Age-dependent Expression Changes Of HCN4 And KCNE2 In The Canine Sinoatrial Node

Background:HCN4 (Hyperpolarization-activated Cyclic Nucleotide -gated channels) is the most abundant isoform of the HCN gene family in sinoatrial node. KCNE2 (Mink-related peptide 1, MiRP1) is a member of a family of KCNE and proposed to be a regulatoryβ-subunit of HCN channels. SAN dysfunction increases with age regardless of gender and occurs with peak prevalence in the elderly population. It has been suggested that the age-dependent decline in SAN function is the result of structural remodeling of SANAim:The aim of this study was to study the changes in the expression of HCN4 and KCNE2 and the function of the canine sinoatrial node (SAN) during ageing to investigate the possible mechanism leading to nodal dysfunction.Methods:We assessed the spontaneous sinus rate (IHR), sinus node recovery time (SNRT) and corrected sinus node recovery time (CSNRT) from young (1 to 3 months) to adult (2 to 5 years) and further to aged (8 to 10 years) normal mongrel dogs (n=5) by invasive cardiac electrophysiological study during autonomic blockade. Competitive RT-PCR and Western blot were used to quantify HCN4 and KCNE2 subunit mRNA and protein expression in the sinoatrial node.Results:Histological sections illustrated that pacemaker cells reduced with age and adipose tissue got more and more in SAN. The IHR decreased from 168±11 beats min-1 in the young to 120±9 beats min-1 in the adult and to 88±9 beats min-1 in the aged animals (p<0.05). The SNRT increased from 412±32 ms in the young to 620±56 ms in the adult and to 838±120 ms in the aged animals (p<0.05). The CSNRT increased from 55±12 ms in the young to 117±27 ms in the adult and to 171±37 ms in the aged animals (p<0.05). RT-PCR showed that marked down-regulated HCN4 and KCNE2 mRNA expression in the SAN from young to adult and to aged mongrel dogs. HCN4 mRNA in adult and aged group were reduced by 36% and 51% respectively than young group.KCNE2 mRNA in adult and aged group were reduced by 59% and 72% respectively than young group. Western blot provided verification that HCN4 and KCNE2 protein expression within the SAN declined during aging. Adult and aged dogs SAN protein expression was reduced for HCN4 by 22% and 51% respectively and for the KCNE2 by 18% and 74% compared with young dogs (p<0.05). The SAN showed greater expression of HCN4 mRNA and protein than KCNE2. Expression of ion channel proteins was in agreement with expression of the corresponding mRNAs. Conclusions:The present data show the expression levels of HCN4 and its purportedβ-subunit, KCNE2 changed with age and in parallel with the age-dependent deterioration of SA node function, which provides further evidence of mechanisms underlying the age-related deterioration of the SAN.