CA2+ CALMODULIN-DEPENDENT CYCLIC-GMP PHOSPHODIESTERASE ACTIVITY IN GRANULE NEURONS AND ASTROCYTES FROM RAT CEREBELLUM/

Cyclic GMP (cGMP) formation induced by agonist stimulation of Ca2+/cal modulin-dependent nitric oxide (NO) synthase type I is known to occur in both granule cell and astrocyte cultures from rat cerebellum. Here we show that in these same cells cGMP is predominantly hydrolyzed by a Ca2+/calmodulin-dependent phosphodiesterase. At 10 mu M cGMP, Ca2+ (2 5 mu M) stimulated basal (Ca2+-independent) phosphodiesterase activity about 6 times in granular neurons and 15 times in astrocytes. The cal modulin antagonist calmidazolium blocked the Ca2+-dependent phosphodie sterase activity and exogenous calmodulin increased 3-4-fold the stimu latory potency of Ca2+ in both cell types (EC,, values 1.26 +/- 0.20 a nd 1.50 +/- 0.42 mu M in the absence and 0.38 +/- 0.11 and 0.39 +/- 0. 14 mu M in the presence of 1 mu M calmodulin, for neurons and astrocyt es, respectively). In both cell types K-m values for cGMP at 25 mu M C a2+ were similar (1.72 +/- 0.20 and 1.92 +/- 0.09 mu M) and phosphodie sterase activities were inhibited by isozyme-selective phosphodiestera se inhibitors with potencies analogous to tho se de scribed for Ca2+ / calmodulin-phosphodiesterases or phosphodiesterase type 1 isoforms in other preparations. The nonselective phosphodiesterase inhibitor 3-iso butyl-l-methylxantine (IBMX) effectively blocked the Ca2+/calmodulin-p hosphodiesterase activity in granule cell and astrocyte extracts (IC50 values at 1 mu M cGMP: 31 +/- 10 mu M and 46 +/- 6 mu M, respectively ), in contrast to the apparent inability of this compound to inhibit t he Ca2+-dependent activity reported in whole brain extracts. These res ults demonstrate that comparable phosphodiesterase type 1 activities a re found in the cytosols of cerebellar granule cells and astrocytes an d suggest that these activities may play an important role in controll ing cGMP levels in cells where the Ca2+-dependent NO synthase type I i s stimulated. (C) 1997 Elsevier Science B.V.