Interaction between DPI 201-106 enantiomers at the cardiac sodium channel

Abstract

The modification of cardiac sodium channels by DPI 201-106, its S-enantiomeric form (S)-DPI, and its R-enantiomeric form (R)-DPI was investigated with whole-cell voltage-clamp recording in single cultured ventricular myocytes obtained from late-fetal rats. From a holding potential of -100 mV, depolarizing pulses to -30 mV of 50-msec duration were applied at 0.2 Hz. Extracellular [Na] was reduced to 70 mM; temperature was 20 degrees. Drugs were administered directly on the cell by a double-barrelled microsuperfusion system. Sodium current inactivation was progressively slowed when the concentration of DPI 201-106 was increased from 0.3 to 3 microM. At 10 microM DPI 201-106, this effect was followed by a blocking effect on peak inward sodium current (INa), and at 30 microM inward sodium current was fully blocked within 2 min. The slowing of inactivation was produced by (S)-DPI (maximally effective at 3 microM), whereas (R)-DPI had little effect on inactivation at 3 microM. Conversely, (R)-DPI reduced INa at 10 microM, whereas (S)-DPI did not reduce INa at 3 microM. The effects of both (S)-DPI and (R)-DPI were partially reversed by washout. (R)-DPI retained its blocking activity on INa when the interval between depolarizing pulses was prolonged to 90 sec. In order to test whether the different sodium channel modifications produced by (S)-DPI and (R)-DPI were mutually exclusive, the INa-reducing activity of (R)-DPI was measured in the absence of (S)-DPI and after equilibration with a maximally effective (S)-DPI concentration. In the absence of (S)-DPI, 3 microM (R)-DPI reduced INa by 35% and in the presence of 3 microM (S)-DPI, by 51%. Thus, modification by (S)-DPI of sodium channels did not prevent their block by (R)-DPI. The INa-reducing activity of (R)-DPI was even significantly augmented by (S)-DPI after a 1-sec depolarization to -30 mV. During such prolonged pulses, (R)-DPI accelerated the monoexponential decay of the (S)-DPI-induced slow phase of sodium current inactivation. The results are consistent with an irreversible binding reaction between (R)-DPI and (S)-DPI-modified open sodium channels (association rate constant, 4.7 x 10(5) M-1sec-1). We conclude that (R)-DPI reduces INa by interacting both with resting sodium channels and with (S)-DPI-modified open sodium channels. The corresponding receptor site is stereoselective and distinct from and allosterically coupled to the (s)-DPI receptor that mediates slowing of inactivation.(ABSTRACT TRUNCATED AT 400 WORDS)