K-CHANNEL OPENING ACTIVITY OF DIHYDROPYRIDINE ZM244085 - EFFECT ON (86)RB EFFLUX AND H-3 P1075 FINDING IN URINARY-BLADDER SMOOTH-MUSCLE

Zeneca ZM244085, 9-(3 cyanophenyl)hexahydro-1,8 acridinedione, is a no vel dihydropyridine (DHP) which relaxes KCl precontracted urinary blad der smooth muscle in vitro. The effect of ZM244085 on low and high KCl induced contractions, Rb-86 efflux and [H-3]-P1075 binding in guinea pig bladder strips was investigated to characterize the K-channel open ing properties of this compound. Since ZM244085 is a dihydropyridine i ts effect on DHP binding sites on Ca2+ channels was also investigated. ZM244085 was found to be more potent in relaxing detrusor strips prec ontracted with 15 mM KCl than strips precontracted with 80 mM KCl (Li et al., 1995). This functional profile of ZM244085 is similar to that exhibited by typical K-channel openers (PCO). In addition, inhibition of ZM244085 induced relaxation of detrusor strips by glibenclamide sug gests that ZM244085 opens ATP sensitive K-channels (K-ATP) in urinary bladder (Li et al., 1995). Since the glibenclamide sensitive smooth mu scle relaxation activity of ZM244085 could still be an indirect effect of this compound on K-ATP channels we carried out Rb-86 efflux studie s and [H-3]-P1075 binding studies to further confirm these findings. T he Rb-86 efflux assay is a direct method for monitoring the movement o f K+ ions across the cell membranes. Displacement of [H-3]-P1075 bindi ng to bladder membranes supports a direct action of the compound on th e K-ATP channel. The present study demonstrates that ZM244085 in a con centration dependent manner increases the rate of Rb-86 efflux from gu inea pig bladder strips. This effect was inhibited by glibenclamide (3 0 uM), a known K-ATP channel blocker. In addition, interaction of ZM24 4085 with K-ATP channels was also confirmed in human bladder smooth mu scle cells using a K-42 efflux assay. Furthermore, we were able to dem onstrate that ZM244085, structurally distinct PCO, inhibited the bindi ng of H-3-P1075 to urinary bladder strips in a manner similar to other K-ATP openers such as cromakalim and pinacidil. Inhibition of H-3-P10 75 binding by ZM244085 and other PCO's correlates well with increases in Rb-86 efflux and bladder muscle relaxation studies. Finally, ZM2440 85 did not exhibit any significant affect on VSCC as evidenced by very weak inhibition of [H-3]-PN200,110 binding to bladder membranes by ZM 244085. It is concluded that Zeneca ZM244085 is a PCO which activates K-ATP channels in urinary bladder.