cyclic-gmp and cyclothiazide

We investigated the role of nitric oxide (NO) on mitochondrial complexes activity, following short-term non-desensitizing activation of AMPA receptors with kainate (KA) plus cyclothiazide (CTZ), in cultured rat hippocampal neurons. In these conditions, we observed a decrease in the activity of mitochondrial complexes I, II/III, and IV. A selective neuronal nitric oxide synthase inhibitor, 7-Nitroindazole, prevented the decrease in the activity of mitochondrial complex I, but not for the other complexes. Exposure to KA plus CTZ also increased cyclic GMP levels significantly, and led to increased levels of 3-nitrotyrosine, a biomarker for peroxynitrite production. Taken together, our results suggest that non-desensitizing activation of AMPA receptors causes inhibition of mitochondrial complex I via peroxynitrite.

Topics: Analysis of Variance; Animals; Antihypertensive Agents; Benzothiadiazines; Cells, Cultured; Cyclic GMP; Drug Interactions; Embryo, Mammalian; Enzyme Inhibitors; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Hippocampus; Immunohistochemistry; Kainic Acid; Microtubule-Associated Proteins; Neurons; NG-Nitroarginine Methyl Ester; Nitric Oxide; Peroxynitrous Acid; Proton Pumps; Quinoxalines; Rats; Receptors, AMPA; Tyrosine

Monitoring of extracellular cGMP during intracerebral microdialysis in freely moving rats permits the study of the functional changes occurring in the glutamate receptor/nitric oxide (NO) synthase/guanylyl cyclase pathway and the relationship of these changes to animal behaviour. When infused into the rat hippocampus in Mg2+-free medium, cyclothiazide, a blocker of desensitization of the AMPA-preferring receptor, increased cGMP levels. The effect of cyclothiazide (300 microM) was abolished by the NO synthase inhibitor L-NARG (100 microM) or the soluble guanylyl cyclase inhibitor ODQ (100 microM). During cyclothiazide infusion the animals displayed a pre-convulsive behaviour characterized by frequent "wet dog shakes" (WDS). Neither L-NARG nor ODQ decreased the WDS episodes. Both cGMP and WDS responses elicited by cyclothiazide were prevented by blocking NMDA receptor function with the glutamate site antagonist CGS 19755 (100 microM), the channel antagonist MK-801 (30 microM) or Mg2+ ions (1 mM). The AMPA/kainate receptor antagonists DNQX (100 microM) and NBQX (100 microM) abolished the WDS episodes but could not inhibit the cyclothiazide-evoked cGMP response. DNQX or NBQX (but not MK-801) elevated, on their own, extracellular cGMP levels. The cGMP response elicited by the antagonists appears to be due to prevention of a glutamate-dependent inhibitory GABAergic tone, since infusion of bicuculline (50 microM) caused a strong cGMP response. The results suggest that (a) AMPA/kainate receptors linked to the NO/cGMP pathway in the hippocampus (but not NMDA receptors) are tonically activated and kept in a desensitized state by endogenous glutamate; (b) blockade of AMPA/kainate receptor desensitization by cyclothiazide leads to endogenous activation of NMDA receptors; (c) the hippocampal NO/cGMP system is under a GABAergic inhibitory tone driven by non-NMDA ionotropic receptors; (d) the pre-convulsive episodes observed depend on hippocampal NMDA receptor activation but not on NO and cGMP production.

Topics: Animals; Behavior, Animal; Benzothiadiazines; Cyclic GMP; Diuretics; Drug Interactions; Excitatory Amino Acid Antagonists; gamma-Aminobutyric Acid; Hippocampus; Male; Nitric Oxide; Nitric Oxide Synthase; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, Glutamate; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Sodium Chloride Symporter Inhibitors

1. Desensitization is an important characteristic of glutamate receptors of the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) type. 2. Stimulation of N-methyl-D-aspartate (NMDA) or AMPA receptors in cerebellum results in increased production of cyclic GMP. We have investigated AMPA receptor desensitization in vivo by monitoring extracellular cyclic GMP during intracerebellar microdialysis in conscious unrestrained adult rats. 3. Local infusion of AMPA (10 to 100 microM) caused dose-related elevations of cyclic GMP levels. The effect of AMPA was prevented by the non-NMDA receptor antagonist, 6,7-dinitroquinoxaline-2,3-dione (DNQX) and by the nitric oxide (NO) synthase inhibitor NG-nitro-L-arginine (L-NOARG). 4. In the absence of AMPA, DNQX lowered the basal levels of cyclic GMP whereas the NMDA receptor channel antagonist dizocilpine (MK-801) was ineffective. 5. Cyclothiazide, a blocker of AMPA receptor desensitization, potentiated the cyclic GMP response to exogenous AMPA. Moreover, cyclothiazide (100-300 microM) produced on its own dose-dependent elevations of extracellular cyclic GMP. The cyclothiazide-induced response was prevented not only by DNQX but also by MK-801. 6. While the cyclic GMP response elicited by AMPA was totally insensitive to MK-801, the response produced by AMPA (10 microM) plus cyclothiazide (30 microM) was strongly attenuated by the NMDA receptor antagonist (30 microM). 7. The results suggest that (a) AMPA receptors linked to the NO-cyclic GMP pathway in the cerebellum can undergo desensitization in vivo during exposure to exogenous AMPA; cyclothiazide inhibits such desensitization; (b) AMPA receptors (but not NMDA receptors) are 'tonically' activated and kept in a partly desensitized state by endogenous glutamate; (c) if cyclothiazide is present, activation of AMPA receptors may permit endogenous activation of NMDA receptors.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Benzothiadiazines; Cerebellum; Cyclic GMP; Dizocilpine Maleate; Male; Microdialysis; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate