Atrial fibrillation (AF) is the most common cardiac arrhythmia with a prevalence of 2-4% in adults (Hindricks et al., 2021). Emerging evidence indicates that autonomic dysfunction is a key driver of atrial arrhythmogenesis. Conditions associated with chronic hypoxia (CH), such as chronic obstructive pulmonary disease and heart failure, are independent risk factors for AF (Grymonprez et al., 2018). In addition to sympathetic activation secondary to peripheral hypoxia sensing, it is possible that CH causes remodelling of left atrial sympathetic neurones, pre-disposing to arrhythmia. At sympathetic terminals, efficacy of noradrenergic outflow is governed by multiple regulatory processes, including the noradrenaline transporter (NAT). As yet, little is known about the regional differences in NAT function in the left atrium and whether this is modified by CH. Here, we have studied NAT kinetics in the left atrial appendage (LAA), left atrial posterior wall (LAPW) and pulmonary veins (PVs) from animals exposed to normoxia (N) and CH.

A recently developed method for dynamic analysis of single-terminal NAT rate (Cao et al. 2020) was employed in atrial tissue obtained from adult male Wistar rats (200-300g). All experiments and procedures were performed in accordance with the UK Animals (Scientific Procedures) Act 1986. Hearts were excised under non-recovery, terminal inhalation isoflurane (3-5% in O₂, flow rate 1.5L min^-1, death by cervical dislocation) and immediately transferred to a superfusion chamber. Atria were dissected free and separated into regions of interest. Tissues were subsequently loaded with a low-concentration solution (1:100 dilution) of Noradrenaline Transporter Assay (NTUA, Molecular Devices), a fluorescent substrate for the NAT, and imaged with confocal microscopy to identify sympathetic terminals. Tissues were then superfused with a 1:20 solution of NTUA, and image stacks were recorded over 15 minutes to monitor uptake rates. Images were analysed using FIJI (v1.53t) to allow quantification of single-terminal NAT activity. Experiments were performed on N (n=6-11; F_iO₂=0.21) and CH (n=3-5; FiO₂=0.12, 9-10 days) animals. Values are expressed as mean ± SEM. Significance was taken as p<0.05, two-way ANOVA with Tukey post-hoc analysis.

Analysis of fluorescence traces revealed a qualitatively similar uptake profile in all 3 regions regardless of CH exposure, demonstrating a linear 6-minute rising slope followed by a plateau phase. CH caused a marked increase in the magnitude of the rising slope in the LAA (N 6.52 ± 0.50% minute^-1 vs CH 10.39 ± 0.66% minute^-1, p=0.001) and LAPW (N 5.93 ± 0.47% minute^-1 vs CH 8.81 ± 0.90% minute^-1, p=0.02), alongside 15-minute plateau fluorescence peaks (LAA N 68.0 ± 4.7% vs CH 102.3 ± 6.7%, p=0.0004; LAPW N 51.3 ± 4.7% vs CH 87.9 ± 6.0%, p=0.003). NAT kinetics were unaffected by CH in the PV (p>0.05).

Our data demonstrate that exposure to CH augments NAT activity in the LAA and LAPW but not in the PV. This could be an adaptive response to a chronic rise in atrial sympathetic nerve firing frequency. Further investigation is required to reveal the mechanism underpinning this upregulation and to what extent this may be of relevance to CH-related atrial arrhythmogenesis.