Saturation characteristics, Ca2+ ions and drug-induced block of cardiac Isi-mediated action potentials.

Abstract

Isi saturation characteristics and the interaction of Ca2+ with the slow channel blockers verapamil and nifedipine were studied in microelectrode experiments on papillary muscles of guinea pigs by analyzing \(\dot V_{\max } \) of Isi-mediated action potentials. 1. A non-linear response of \(\dot V_{\max } \) on external Ca2+ variations between 2 and 6 mmol/l was obtained strongly suggesting saturation characteristics of the Isi system. Transformation of each individual \(\dot V_{\max } \)-[Ca2+]0 relationship into the Lineweaver-Burk form revealed apparent Km's for Ca2+ which varied from 1.5 to 31.2 mmol/l (mean: 7.96±1.07 mmol/l). Values of 20–108 V/s were calculated for the saturated \(\dot V_{\max } \), i.e. \(\dot V_{\max } \) at an infinitely high [Ca2+]0. 2. Histamine did not alter the apparent Km for Ca2+ but produced a concentration-dependent rise of the saturated \(\dot V_{\max } \). 3. Alkalinization of the medium lowered the apparent Km for Ca2+ from 9.2±1.1 mmol/l (at pH 7.4) to 2.7±0.2 mmol/l (at pH 9.0). 4. Sr2+ and Na+ compete with Ca2+ for common anionic sites. The ratio Ksr2+: KCa2+ amounted to 1.75±0.21. The apparent Km for Na+ should be much smaller than KCa2+ because external Na+ reduction from 150 to 75 mmol/l led to an increase of \(\dot V_{\max } \). 5. Nifedipine and, when analyzed under rested-state conditions, verapamil induced alterations of the \(\dot V_{\max } \)-[Ca2+]0 relationship which perfectly fit the formalism of competitive interaction. The y-intercept of the straight relating 1/\(\dot V_{\max } \) to 1/[Ca2-]0 remained either unchanged or varied insignificantly whilst the x-intercept strongly shifted to higher values. Repetitive stimulation changed the mode of verapamil action from a competitive to a non-competitive type. 6. Ca2+ modulates both the verapamil- and the nifedipine-induced Isi block in that excess Ca2+ attenuates drug action and vice versa. The same decline of block develops after adding Sr2+ to the superfusate. 7. The Ca2+ sensitivity of the both types of varapamil-induced Isi inhibition, tonic (rested-state) and phasic (use-dependent) block, differs considerably. The former declined in response to a threefold increase in external Ca2+ by a factor of 3.18, but the latter (analyzed at a driving rate of 12/min) by a factor of only 1.18. Consistent with this pecularity, concentration-response analysis of the verapamil action yielded individual apparent dissociation constants and nH coefficients being (at 3 mmol/l Ca2+) 6.4×10−6 mol/l and 3.91 for tonic Isi blockade, but 1×10−6 mol/l and 1.06 for phasic (frequency 12/min) Isi blockade.