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Functional comparison of HCN isoforms expressed in ventricular and HEK 293 cells
Jihong Qu1, Claudia Altomare, Annalisa Bucchi
1Department of Pharmacology and Center for Molecular Therapeutics, Columbia University, 630W 168th St., New York, NY 10032, USA.
Insights
The cellular environment significantly impacts HCN channel function, affecting cardiac rhythm. Neonatal myocytes and HEK 293 cells show distinct HCN2 and HCN4 isoform gating properties.
Area of Science:
- Cardiology
- Molecular Biology
- Biophysics
Background:
- The pacemaker current (I(f)), encoded by the HCN gene family, is crucial for cardiac rhythm.
- I(f) biophysical characteristics, like voltage dependence, vary with cardiac region, development, and disease.
- Heterologous expression of individual HCN isoforms has not fully explained native current functionality.
Purpose of the Study:
- To investigate how the cellular environment influences the gating of HCN channels.
- To compare the functional characteristics of HCN2 and HCN4 isoforms in different cellular contexts.
Main Methods:
- Over-expression of HCN2 and HCN4 isoforms in neonatal myocytes (normal context) and HEK 293 cells (heterologous context).
- Comparison of functional characteristics, including voltage dependence and activation kinetics, between the two cell types.
Main Results:
- HCN4 consistently activates slower than HCN2, with a less negative half-maximum activation voltage, irrespective of cell type.
- Both HCN2 and HCN4 isoforms exhibit more positive activation in myocytes compared to HEK 293 cells.
- Cell type significantly influences HCN voltage dependence, impacting both isoforms similarly.
Conclusions:
- Cellular context, not just inherent isoform properties, dictates HCN channel gating.
- Understanding cell-type specificity is essential for accurately modeling I(f) function in cardiac physiology and disease.
Abstract:
Pacemaker current (I(f)) encoded by the HCN gene family contributes importantly to cardiac rhythm. That contribution depends on the biophysical characteristics of I(f), such as voltage dependence, which vary markedly with cardiac region, development and disease. Heterologous expression studies of individual HCN isoforms have failed to account for the diverse functionality of the native current. To investigate the influence of cellular environment on the gating of HCN channels, we compared the functional characteristics of HCN2 and HCN4, the two major ventricular isoforms, when over-expressed in a normal context (neonatal myocytes) and in a heterologous context (HEK 293 cells). Independent of cell type, HCN4 activates substantially slower than HCN2 and with a half-maximum activation voltage approximately equal 10 mV less negative. However, both isoforms activate more positively in myocytes than in HEK 293 cells. The latter result suggests a context dependence (i.e. cell-type specificity) to HCN voltage dependence that exerts a comparable influence on these two isoforms. This is distinct from the inherent difference in the biophysical properties of HCN2 and HCN4.