J Mol Cell Cardiol23
INTRACELLULAR FREE CALCI'JM DYNAMICS AND TAURINE PREVENTION DURING THE CALCIUM PARADOX OF CULT'JRED MYOCARDIAL CELLS. K. Takahashl, K. Boku. Y. Ihara, H. Harada, A. Azuma. Third Sawamura, J. Department of Internal Medlclne Osaka Unlverslty Medlcal School, Osaka, Jdpan. Intracellular free calcium concentration [Ca'+], was continuously measured in spontaneously beating cultured myocytes durlnq the caLcium paradox using mlcroscope~fluorescent system, and preventive effect of taurlne was evaluated. Myocyte superfused with control culture medium (Ca'+: 2mM) exhlblted typ1ca1 C.Eiz+ transient (Ca-T) associated with spontaneous beating. Cd"-free medium lmmedlately ceased Ca-T and beatlnq, and decreased [Ca'+],, eventually reached about lo-' M. Reperfuslon with C.32 + Increased [Ca'+] , to 4-10 times the control level wlthln 20 sec. The recovery of spontaneous beating and Ca-T required over 7 min. Tallr1ne pretreatment (20 mM, 15 mln) reduced the excess accumulation of returned Ca-T and beating wlthln 5 mln [Ca*+l I, and after caZ+ reperfus1on. It was proposed that taurlne plays a role in regulation of CaZt homeostasls dlurlnq Ca'+ paradox.
!3FF3FF;g OF 2, 3TBUT+NXbIONX “NOX~ic;;D14)O~o VXNa~~CUL+X ““GE;;;; : COKPAkISON If-I:; Hlroyuki Department . Toshlyukl Takasago, Cardiovascular Dynamics, National Cardiovascular Center Research Institute, Suita. Osaka, Japan . It is known that BDM at a low concentration (up to 5mM) gives a negative inotropic
change in the calcium
The mechanism of this negative inotropic effect has been considered to be a direct inhibition of crossbridges. We attempted to clarify the mechanism of this effect of BDM in the isolated. blood-nerfused left ventricle of the doe from the viewooint of cardiac energetics.’ First, ‘we obtained the relationship beyween left ven;ricular oxygen consumption (Vo,) and systolic pressure-volume area (PVA) in a control steadystate contractility. Next, we depressed ventricular contractility in steps with BDM (up to 5mM) given into coronary circulation and obtained the Vos-PVA relation at a constant volume. Both Vos and PVA decreased with the gradually decreased contractility in terms of Rmax (an index of ventricular contractility). Finally, we enhanced ventricular contractility in steps with calcium and obtained the Vos-PVA relation. We calculated PVA-indeoendent Vo, values at varied Emax levels in both BDM and calcium runs and determined’ oxvsen c&t of Fmax as the slope of the relation between PVA-
independent different is the
Voa and XmaxT We found
of Fmsx was not significantly
between BDM and calcium. We have sum of basal metabolic Vos and the
alreadv shown that the PVA-indeoendent Vo, Vos fo> excitation-contraction-relaxatiori we conclude that the mechanism of the cycle which varies with Emax. Therefore, negative inotropic effect of BDM is mainly suppression of the total calcium handling in the excitation-contraction-relaxation cycle.
5-HYDROKYTRYP’li%MINE STlMULATES SPONTANEOUS BEA’DNG VlA PROTEIN KlNASE C IN CULTURED PETAL MOUSE VENTRICULAR MYOCYTES. Y. Hamamorl. M. Okamoto’. K. Gosbima’. M. Yamada. H. Akita. T. Nakarnura**, M. Yokoyama. The 1st Dept. of Med., Kobe Univ. School of Med., * Kobe-Gakuin Univ.. Kobe . l * Osaka Univ.. Osaka, Japan. 5-Hydroxytryptsmlne(5-HT) has positive lnotroplc and chronotroplc effects in the isolated heart and the molecular mechanisms of the 5-HT actions remain unknown. We studied signal transduction for the 5-l-D actions and the accompanied change in intracellular free calcium concentration(lCa2+11) in cultured fetal mouse ventrtcular myocytes. 5-HT stkrmlated phospholnositldes hydrolysis tlmeand dose-dependently (Berrldge’s method). 5-HT dld not affect CAMP level(radiohnmunoassay1. 5-HT stimulated rate and force of spontaneous beattng[SB) of the cultured myocytesfan optical edge tracking method) and these effects were mimicked by protein kinase C activators, TPA. PDBu and OAG. 5-HT effects on both PI hydrolysis and SB were inhibited by ketanserln. a specific 5-HT2 antagontst. but not by al- and 5-blockers. Staurosporine. a protein klnase C lnhlbitor, abolished most of the effects of 5-HT. TPA and PDBu on SB. 5-HT-stimulated SB force was accompanted by the mild increase in systolic [Ca2+]i from 358f81 to 401+87nM(Pc 0.001. n=ll). as measured by fluo-3 AM, while TPA and PDBu stimulated SB force without increasing [Ca2+]i. These results suggest that 5HT induced PI hydrolysis and that subsequent activation of protein klnase C may, m part, be responsible for 5-l-ll%stlmulated SB. Protein kinase C may stimulate SB force by increasing myoftbril sensitlvlty to Ca2+ and this mechanism may also operate In the 5-HT action. A role of inositol trlsphosphate in the 5-HT actions remams to be studied. S.28