Mechanisms underlying the diurnal rhythm in sinoatrial node pacemaking

Abstract

Heart rate displays a prominent diurnal rhythm, priming the heart in anticipation of daily patterns of physiological demand. These temporal heart rate changes are regulated by the body’s inherent molecular timekeeping mechanism known as the circadian clock. For >90 years, the diurnal variation in heart rate has been attributed to daily rhythms in autonomic nervous system (ANS) signalling to the sinoatrial node (SAN, the pacemaker of the heart), particularly high vagal tone during sleep. However, there is a lack of mechanistic information regarding the regulation of the diurnal rhythm in heart rate. Whilst autonomic tone has long-been regarded as the key mediator of this, observations of daily rhythms in (i) cardiac ion channel expression and (ii) circadian clock genes in other regions of the heart suggest that there may be an alternative mechanism that is intrinsic to the SAN. In this thesis, I investigated the role and regulation of cardiac ion channels in the regulation of the diurnal rhythm in SAN pacemaking and cardiac excitability. I hypothesised that in mice (that recapitulate cardiovascular diurnal rhythms seen in humans) daily heart rate rhythms occur due to intrinsic diurnal remodelling of SAN coupled-clock transcripts resulting in faster intrinsic pacemaking during the active phase of the animal. The specific contribution of the hyperpolarisation-activated cyclic nucleotide-gated channel 4 (HCN4), an important pacemaking ion channel, in mediating heart rate rhythms was also investigated. A series of approaches were used to explore this, ranging from the whole animal to isolated tissue to the cardiomyocyte nucleus, encompassing electrophysiological recordings, pharmacological manipulation, quantitative polymerase chain reaction, Western blotting, microRNA manipulation, site-directed mutagenesis and luciferase reporter assays were utilised. Using these methods, a number of novel observations were made: (i) There is an intrinsic day-night rhythm SAN pacemaking, which is independent from acute changes in autonomic tone and physical activity; (ii) Several components of the coupled-clock mechanism of the SAN that underlie spontaneous diastolic depolarisation display a diurnal rhythm at the transcript level, appropriate to explain the daily variation in intrinsic heart rate; (iii) Daily rhythms in the expression and function of Hcn4 and If are a key contributor to the day-night variation in SAN pacemaking; (iv) The sympathetic nervous systemic plays a non-canonical role in mediating diurnal rhythms in SAN gene expression to regulate long-range, daily variation in SAN pacemaking. In probing the mechanisms that regulate ion channel transcriptional rhythms, novel roles for (i) the sympathetic nervous system, (ii) tissue-specific transcription factors, (iii) cardiomyocyte chromatin accessibility, and (iv) diurnal microRNA-gene targeting were identified. This work questions the commonly accepted ANS-centric view of the heart rate rhythm, and provides unprecedented new insight into the complex set of rhythmic inputs that regulate diurnal cardiac excitability and SAN pacemaking.

Date of Award	31 Dec 2023
Original language	English
Awarding Institution	The University of Manchester
Supervisor	Delvac Oceandy (Supervisor) & Alicia D'Souza (Supervisor)