Diltiazem causes open channel block of recombinant 5-HT3 receptors

1. To extend our knowledge of the site and mechanism of action of L-type Ca 2+ channel antagonists on 5-HT3 receptors, whole-cell voltage clamp electro physiology was used to investigate the action of one of these compounds, di ltiazem, on the recombinant receptor expressed in human embryonic kidney (H EK) 293 cells. 2. Application of diltiazem with 5-HT (30 mu M) caused an increase in the r ate of receptor current decay, but did not significantly affect peak curren t (I-p), the EC50 or the Hill coefficient, indicating a non-competitive mec hanism of action. Pre-application of the antagonist had no effect indicatin g that diltiazem mediates its effects by binding preferentially to the open state of the 5-HT3 receptor. 3. To examine the effects of diltiazem on the open state of the receptor in more detail we used 10 mM 5-hydroxyindole (5-OHi) to reduce receptor desen sitisation. These experiments showed that diltiazem causes a rapid, reversi ble, block in the presence of agonist but can become trapped in the unligan ded state of the receptor by prior washout of agonist. Dose-inhibition data yielded an IC50 of 5.5 mu M and a Hill coefficient of 0.96; inhibition was slightly voltage dependent as the degree of blockade at +60 mV was reduced . 4. The Hill coefficient of near unity suggests a single molecule of diltiaz em mediates inhibition and, indeed, kinetic analysis verified that the inte raction of diltiazem with the 5-HT3 receptor was well described by a bimole cular reaction scheme. The results suggest that diltiazem acts by causing o pen-channel block of the 5-HT3 receptor.