Coordinating heart morphogenesis: A novel role for hyperpolarization-activated cyclic nucleotide-gated (HCN) channels during cardiogenesis inXenopus laevis

Abstract

Hyperpolarization-activated cyclic-nucleotide gated channel (HCN) proteins are important regulators of both neuronal and cardiac excitability. Among the 4 HCN isoforms, HCN4 is known as a pacemaker channel, because it helps control the periodicity of contractions in vertebrate hearts. Although the physiological role of HCN4 channel has been studied in adult mammalian hearts, an earlier role during embryogenesis has not been clearly established. Here, we probe the embryonic roles of HCN4 channels, providing the first characterization of the expression profile of any of the HCN isoforms during Xenopus laevis development and investigate the consequences of altering HCN4 function on embryonic pattern formation. We demonstrate that both overexpression of HCN4 and injection of dominant-negative HCN4 mRNA during early embryogenesis results in improper expression of key patterning genes and severely malformed hearts. Our results suggest that HCN4 serves to coordinate morphogenetic control factors that provide positional information during heart morphogenesis in Xenopus.

HCN4 ion channel function is required for early events that regulate anatomical left-right patterning in a Nodal- and Lefty asymmetric gene expression-independent manner

“…To molecularly validate whether HCN4 channels are specifically involved in organizing left-right organ laterality, we used an HCN4-DN (hyperpolarization-activated cyclic nucleotide-gated channel 4-dominant negative) mRNA construct that has been previously molecularly characterized and shown to inhibit the HCN4 channel current in mammalian cell culture by direct electrophysiology ( Pitcairn et al, 2017 ). We confirmed that expression of HCN4-DN protein blocks HCN4 channel function and depolarizes the membrane voltage in Xenopus embryos ( …”

“…Here we extend our recent work on HCN4 channels in cardiogenesis ( Pitcairn et al, 2017 ) using a different set of misexpression conditions to target earlier events. We show that HCN4 channels are present in Xenopus embryos beginning at the first cleavage event (2-cell stage).…”

“…Our recent study explored HCN4 function in embryonic cardiac tissue development ( Pitcairn et al, 2017 ). That study and the experiments presented here used drastically different injection and culture conditions to target two different lineages/tissues: the animal cap versus the mesodermal heart lineage.…”

“…They open at hyperpolarized membrane voltages (negative), giving rise to currents which are a mix of sodium and potassium ion fluxes. Among the four HCN isoforms (HCN1-4), HCN4 channels have been primarily studied in adult hearts as pacemaker channels ( Scicchitano et al, 2012 ; Verkerk and Wilders, 2015 ), but recent evidence has demonstrated their presence in human and mouse embryonic cells ( Cerbai et al, 1999 ; Qu et al, 2008 ; Robinson et al, 1997 ; Später et al, 2013 ; Vicente-Steijn et al, 2011 ; Yasui et al, 2001 ), and implicated them in cardiac patterning ( Pitcairn et al, 2017 ). Because these channels have not been studied in the context of developmental bioelectricity ( Adams and Levin, 2013 ; Levin, 2013 , 2014a ,b ) or control of embryonic axial patterning, we characterized the role of HCN4 channels in embryonic left-right asymmetry establishment in Xenopus embryos.…”

Correlation between HCN4 gene polymorphisms and lone atrial fibrillation risk

“…As is well known, HCN family plays an important role in controlling neuronal and cardiac excitability. HCN4 serves as a pacemaker channel, which could maintain periodicity of contractions in vertebrate hearts [29]. Rs498005 SNP might influence the activity of HCN4 gene, thus, contributing to initiation of AF.…”