Mitochondrial NAD(P)H, ADP, oxidative phosphorylation, and contraction in isolated heart cells

To examine the relationship between mitochondrial NADH (NADHm) and cardiac work output, NADHm and the amplitude and frequency of the contractile response of electrically paced rat heart cells were measured at 25°C. With 5.4 mM glucose plus 2 mM β-hydroxybutyrate, NADHm was reversibly decreased by 23%, and the amplitude of contraction was reversibly decreased by 27% during 4-Hz pacing. With glucose plus 2 mM pyruvate or with 10 mM 2-deoxy-d-glucose, NADHm was maintained during rapid pacing, and the contractile amplitude remained high. Phosphocreatine levels decreased with 2-deoxy-d-glucose administration but not with rapid pacing. Respiration increased to meet the increased ATP demand at 30°C. The data suggest that 1) when NADHm is decreased during rapid pacing with defined substrates, the amplitude of contraction is decreased; 2) the amplitude of contraction during electrical pacing does not change with rate of pacing when both the ATP and NADHm levels are continuously replenished; and 3) the replenishment of NADHm during pacing with physiological substrates may be rate-limited by substrate supply to mitochondrial dehydrogenases. During activation of mitochondrial dehydrogenases, or a significant increase in free ADP induced by 2-deoxy-d-glucose, this rate limitation is bypassed or overcome.

Role of Cardiac KATP Channels During Anoxia and Ischemia

In isolated heart cells, maintained anoxia causes a transient opening of KATP channels. In the ischemic myocardium, this extra K+ conductance results in a decreased contractility and may be arrhythmogenic. Recent studies provide further evidence that the transient activity of KATP channels during anoxia is correlated with the time course of extracellular K+ accumulation during ischemia.

Immunofluorescence localization of the Na-Ca exchanger in heart cells

We investigated the localization of the Na-Ca exchanger in fixed, isolated heart cells from rat and guinea pig using immunocytochemical methods with epifluorescence and confocal microscopy. We found that the Na-Ca exchanger is distributed throughout all membranes in contact with the extracellular space, including the sarcolemma, the transverse tubules (T-tubules), and the intercalated disks. Microscopic nonuniformities in the fluorescent labeling appear to reflect varying views of the membranes containing Na-Ca exchanger protein. Confocal thin-section imaging reveals a regular grid of discrete foci of fluorescence, which represent Na-Ca exchanger in T-tubules viewed en face. These foci are 1.80 +/- 0.01 microns apart from sarcomere to sarcomere and are aligned with the Z-line. Along each Z-line, these foci are spaced at 1.22 +/- 0.11-microns intervals. Longitudinal sections of the sarcolemma-T-tubule junction show a comblike appearance, with T-tubules extending inward from the heavily labeled sarcolemma. Our finding that the Na-Ca exchanger is widely distributed over the cell surface may provide further insight into the role of Na-Ca exchange in the heart.

FMRFamide effects on membrane properties of heart cells isolated from the leech, Hirudo medicinalis

1. The effects of the cardioactive peptide FMRFamide were tested on enzymatically dissociated muscle cells isolated from hearts of the leech. These cells were normally quiescent, with resting potentials near -60 mV. 2. Superfusion of FMRFamide induced a strong depolarization in isolated heart cells (e.g., greater than 40 mV with 10(-6) M FMRFamide). The depolarization was maintained in the continued presence of peptide and persisted long after its removal. Less frequently, FMRFamide superfusion elicited an episodic polarization rhythm. 3. The response of isolated heart cells to bath-applied FMRFamide showed a 1- to 2-min latency. The latency decreased with repeated applications of FMRFamide. 4. The FMRFamide response was diminished by Na+ replacement but persisted with Ca2+ channel blockade. 5. In voltage-clamped heart cells (-60 mv), superfusion of FMRFamide elicited a slow inward current with a transient and a sustained component. 6. Current-voltage (I-V) curves during FMRFamide superfusion in normal leech saline showed that FMRFamide also enhanced voltage-dependent outward currents activated at depolarized levels. 7. Under conditions in which K+ currents were substantially blocked, the FMRFamide-dependent I-V curve was net inward from -90 to +50 mV. A voltage-dependent component was blocked by Co2+ and a linear component by Na+ replacement. 8. We conclude that FMRFamide elicits a persistent inward current with a Na+ component and in addition modulates both voltage-dependent Ca2+ and K+ currents that may contribute to the normal myogenic activity of leech heart muscle cells.