THE EFFECTS OF DIAZOXIDE AND BENZOTHIADIAZINE DIURETICS UPON PHOSPHODIESTERASE

Diazoxide causes hyperglycemia,' hypotension,2 tachycardia: and stimulaLion of lip01ysis. ~ It also non-specifically antagonizes contractions of isolated intestinal and aortic muscle induced by any number of spa smog en^^,^ and increases water transfer in the isolated toad bladder.6 All of these actions may be ascribed to elevating intracellular levels of cyclic AMP in various specific tissues. A significant portion of this profile is associated with the phosphodiesterase inhibitor, theophylline. Schultz and coworkers7 first reported that diazoxide inhibited phosphodiesterase. They proposed that a principal biochemical basis for the hyperglycemic action of diazoxide was inhibition of phosphodiesterase, which produced increased levels of cyclic AMP and, in turn, promoted glycolysis by stimulating glycogen phosphorylase and inhibiting glycogen synthetase. This inhibition of phosphodiesterase also caused potentiation of the hyperglycemic effect of endogenous catecholamines. This paper caused us to re-examine our results, which had indicated that diazoxide did not inhibit phosphodiesterase. A comparison of methods showed that both studies had used enzymes prepared from beef heart as outlined by Butcher and Sutherland.8 In our studies, we used the same analytical procedure and substrate level of cyclic AMP as Butcher and Sutherland,* namely, 4 X M. Schultz and as~ociates,~ however, had carried out their studies at substrate concentrations of 0.28 and 1.38 X lO-4M. Only when we reduced the substrate concentration to 1 X 10-4M was it possible for us to observe inhibition of phosphodiesterase by diazoxide. FIGURE 1 is a graphic illustration of the results of a study of theophylline, caffeine and various benzothiadiazine derivatives as inhibitors of phosphodiesterase. The study was carried out at a substrate concentration of 1 X 10-4M. It is conceivable that these agents would inhibit the enzyme at higher substrate concentrations, if their concentration in the media could also be elevated. However, their solubility in water at pH 7.5 was the limiting factor in this regard. Hydroflumethiazide, bendroflumethiazide and hydrochlorothiazide were also studied in this same experiment, but their activities could only be observed at two levels. In TABLE 1, the compounds have been arranged in decreasing order of approximate potency as inhibitors of phosphodiesterase. The relative potencies were determined by statistical analysis of the regression lines as outlined by Finney? without regard to deviations from parallelism. Although the paucity of data available for the last three compounds makes their relative potency values rather arbitrary, they are, nonetheless, important to the overall body of data, which indicates that all of the benzothiadiazines tested did, in fact, inhibit the enzyme. In man, the diazoxide-induced hyperglycemia is quite pronounced, and occurs upon acute administration of the drug, whereas the hyperglycemia associated with the diuretic benzothiadiazines generally occurs only after chronic administration of the compound. There are probably a number of explanations for this. The first consideration must be that in the in vitro test, the drug is placed in physical