Inhibition of Ca2+ current in neonatal and adult rat ventricular myocytes by the tyrosine kinase inhibitor, genistein.
Yokoshiki et al. (Yokoshiki, H., Sumii, K., Sperelakis, N., 1996. Inhibition of L-type calcium current in rat ventricular cells by the tyrosine kinase inhibitor, genistein and its inactive analog, daidzein. J. Mol. Cell. Cardiol. 28, 807-814) reported that genistein and daidzein inhibited L-type Ca2+ current (I(Ca)(L)) in young rat ventricular cells. Therefore, we investigated the developmental differences in the effect of genistein, an inhibitor of tyrosine kinases, on I(Ca)(L) in freshly-isolated neonatal (3-7 days) and adult (2-5 months) rat ventricular myocytes using whole-cell voltage clamp and single-channel recordings (cell-attached configuration). For whole-cell voltage clamp, I(Ca)(L) was measured as the peak inward current at a test potential of +10 mV by applying a 300 ms pulse from a holding potential of -40 mV. To isolate I(Ca(L), the pipette solution was Cs+-rich and the bath solution was Na+-, K+-free. Ca2+ (1.8 mM) was used as charge carrier. Bath application of 100 microM genistein (sufficient for maximal effect) decreased the basal I(Ca)(L) by 43.3% (n = 27) in neonatal cells and by 30.6% (n = 14) in adult cells (P < 0.05). In the current/voltage relationships, the potential of peak I(Ca)(L) was shifted to the right by genistein by 8.6 mV in neonatal and by 9.3 mV in adult cells. Genistein produced a shift of the steady-state inactivation curve (to the left) in neonatal cells (from -16.0 +/- 3.9 mV to -26.1 +/- 4.2 mV; P < 0.05) and in adult cells (-15.9 +/- 3.2 mV to -22.9 +/- 3.3 mV; P < 0.05); the slope factor was not affected. For single-channel recordings in cell-attached patches, Ca2+ currents were evoked by applying a 150 ms pulse from a holding potential of -40 mV to a test potential of 0 mV. The pipette solution contained 110 mM Ba2+ (as charge carrier), and the bath solution contained 150 mM K+ (to bring resting potential to near zero). Genistein (50 microM) decreased the open probability of the channels from 2.8% to 0.75% (P < 0.05) in absence of Bay K 8644, and from 24% to 7.9% (P < 0.05) in presence of Bay K 8644; the mean open time and the slope conductance of the currents were not affected. In conclusion, (1) genistein inhibits the basal I(Ca)(L) in rat ventricular cells and (2) the inhibition of I(Ca)(L) by genistein is greater in immature cells than in adult cells.
Katsube
Y
Y
Department of Pediatrics, Nippon Medical School Hospital, Tokyo, Japan.
Yokoshiki
H
H
Nguyen
L
L
Yamamoto
M
M
Sperelakis
N
N
D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester / Q000494:pharmacology; D000818:Animals; D000831:Animals, Newborn; D002120:Calcium Channel Agonists / Q000494:pharmacology; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D019833:Genistein / Q000494:pharmacology*; D006321:Heart / Q000187:drug effects; D066298:In Vitro Techniques; D008564:Membrane Potentials / Q000187:drug effects; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors*; D051381:Rats; D017207:Rats, Sprague-Dawley
D002120:Calcium Channel Agonists; D002121:Calcium Channel Blockers; D004791:Enzyme Inhibitors; D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; D019833:Genistein; D011505:Protein-Tyrosine Kinases