Nitric oxide-dependent efflux of cGMP in rat cerebellar cortex: An in vivo microdialysis study

Abstract

The stimulation of excitatory amino acid receptors in the cerebellar cortex results in the Ca2+/calmodulin-dependent activation of nitric oxide synthase. This leads to an increase in tissue levels of cGMP following the interaction of nitric oxide with soluble guanylyl cyclase. The cerebellar cortex has the highest levels of nitric oxide synthase and cGMP in the brain; however, the levels of guanylyl cyclase and cGMP-phosphodiesterase are remarkably low. Thus, the mechanisms regulating cGMP levels in cerebellar cells are unclear. One report has noted that cGMP can be released from cerebellar slices. We have therefore used intracerebellar microdialysis in awake, freely moving rats to test the hypothesis that activation of nitric oxide synthase in the cerebellar cortex results in the release of cGMP. Climbing fibers, which release excitatory amino acids in the cerebellum, were activated with systemic harmaline. This resulted in an immediate increase in extracellular cGMP, which was blocked by TTX or the removal of extracellular Ca2+, and attenuated by prior lesion of the climbing fibers. Blockade of N- type calcium channels with ω-conotoxin also antagonized the harmaline- induced increase. In contrast, blockade of L-type calcium channels, or inhibition of anion transport with probenecid or bromosulfophthalein, potentiated the increase in cGMP seen in response to harmaline. Inhibitors of nitric oxide synthase or guanylyl cyclase prevented the harmaline-induced increase in extracellular cGMP, while phosphodiesterase inhibitors potentiated the increase. Local application of the NMDA antagonist 2-amino- 5-phosphonopentanoic acid or the AMPA receptor antagonist 6-cyano-7- nitroquinoxaline-2,3-dione attenuated the effect of harmaline. In the presence of TTX, local application of NMDA, AMPA, or the metabotropic agonist trans-1-amino-1,3-cyclopentane-dicarboxylic acid (t-ACPD) elevated extracellular cGMP. Thus, activation of excitatory amino acid receptors results in a nitric oxide-dependent increase in the extracellular levels of cGMP in the cerebellar cortex. This may indicate the presence of an intercellular metabolic pathway for the regulation of intracellular cGMP levels. It is also possible that extracellular cGMP plays a role in intercellular communication in the cerebellar cortex.