Ca2+ SIGNALLING IN MYOCYTES: TARGETING BY A NOVEL HORMONE-LIKE COMPOUND EXERTING POSITIVE INOTROPIC EFFECTS.

M. Keller (1), B. Vacher (2), C. Pignier (2), E. Niggli (1), M. Egger (1).

(1) Dept. of Physiology, University of Bern, Switzerland. (2) Centre de Recherche Pierre-Fabre, Castres, France.  

Heart failure is correlated with a loss of efficiency of cardiac function. A pharmaceutical compound increasing the cardiac function (positiv-inotropic effect) in a specific manner could therefore be used to treat this loss of function. For a newly developed compound (F90927) with structural similarities to steroid hormones, positive inotropic effects were found on the systemic level. Application of F90927 in microM concentrations increased the left ventricular developed pressure (LVDP) reversibly by 50 %, whereas the heart rate remained unchanged. However, the underlying cellular and molecular mechanisms by which of F90927 exerts these beneficial effects are unknown. On the cellular level, positive inotropic effects on cardiac myocytes can be mediated by larger Ca2+ influx or reduced Ca2+ efflux, consequently increased Ca2+ transients by augmented Ca2+ release from the sarcoplasmic reticulum or by modifying the Ca2+ sensitivity of the myofilaments. In this study, the L-type Ca2+ channel as a possible target for the F90927 compound and a key element of excitation-contraction-coupling in ventricular myocytes was examined in detail. Experiments were carried out with ventricular myocytes isolated from guinea-pig hearts. L-type Ca2+ currents were examined with the whole-cell patch clamp technique. In addition, intracellular Ca2+ transients were examined with laser-scanning confocal microscopy of fluo-3 fluorescence. In single cells, F90927 increased ICaL about 2.5 fold at 10 microM, while Na+-currents and the Na+-Ca2+ exchanger activity remained unaffected. The dose-response curve showed a biphasic increase of ICaL with two Kd of approximately 10 nM and approximately 2 microM, respectively. The ICaL/V relationship exhibited a leftward shift by 8.5 mV. The voltage-dependence of ICaL activation and inactivation revealed a shift of the activation curve by 20.4 mV towards negative potentials (at 10 microM). We conclude that F90927 exerts its positive inotropic effect as an L-type Ca2+ channel agonist. The current is increased by shifting the voltage-dependent activation towards more negative potentials. In addition, there appear to be two binding sites for F90927 on the channel (or on regulatory proteins), exhibiting distinctly different affinities.