cord play a role in modulating the muscle pressor reflex. P2X stimulation can alter release of the excitatory amino acid, glutamate (Glu). In this report, we tested the hypothesis that stimulation of P2X receptors enhances the concentrations of Glu ([Glu]) in the dorsal horn, and that blocking P2X receptors attenuates contraction-induced Glu increases and the resultant reflex pressor response. Contraction was elicited by electrical stimulation of the L7 and S1 ventral roots of fourteen cats. Glu samples were collected from microdialysis probes inserted in the L7 level of the dorsal horn of the spinal cord, and dialysate [Glu] was determined using the HPLC method. First, microdialyzing ,-methylene ATP (0.4 mM) into the dorsal horn significantly increased [Glu]. In addition, contraction elevated [Glu] from baseline of 536±53 nM to 1179±192 nM (P<0.05, vs. baseline), and mean arterial pressure (MAP) by 39±8 mmHg in control experiment. Microdialyzing the P2X receptor antagonist pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid (PPADS; 10 mM) into the dorsal horn attenuated the contraction induced-Glu increase (610±128 nM to 759±147; P>0.05) and pressor response (16±3 mmHg P<0.05 vs. control). Our findings demonstrate that P2X modulates the cardiovascular responses to static muscle contraction by affecting the release of Glu in the dorsal horn of the spinal cord.