Stretch‐Induced Increase in Cardiac Contractility Is Independent of Myocyte Ca\u3csup\u3e2+\u3c/sup\u3e While Block of Stretch Channels by Streptomycin Improves Contractility After Ischemic Stunning

Stretching the cardiac left ventricle (LV) enhances contractility but its effect on myoplasmic [Ca2+] is controversial. We measured LV pressure (LVP) and [Ca2+] as a function of intra-LV stretch in guinea pig intact hearts before and after 15 min global stunning ± perfusion with streptomycin (STM), a stretch activated channel blocker. LV wall [Ca2+] was measured by indo-1 fluorescence and LVP by a saline-filled latex balloon inflated in 50 μL steps to stretch the LV. We implemented a mathematical model to interpret crossbridge dynamics and myofilament Ca2+ responsiveness from the instantaneous relationship between [Ca2+] and LVP ± stretching. We found that: (1) stretch enhanced LVP but not [Ca2+] before and after stunning in either control (CON) and STM groups, (2) after stunning [Ca2+] increased in both groups although higher in STM versus CON (56% vs. 39%), (3) STM-enhanced LVP after stunning compared to CON (98% vs. 76% of prestunning values), and (4) stretch-induced effects on LVP were independent of [Ca2+] before or after stunning in both groups. Mathematical modeling suggested: (1) cooperativity in cross-bridge kinetics and myofilament Ca2+ handling is reduced after stunning in the unstretched heart, (2) stunning results in depressed myofilament Ca2+ sensitivity in the presence of attached cross-bridges regardless of stretch, and (3) the initial mechanism responsible for increased contractility during stretch may be enhanced formation of cross-bridges. Thus stretch-induced enhancement of contractility is not due to increased [Ca2+], whereas enhanced contractility after stunning in STM versus CON hearts results from improved Ca2+ handling and/or enhanced actinomyosin cross-bridge cycling

Mg\u3csup\u3e2+\u3c/sup\u3e Differentially Regulates Two Modes of Mitochondrial Ca\u3csup\u3e2+\u3c/sup\u3e Uptake in Isolated Cardiac Mitochondria: Implications for Mitochondrial Ca\u3csup\u3e2+\u3c/sup\u3e Sequestration

The manner in which mitochondria take up and store Ca2+ remains highly debated. Recent experimental and computational evidence has suggested the presence of at least two modes of Ca2+ uptake and a complex Ca2+ sequestration mechanism in mitochondria. But how Mg2+ regulates these different modes of Ca2+ uptake as well as mitochondrial Ca2+ sequestration is not known. In this study, we investigated two different ways by which mitochondria take up and sequester Ca2+ by using two different protocols. Isolated guinea pig cardiac mitochondria were exposed to varying concentrations of CaCl2 in the presence or absence of MgCl2. In the first protocol, A, CaCl2 was added to the respiration buffer containing isolated mitochondria, whereas in the second protocol, B, mitochondria were added to the respiration buffer with CaCl2 already present. Protocol A resulted first in a fast transitory uptake followed by a slow gradual uptake. In contrast, protocol B only revealed a slow and gradual Ca2+ uptake, which was approximately 40 % of the slow uptake rate observed in protocol A. These two types of Ca2+ uptake modes were differentially modulated by extra-matrix Mg2+. That is, Mg2+ markedly inhibited the slow mode of Ca2+ uptake in both protocols in a concentration-dependent manner, but not the fast mode of uptake exhibited in protocol A. Mg2+ also inhibited Na+-dependent Ca2+ extrusion. The general Ca2+ binding properties of the mitochondrial Ca2+ sequestration system were reaffirmed and shown to be independent of the mode of Ca2+ uptake, i.e. through the fast or slow mode of uptake. In addition, extra-matrix Mg2+ hindered Ca2+ sequestration. Our results indicate that mitochondria exhibit different modes of Ca2+ uptake depending on the nature of exposure to extra-matrix Ca2+, which are differentially sensitive to Mg2+. The implications of these findings in cardiomyocytes are discussed

Ischemia reperfusion dysfunction changes model-estimated kinetics of myofilament interaction due to inotropic drugs in isolated hearts

BACKGROUND: The phase-space relationship between simultaneously measured myoplasmic [Ca(2+)] and isovolumetric left ventricular pressure (LVP) in guinea pig intact hearts is altered by ischemic and inotropic interventions. Our objective was to mathematically model this phase-space relationship between [Ca(2+)] and LVP with a focus on the changes in cross-bridge kinetics and myofilament Ca(2+ )sensitivity responsible for alterations in Ca(2+)-contraction coupling due to inotropic drugs in the presence and absence of ischemia reperfusion (IR) injury. METHODS: We used a four state computational model to predict LVP using experimentally measured, averaged myoplasmic [Ca(2+)] transients from unpaced, isolated guinea pig hearts as the model input. Values of model parameters were estimated by minimizing the error between experimentally measured LVP and model-predicted LVP. RESULTS: We found that IR injury resulted in reduced myofilament Ca(2+ )sensitivity, and decreased cross-bridge association and dissociation rates. Dopamine (8 μM) reduced myofilament Ca(2+ )sensitivity before, but enhanced it after ischemia while improving cross-bridge kinetics before and after IR injury. Dobutamine (4 μM) reduced myofilament Ca(2+ )sensitivity while improving cross-bridge kinetics before and after ischemia. Digoxin (1 μM) increased myofilament Ca(2+ )sensitivity and cross-bridge kinetics after but not before ischemia. Levosimendan (1 μM) enhanced myofilament Ca(2+ )affinity and cross-bridge kinetics only after ischemia. CONCLUSION: Estimated model parameters reveal mechanistic changes in Ca(2+)-contraction coupling due to IR injury, specifically the inefficient utilization of Ca(2+ )for contractile function with diastolic contracture (increase in resting diastolic LVP). The model parameters also reveal drug-induced improvements in Ca(2+)-contraction coupling before and after IR injury