Exogenous nitric oxide (NO) donors (S-nitroso-N-acetyl-DL-penicillamine and (E)-4-ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexenamide) and simultaneous application of high potassium and glutamate led to the phosphorylation of extracellular signal-regulated kinase (ERK)1/2 in cerebellar Purkinje cells. NO-induced ERK1/2 phosphorylation was abolished by the addition of MEK (MAPK ERK kinase) inhibitor (PD98059) or oxyhemoglobin, and was significantly reduced by the PKG (cGMP-dependent protein kinase) inhibitor, KT5823. KT5823 also reduced the peak levels of ERK1/2 activation induced by high K-glutamate or NO in Purkinje cell somata. NO-induced ERK1/2 phosphorylation remained above basal levels for at least 1 h after NO-donor removal. To address the physiological role of NO-induced ERK activation in Purkinje cells, we examined the effects of NO and 8-bromoadenosine 3', 5'-cyclic monophosphate (8-Br-cGMP) on PKC-dependent receptor declustering, which underlies cerebellar long-term depression. TPA (12-Ø-tetradecanoylphorbol 13-acetate), a potent activator of PKC, induced GluR2/3 receptor declustering. The 8-Br-cGMP or NO-donors enhanced GluR2/3 declustering, but did not induce it. These results suggest an important role for the NO-cGMP-PKG pathway in the activation of ERK1/2 and GluR2/3 (ionotropic glutomate receptor subunit 2/3) declustering in Purkinje cells.