Characterization of ionotropic glutamate receptor-mediated nitric oxide production in vivo in rats

BACKGROUND AND PURPOSE: Glutamate receptor activation can stimulate nitric oxide (NO) production and possibly play a role in long-term potentiation and excitotoxic-mediated injury. We studied the differential effect of agonist-induced activation of ion channel-linked N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor subtypes on NO production in vivo in rat hippocampus. We also studied whether dantrolene, a ryanodine calcium channel inhibitor previously shown to attenuate metabotropic glutamate receptor stimulation of NO production, also attenuated ionotropic glutamate receptor-mediated stimulation of NO production. METHODS: Microdialysis probes were placed bilaterally into the CA3 region of the hippocampus of pentobarbital-anesthetized adult Sprague-Dawley rats and were perfused for 5 hours with artificial cerebrospinal fluid (CSF) containing 3 mumol/L 14C]L-arginine. Recovery of 14C]L-citrulline in the effluent was used as a marker of NO production. In 13 groups of rats, increases in 14C]L-citrulline recovery were compared between right- and left-sided probes perfused with no additional drugs versus combinations of NMDA, AMPA, the NO synthase inhibitor N omega-nitro-L-arginine methyl ester (L-NAME), the non-competitive glutamate receptor blocker MK-801, the AMPA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), and dantrolene. RESULTS: Recovery of 14C]L-citrulline during perfusion with artificial CSF progressively increased to 272 +/- 73 fmol/min (+/-SEM) over 5 hours. Contralateral perfusion with 1 mmol/L L-NAME inhibited 14C]L-citrulline recovery. Perfusion with 1 mmol/L MK-801 or 1 mmol/L CNQX reduced 14C]L-citrulline recovery compared with contralateral perfusion with CSF alone. Perfusion with 1 mmol/L NMDA enhanced 14C]L-citrulline recovery, and this enhancement was attenuated by L-NAME, MK-801, and CNQX but not by dantrolene. Perfusion with 1 mmol/L AMPA enhanced 14C]L-citrulline recovery, and this enhancement was also attenuated by L-NAME, MK-801, and CNQX but not by dantrolene. CONCLUSIONS: Through an indirect method of assessing NO production in vivo, results with MK-801 and CNQX indicate that NMDA and AMPA receptor activation contribute to basal NO production in the rat hippocampus. Enhanced NO production with NMDA and AMPA agonists appears to involve a complex neuronal interaction because the effect of NMDA was attenuated by both MK-801 and CNQX and because the effect of AMPA was attenuated by both CNQX and MK-801. In contrast to metabotropic glutamate receptor activation, release of calcium from intracellular ryanodine calcium channels does not appear to be a prominent mediator of ionotropic glutamate receptor stimulation of NO production.