calcimycin and malic-acid

The effects of intracellular free calcium (Ca2+) ions on the malate-aspartate shuttle were studied in slow-cooled boar spermatozoa. The capacity of the malate-aspartate shuttle was assessed by an indirect method on the basis of accumulation of lactate relative to pyruvate when ethanol is provided as substrate. The capacity of the malate-aspartate shuttle at 37 degrees C was dependent on the presence of Ca2+ ions and was stimulated by an influx of Ca2+ ions induced by the combination of the Ca2+ ionophore A23187 and 100 microM CaCl2. When washed spermatozoa were cooled slowly to 15 degrees C, the percentage of progressive motile spermatozoa was about half that at 37 degrees C in 100 microM Ca(2+)-containing medium, while the capacity of the malate-aspartate shuttle remained equal to that at 37 degrees C. The motility decreased further at higher concentrations of Ca2+ ions. Spermatozoa in EGTA-containing medium barely moved at 15 degrees C and the capacity of the malate-aspartate shuttle decreased. Even in Ca(2+)-containing medium, LaCl3 caused a decrease in the capacity of the malate-aspartate shuttle at 15 degrees C. These results suggest that an influx of a low concentration of Ca2+ ions activates the malate-aspartate shuttle at 15 degrees C, with a subsequent increase in the proportion of cells that maintain progressive motility.

Topics: Adenosine Diphosphate; Adenosine Triphosphate; Animals; Aspartic Acid; Calcimycin; Calcium; Cold Temperature; Egtazic Acid; Ethanol; Lanthanum; Malates; Male; Sperm Motility; Spermatozoa; Swine

Mitochondrial dysfunction has been implicated as the cause of irreversible injury in the ischemic heart. To circumvent artifacts associated with organelle isolation, mitochondrial function was studied in intact isolated, Ca2+-tolerant rat ventricular myocytes. After 30 min of anaerobic incubation, myocyte viability decreased from 76 +/- 1 to 33 +/- 4%. Basal O2 consumption rates (nanoatoms . mg cell protein-1 . min-1) were 17.1 +/- 1.3 in aerobic cells and 51.0 +/- 9.8 in anoxic cells. Carbonylcyanide-p-trifluoromethoxyphenyl hydrazone (FCCP)-stimulated rates were 65.5 +/- 9.2 and 84.5 +/- 15.3 in aerobic and anoxic cells, respectively. Respiratory control ratio was lower in anoxic cells: 2.3 +/- 0.3 versus 4.2 +/- 0.4 in aerobic cells. These data suggest that early anoxic mitochondrial injury is due to increased permeability of the inner membrane. Addition of pyruvate, malate, and FCCP to cells made permeable by digitonin resulted in similar maximal O2 consumption rates: 276.5 +/- 31.8 in aerobic and 299.3 +/- 31.9 in anoxic cells, suggesting the electron transport chain is intact in anoxic cells. For purposes of investigating whether anoxic mitochondrial dysfunction is secondary to cellular or mitochondrial Ca2+ overload, total cell Ca2+, cytosolic free Ca2+ levels (measured by null-point titration), and mitochondrial Ca2+ contents (measured as FCCP-releasable Ca2+) were measured. There were no differences in these three parameters between aerobic and anoxic cells, suggesting that mitochondrial dysfunction and irreversible hypercontraction of isolated cardiac myocytes exposed to 30 min of anoxia are not related to Ca2+ overload.

Topics: Animals; Calcimycin; Calcium; Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; Cell Membrane Permeability; Digitonin; Malates; Mitochondria, Heart; Myocardial Contraction; Oligomycins; Oxygen; Oxygen Consumption; Pyruvates; Pyruvic Acid; Rats; Ruthenium Red; Time Factors