Mechanism Of Triggering Excitation-contraction Coupling In Guinea-pig Ventricular Myocytes

Mechanism of triggering excitation-contraction coupling in guinea-pig ventricular myocytesOBJECTIVE: We studied use-dependent and use-independent components of L-type calcium current (Ica(L)) block by nifedipine (Nif) and compared the characteristics of Ca2"1" release from the sarcoplasmic reticulum (SR) triggered by L-type calcium current and by reverse-mode Na/Ca exchange during depolarizing steps in single guinea-pig ventricular myocytes.METHODS: Whole-cell membrane-potential, membrane-current and cell-shortening data were simultaneously acquired during whole-cell voltage clamp protocols. Membrane current and cell contraction were measured by patch pipettes (K+-baesd pipette solution with 10 mmol/L [Na]p ) and edge detection. Conditioning pulses to +10mV or +30mV ensured comparable Ca2+ loading of the SR. Voltage clamp pulses elicited Ica(L) at +10mV, +50mV, +100mV and evoked contractions in myocytes supervised with Tyrode's solution at 35癈. Ica(L) is taken as Cd2"1" (0.1-0.3mM)-sensitive current.RESULTS: With Cs+-rich pipette solution (-80mV holding potential), half-maximal inhibition dose (ICso) of nifedipine for the peak at +10mV was 0.3 uM. At holding potential -40mV, the ICso decreased to O.OSuM indicating preferential block of inactivated L-type Ca2+ channels. With K+-rich pipette solution, use-independent block of Ica(L) increased in proportion to nifedipine concentration (lO-lOOjiM) and rest intervals (5-30s). The magnitude of use-dependent block decreased as the nifedipine concentration increased from 10 to 100 uM, and the combination of Nif30/Cd2+nM. Nifedipine (3 and 30 u,M) accelerated ICa(Dinactivation on the first test pulse. The greater the inhibition of Ica(L), the more likely contraction will be abolished at +10mV test potential. Nif 30/Cd2+ 30uM was more effective than 100 uM Nif alone as a suppressor of contractions. Nifedipine and Nif 30/Cd2+ 30 uM had no detectable effect on Na/Ca exchange current (iNa/ca) at +10 to +100mV test potentials. At potential positive to +50mV, contractions were partially suppressed by lOOuM nifedipine or Nif 30/Cd2+ 30 uM. The residual contraction was significantly delayed in onset. At +100mV test potential, contractions were delayed in onset compare to +50 mV and resistant to Nif lOOuM or Nif 30/Cd2+ 30 uM. The greater the inhibition of L-type calcium current, the more cell shortening will be decreased under voltage clamp conditions.CONCLUSIONS: Use-independent block of Ica(L) by nifedipine increases with increasing concentration and application time, and use-dependent block of Ica