Aims: Cellular excitability is not only determined by the type but also by the number of ion channels in the plasma membrane. Recent evidence indicates that cell surface expression of cardiac pacemaker channels might be controlled beyond the level of biosynthesis by regulating their surface transport. However, neither the underlying trafficking pathways nor their molecular control have yet been investigated.
Methods and results: We have studied endocytic trafficking of hyperpolarization-activated cyclic nucleotide-gated (HCN) ion channels expressed as fusions with green fluorescent protein or tagged with an extracellular haemagglutinin epitope in opossum kidney cells, dissociated rat hippocampal neurons, and ventricular cardiomyocytes. After being internalized from the plasma membrane, HCN2 and HCN4 are sorted to the Rab11-positive endocytic recycling compartment (ERC). From there, they are transported back to the cell surface depending on active phospholipase D2 (PLD2). The peptide hormone angiotensin II, which is upregulated in a number of cardiac pathologies and a known activator of PLD2, stimulates ERC trafficking of HCN4 channels. It significantly increases HCN surface expression independent of their biosynthesis.
Conclusion: Recycling endosomes serve as an intracellular storage compartment for the cardiac pacemaker channels HCN2 and HCN4. They are not only crucial for maintaining a homeostatic surface expression but also supply channels for rapid adaptation of their surface expression in response to extracellular stimuli.