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Development of the fast sodium current in early embryonic chick heart cells
S Fujii1, R K Ayer, R L DeHaan
1Department of Anatomy and Cell Biology, Emory University Health Science Center, Atlanta, Georgia 30322.
Insights
Fast sodium current (INa) in developing chick heart cells increases significantly with age. This age-related increase in INa contributes to the shift in action potential upstroke dependence from calcium to sodium during embryonic development.
Area of Science:
- Cardiovascular Physiology
- Developmental Biology
- Electrophysiology
Background:
- During embryonic development, cardiac action potentials undergo significant changes.
- The ionic mechanisms underlying these changes, particularly the role of fast sodium current (INa), are not fully elucidated.
Purpose of the Study:
- To investigate the developmental changes in fast sodium current (INa) in embryonic chick ventricular cells.
- To determine the contribution of INa to the developmental shift in action potential upstroke dependence.
Main Methods:
- Dissociation and in vitro culture of single ventricle cells from chick embryos at various developmental stages (2, 3, 4, and 7 days).
- Whole-cell patch-clamp electrophysiology to measure fast sodium current (INa).
- Application of voltage-command protocols to assess INa magnitude, voltage dependence, and kinetics.
- Tetrodotoxin (TTX) sensitivity was evaluated.
Main Results:
- Fast sodium current (INa) was detected in 2-day-old embryonic chick ventricular cells and was present in all cells from 3 days onwards.
- INa activation kinetics, voltage dependence, and TTX sensitivity (0.5-1.0 nM) remained consistent across developmental stages.
- Between 2 and 7 days of development, INa inactivation rate doubled, and channel density increased approximately eightfold.
- The resting membrane potential (diastolic potential) shifted to more negative values with development.
Conclusions:
- The limited Na+ dependence of the action potential upstroke on day 2 is due to a depolarized diastolic potential and insufficient INa activation.
- The developmental shift from Ca2+ to Na+ dependence of the action potential upstroke between days 3 and 7 is attributed to both the hyperpolarizing shift in diastolic potential and the increased available Na+ conductance.
Abstract:
Single ventricle cells were dissociated from the hearts of two-, three-, four- or seven-day-old chick embryos, and were maintained in vitro for an additional 6 to 28 hr. Rounded 13 to 18 micron cells with input capacitance of 5 to 10 pF were selected for analysis of fast sodium current (INa). Voltage command protocols designed to investigate the magnitude, voltage dependence, and kinetics of INa were applied with patch electrodes in the whole-cell clamp configuration. INa was present in over half of the 2d, and all 3d, 4d and 7d cells selected. The current showed no systematic differences in activation kinetics, voltage dependence, or tetrodotoxin (TTX) sensitivity with age or culture conditions. Between the 2d and 7d stages, the rate of current inactivation doubled and channel density increased about eightfold. At all stages tested, INa was blocked by TTX at a half-effective concentration of 0.5 to 1.0 nM. We conclude that the lack of Na dependence of the action potential upstroke on the second day of development results from the relatively depolarized level of the diastolic potential, and failure to activate the small available excitatory Na current. The change from Ca to Na dependence of the upstroke during the third to the seventh day of incubation results partly from the negative shift of the diastolic potential during this period, and in part from the increase in available Na conductance.