PURPOSE: Phosphodiesterase (PDE) inhibitors are important therapeutic agents, but their effects on photoreceptor PDE (PDE6) and photoreceptor cells are poorly understood. The potency and selectivity of various classes of PDE inhibitors on purified rod and cone PDE6 and on intact rod outer segments (ROS) were characterized. METHODS: The inhibition constant (K(i)) of isozyme-selective PDE inhibitors was determined for purified rod and cone PDE6. Perturbations of cGMP levels in isolated ROS suspensions by PDE inhibitors were quantitated by a cGMP enzyme-linked immunoassay. RESULTS: Most PDE5-selective inhibitors were excellent PDE6 inhibitors. Vardenafil, a potent PDE5 inhibitor (K(i) = 0.2 nM), was the most potent PDE6 inhibitor tested (K(i) = 0.7 nM). Zaprinast was the only drug that inhibited PDE6 more potently than did PDE5. PDE1-selective inhibitors were equally effective in inhibiting PDE6. In intact ROS, PDE inhibitors elevated cGMP levels, but none fully inhibited PDE6. Their potency for elevating cGMP levels in ROS was much lower than their ability to inhibit the purified enzyme. Competition between PDE5/6-selective drugs and the inhibitory gamma-subunit for the active site of PDE6 is proposed to reduce the effectiveness of drugs at the enzyme-active site. CONCLUSIONS: Several classes of PDE inhibitors inhibit PDE6 equally as well as the PDE family to which they are targeted. In intact ROS, high PDE6 concentrations, binding of the gamma-subunit to the active site, and calcium feedback mechanisms attenuate the effectiveness of PDE inhibitors to inhibit PDE6 and disrupt the cGMP signaling pathway during visual transduction.
