2--3--o-(2-4-6-trinitrophenyl)adenosine-5--triphosphate and 1-3-dipropyl-8-cyclopentylxanthine

Our electrophysiological studies have shown that both purinergic and glutamatergic receptors are involved in central sensitization of nociceptive neurons in the medullary dorsal horn (MDH). Here we assessed the effects of intrathecal administration of apyrase (a nucleotide degrading enzyme of endogenous adenosine 5-triphosphate [ATP]), a combination of apyrase and 1,3-dipropyl-8-cyclopentylxanthine (DPCPX, an adenosine A1 receptor antagonist), or 2,3-O-2,4,6-trinitrophenyl-adenosine triphosphate (TNP-ATP, a P2X1, P2X3, P2X2/3 receptor antagonist) on the release of glutamate in the rat MDH evoked by application of mustard oil (MO) to the molar tooth pulp. In vivo microdialysis was used to dialyse the MDH every 5 min, and included 3 basal samples, 6 samples after drug treatment and 12 samples following application of MO. Tooth pulp application of MO induced a significant increase in glutamate release in the MDH. Superfusion of apyrase or TNP-ATP alone significantly reduced the MO-induced glutamate release in the MDH, as compared to vehicle. Furthermore, the suppressive effects of apyrase on glutamate release were reduced by combining it with DPCPX. This study demonstrates that application of an inflammatory irritant to the tooth pulp induces glutamate release in the rat MDH in vivo that may be reduced by processes involving endogenous ATP and adenosine.

Topics: Adenosine; Adenosine Triphosphate; Animals; Apyrase; Central Nervous System Sensitization; Dental Pulp; Glutamic Acid; Irritants; Male; Microdialysis; Molar; Mustard Plant; Plant Oils; Posterior Horn Cells; Purinergic P2X Receptor Antagonists; Rats; Rats, Sprague-Dawley; Receptors, Purinergic P2X; Trigeminal Caudal Nucleus; Xanthines

ATP is released in a vesicular manner from nerve terminals mainly at higher stimulation frequencies. There is a robust expression of ATP (P2) receptors in the brain, but their role is primarily unknown. We report that ATP analogs biphasically modulate the evoked release of glutamate from purified nerve terminals of the rat hippocampus, the facilitation being mediated by P2X1, P2X2/3, and P2X3 [antagonized by 8-(benzamido)naphthalene-1,3,5-trisulfonate and 2',3'-O-(2,4,6-trinitrophenyl)-ATP] and the inhibition by P2Y1, P2Y2, and/or P2Y4 [antagonized by reactive blue 2 and 2'deoxy-N6-methyladenosine-3',5'-bisphosphate and mimicked by P1-(urinine 5'-),P4-(inosine 5'-) tetraphosphate and 2-methylthio-ADP] receptors. The combination of single-cell PCR analysis of rat hippocampal pyramidal neurons, Western blot analysis of purified presynaptic active zone fraction, and immunocytochemical analysis of hippocampal glutamatergic terminals revealed that the P2 receptors expressed in glutamatergic neurons, located in the active zone and in glutamatergic terminals, were precisely P2X1, P2X2, and P2X3 subunits and P2Y1, P2Y2, and P2Y4 receptors. This provides coincident functional and molecular evidence that P2 receptors are present and act presynaptically as a modulatory system controlling hippocampal glutamate release.

Topics: Adenosine Diphosphate; Adenosine Triphosphate; Adenylyl Imidodiphosphate; Animals; Astrocytoma; Calcium; Cell Line; Cell Line, Tumor; Glutamic Acid; Hippocampus; Kidney; Male; Potassium; Pyramidal Cells; Pyridoxal Phosphate; Rats; Rats, Wistar; Receptors, Presynaptic; Receptors, Purinergic P2; Receptors, Purinergic P2X; Receptors, Purinergic P2X2; Receptors, Purinergic P2X3; Receptors, Purinergic P2Y1; Receptors, Purinergic P2Y2; Recombinant Fusion Proteins; RNA, Messenger; Subcellular Fractions; Suramin; Synaptosomes; Transfection; Triazines; Triazoles; Xanthines