REGULATION OF CYCLIC NUCLEOTIDE PHOSPHODIESTERASES OF CEREBRAL CORTEX BY Ca 2+ AND CYCLIC GMP

— Cyclic nucleotide phosphodiesterase activity of porcine cerebral cortical extracts was measured with 0.1–100 μM‐cyclic AMP and cyclic GMP and found to be dependent on both Ca 2+ and added cyclic nucleotides. With decreasing substrate concentration activity with cyclic GMP became more dependent on Ca 2+ whereas hydrolysis of cyclic AMP became less dependent. Cyclic GMP at 3 μM stimulated the hydrolysis of 0.1–10μM‐cyclic AMP in the absence of Ca 2+ (< 10 ‐10 M) but inhibited activity with 200 μM‐Ca 2+ present. This differential, substrate‐ and Ca 2+ ‐dependent regulation was attributed to the presence of at least two types of phosphodiesterase distinguishable by DEAE‐column chromatography. In the absence of Ca 2+ , activity with 1 μM‐cyclic GMP eluted in one minor peak followed by two major peaks, D‐I and D‐II. Activity with 1 μM‐cyclic AMP eluted almost entirely in D‐II. Hydrolysis of cyclic AMP in D‐II was activated by cyclic GMP. With added Ca 2+ plus a Ca 2+ ‐dependent regulator (CDR), activity with 1 μM‐cyclic GMP was markedly increased and eluted entirely at D‐I. Total activity with 1 μM‐cyclic AMP was only moderately increased and eluted as D‐I with a shoulder at D‐II. Elution profiles with 100 μM‐substrate were relatively independent of substrate, with D‐I predominant with Ca 2+ ·CDR present and D‐II predominant in its absence. Kinetic analysis of rechromatographed D‐I showed a 20‐ to 40‐fold activation by Ca 2+ ·CDR that was largely due to an increase in V max , with only 50% decreases in K m Both substrates competitively inhibited hydrolysis of the other with K i values equal to their respective K m values (1.7 μM for cyclic GMP and 48 μM for cyclic AMP with Ca 2+ ‐CDR present). Studies with theophylline and trifluoperazine indicate differential, substrate‐dependent inhibitions of both enzymes. These findings demonstrate that phosphodiesterase activity in neural tissue is subject to regulation by Ca 2+ , cyclic GMP, and inhibitors in a complex, substrate‐specific and concentration‐dependent manner.