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Novel Potassium Channels in Kidney Mitochondria: The Hyperpolarization-Activated and Cyclic Nucleotide-Gated HCN

Daniel León-Aparicio¹, Carolina Salvador², Omar Emiliano Aparicio-Trejo³

¹Departamento de Fisiología, Facultad de Medicina, Universidad Nacional Autónoma de México (UNAM), Mexico City 04510, Mexico. dalebx4@gmail.com.

International Journal of Molecular Sciences

|October 12, 2019

Insights

Pacemaker hyperpolarization-activated cyclic nucleotide-gated (HCN) channels, specifically HCN3, are found in kidney mitochondria. These channels facilitate potassium transport, supporting respiratory chain activity and ATP synthesis.

Area of Science:

Cellular and Molecular Biology
Renal Physiology
Mitochondrial Function

Background:

Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels (HCN1-4) regulate neuronal excitability and ion transport.
In the kidney, HCN1, HCN2, and HCN3 channels are involved in sodium, potassium (K+), and ammonium transport.
HCN3 expression in the kidney is influenced by potassium diets.

Purpose of the Study:

To investigate the role of HCN channels in kidney mitochondria.
To determine if HCN channels are present and functional within renal mitochondria.
To elucidate the impact of HCN channel activity on mitochondrial bioenergetics.

Main Methods:

Proteomic analysis of HCN3 in human embryonic kidney (HEK293) cells.
Immunoblotting and immunogold electron microscopy to detect HCN3 in rat and human kidney mitochondria.
Patch-clamp recordings of mitochondria from renal tissues and HEK293 cells overexpressing HCN channels, coupled with MitoTracker Green FM staining.
Assessment of oxygen consumption, ATP synthesis, and inner mitochondrial membrane potential following ZD7288 treatment.

Main Results:

Proteomic analysis revealed significant mitochondrial association for HCN3 interacting proteins.
HCN3 protein expression was confirmed in both rat and human kidney mitochondria.
Patch-clamp electrophysiology demonstrated inwardly rectifying K+ currents in mitochondria, exclusively mediated by HCN3 and inhibited by ZD7288.
Pharmacological blockade of HCN channels with ZD7288 inhibited mitochondrial oxygen consumption, ATP synthesis, and hyperpolarized the inner mitochondrial membrane.

Conclusions:

This study identifies pacemaker HCN channels, particularly HCN3, as functional components within kidney mitochondria.
HCN3 channels facilitate K+ transport across the inner mitochondrial membrane.
Mitochondrial HCN3 activity is crucial for maintaining the inner mitochondrial membrane potential, thereby supporting respiratory chain function and ATP production.

Read full analysis

Abstract:

Hyperpolarization-activated cationic HCN channels comprise four members (HCN1-4) that control dendritic integration, synaptic transmission and action potential firing. In the kidney, HCN1, HCN2 and HCN3 are differentially expressed and contribute to the transport of sodium, potassium (K⁺) and ammonium into the nephrons. HCN3 is regulated by K⁺ diets in the kidney. In this work we performed a proteomic analysis of HCN3 expressed in human embryonic kidney cells (HEK293 cells). More than 50% of the interacting proteins belonged to mitochondria. Therefore, we explored the presence of HCN channels in kidney mitochondria. By immunoblotting and immunogold electron microscopy HCN3 protein expression was found in rat kidney mitochondria; it was also confirmed in human kidney. Patch-clamp recordings of renal mitochondria and mitochondria from HEK293 cells overexpressing HCN1, HCN2 and HCN3 channels, stained with MitoTracker Green FM, indicated that only HCN3 could produce inwardly K⁺ currents that were inhibited by ZD7288, a specific blocker of HCN channels. Furthermore, ZD7288 caused inhibition of the oxygen consumption coupled to ATP synthesis and hyperpolarization of the inner mitochondrial membrane. In conclusion, we show for the first time that pacemaker HCN channels contribute to K⁺ transport in mitochondria facilitating the activity of the respiratory chain and ATP synthesis by controlling the inner mitochondrial membrane potential.

Keywords:

hyperpolarization-activated cyclic nucleotide-gated cationic HCN channels in mitochondria kidney mitochondria mitochondria uncoupling mitochondrial potassium channel potassium transport in mitochondria

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