Transient cardiac electrophysiological changes in a rat model of subarachnoid haemorrhage: a brain–heart interaction

Abstract

Aims

Subarachnoid haemorrhage (SAH) is one of the causes of sudden cardiac death (SCD). However, the time course of ventricular arrhythmias and potential mechanisms responsible for this effect after SAH remain unknown.

Objective

This study aims to investigate the effect of SAH on ventricular electrophysiological changes and its potential mechanisms in long-term phase.

Methods and results

We examined the ventricular electrophysiological remodelling and potential mechanisms in a Sprague Dawley rat model of SAH at six time points (baseline, and Days 1, 3, 7, 14 and 28) and explored the potential mechanisms. We measured the ventricular effective refractory period (ERP), ventricular fibrillation threshold (VFT) and left stellate ganglion (LSG) activity at different time points before and after SAH. We also detected neuropeptide Y (NPY) levels in plasma and myocardial tissues by enzyme-linked immunosorbent assay, and quantified NPY 1 receptor (NPY1R) protein and mRNA expression levels by western blotting and quantitative real-time reverse transcription-polymerase chain reaction, respectively. Subarachnoid haemorrhage gradually prolonged QTc intervals, shortened ventricular ERP and reduced VFT during the acute phase, peaking at Day 3. However, no significant changes were observed from Days 14 to 28 compared to Day 0. Subarachnoid haemorrhage gradually increased LSG activity, increased NPY concentrations and up-regulated NPY1R expression in the acute phase of SAH, peaking at Day 3. However, no significant variations were found from Days 14 to 28 compared to Day 0.

Conclusion

Subarachnoid haemorrhage increases the transient susceptibility of VAs in the acute phase, and the underlying mechanisms for this response included increased sympathetic activity and up-regulated NPY1R expression.

Graphical Abstract

Graphical Abstract.

What’s new?

• Subarachnoid haemorrhage (SAH) is a known cause of sudden cardiac death. Our study proves the brain–heart interaction with a focus on ventricular arrhythmias (VAs).

• This study expands on previous research by examining the time course of VAs during the acute phase of SAH.

• Our study is the first to demonstrate that SAH increases the transient susceptibility of VAs in the acute phase. We identified the underlying mechanisms behind this response, which include increased sympathetic activity and up-regulated NPY1R expression.

Introduction

The brain and heart are closely connected,¹ with the brain controlling the heart's normal function directly through the autonomic nervous system. An imbalanced brain–heart interaction could have negative consequences for the heart. Individuals who have experienced brain damage following acute stroke are particularly vulnerable to severe cardiac adverse events.² Subarachnoid haemorrhage, a life-threatening type of stroke, increases the risk of malignant ventricular arrhythmias (VAs) and even leads to sudden cardiac death. Subarachnoid haemorrhage (SAH) patients with VAs have a significantly higher mortality rate and worse functional outcomes than those without VAs. Malignant VAs are more frequently detected during the acute phase of SAH.³

Previous studies have indicated that significant cardiac arrhythmias, including complex ventricular premature contractions and ventricular tachycardia, are highly prevalent during the first 72 h after SAH.⁴ However, significant VAs are rarely observed in later stages, suggesting that severe arrhythmias are responsible for the deaths of some patients either before a definitive diagnosis of SAH is made or early after admission to the neurosurgical unit. Currently, few experimental studies have been conducted to explore the time course and mechanisms of VAs caused by brain–heart crosstalk. It is important to determine the time course of electrophysiological changes triggered by SAH. The primary goal of this study was to evaluate the time profile of ventricular electrophysiological instability and its potential mechanistic effects on brain–heart dysregulation of cardiac electrophysiological properties after SAH in a rat model. Investigating the role of the brain–heart interaction in electrophysiological properties after stroke is highly clinically significant.

Materials and methods

Animal preparation

All experimental procedures in this study were conducted in accordance with the ethical standards and guidelines set forth by the Animal Ethics Committee of Wuhan University and Animal Care and Use Committee of Renmin Hospital of Wuhan University. The study was carried out in compliance with the National Institutes of Health's guidelines for the care and use of laboratory animals (NIH Publication, revised 2011). Male Sprague Dawley rats, aged 11–12 weeks and weighing between 211 and 296 g, were obtained from Hunan SJA Laboratory Animal Co., Ltd. and housed in a specific pathogen-free laboratory under animal barrier conditions. Prior to the start of the study, the rats were acclimatized to a 12/12 h light/dark cycle and provided with ad libitum access to food and water for 1 week. All rats were anaesthetized with 80 mg/kg sodium pentobarbital administered intraperitoneally and euthanized by an overdose of sodium pentobarbital.

Experimental protocol

Sprague Dawley rats were randomly divided into two groups: Control group and SAH group. Furthermore, both groups were divided into subgroups at different time points, respectively. The Control group was divided into six subgroups including Day 0 (n = 13), Day 1 group (n = 19), Day 3 group (n = 19), Day 7 group (n = 19), Day 14 group (n = 19) and Day 28 group (n = 19). In the Control group, all rats received an injection of 0.1 mL of physiological saline into the cisterna magna following the withdrawal of cerebrospinal fluid. The SAH group was divided into six subgroups as follows: Day 0 (n = 20), Day 1 group (n = 25), Day 3 group (n = 25), Day 7 group (n = 25), Day 14 group (n = 25) and Day 28 group (n = 25). In the SAH group, all rats received an injection of 0.1 mL autologous arterial blood into the cisterna magna after cerebrospinal fluid withdrawal. All rats were anaesthetized with sodium pentobarbital (30 mg/kg) and attached to a positive pressure ventilator (Anhui Zheng Hua Biological Instrument Co., Ltd, Anhui, China). Electrocardiogram (ECG) recordings were obtained by placing needle electrodes under the limb skin, and the standard limb lead-II was consecutively recorded using a Power Lab data acquisition system. During the whole experiment, the standard limb lead-II was consecutively recorded using a Power Lab data acquisition system (AD Instruments, Bella Vista, Australia). Electrocardiogram parameters (QT interval, QTc interval and RR interval) were measured and averaged at Day 0, Day 1, Day 3, Day 7, Day 14 and Day 28 in both groups. Additionally, a 24-gauge catheter was placed into the right carotid artery to detect blood pressure and withdrawn autologous arterial blood. A thoracotomy at the third intercostal space was used to measure effective refractory period (ERP) measurement and detect left stellate ganglion (LSG) activity detection.

Subarachnoid haemorrhage model induction

The SAH rat model was established using a previously described method. To avoid head movement, a head holder with 30° flexion was utilized to position the rat’s head. The atlanto-occipital membrane was exposed for subarachnoid injection, and a 27-gauge needle was used to puncture the cisterna magna. A 0.1 mL sample of cerebrospinal fluid was withdrawn and discarded, and an equal volume (0.1 mL) of physiological saline or autologous arterial blood was slowly injected into the cisterna magna over a 10-min period. After the injection, the wound was closed, and the rat's head was turned down for 10 min. Rats were allowed to awaken following the procedure.⁵

Measurement of ventricular effective refractory period

A custom-made Ag-AgCl catheter was inserted into the left and right ventricular free walls to measure ventricular ERP at four sites: left ventricular base, left ventricular apex, right ventricular base, and right ventricular apex. The ERP at each site was measured using programmed pacing, which involved eight consecutive stimuli (S1–S1 = 150 ms) followed by a premature stimulus (S2). The S1–S2 interval was progressively shortened by 10 ms and then by 1 ms, starting at 120 ms. The longest S1–S2 interval that failed to capture the ventricle was defined as ERP. The ERP dispersion was determined as the coefficient of variation (standard deviation/mean of ERP) at all four sites, as previously described.⁶ Two researchers who were blinded to the animal group conducted the ERP measurements.

Ventricular fibrillation threshold measurement

Bipolar stimulating electrodes with a diameter of 1 mm and an interelectrode distance of 5 mm made of platinum–iridium were positioned at the right ventricle. The heart was continuously stimulated with rectangular pulses (50 Hz frequency, 4 ms impulse length) for 5 s to determine the ventricular fibrillation threshold (VFT). After each stimulation, at least 30 s was allowed to elapse before administering the next stimulus unless ventricular fibrillation occurred. The stimulating voltage was initially set at 2 V and increased in increments of 1 V until VF occurred. The VFT was defined as the minimum voltage required to produce sustained VT/VF (≥3 s) on three occasions.⁷ Electrocardiograms with arrhythmias were analysed and evaluated by two researchers who were unaware of the animal group.

Measurement of left stellate ganglion neural activity

In each group, a 1 min recording of LSG activity was obtained. The tungsten-coated microelectrode was inserted into the fascia of the LSG, and the chest wall was connected to a group lead. The LSG-generated electrical signals were recorded using a Power Lab data acquisition system (8/35, AD Instruments, New South Wales, Australia) and an amplifier (DP-304, Warner Instruments, Hamden, CT). Prior to recording, bandpass filters (300–1 kHz) were established and amplification was set to 30–50 times. Neural activity was recorded based on frequency and amplitude, with deflections having a signal-to-noise ratio >3:1 defined as neural activity. The amplitude and frequency were manually determined, as previously described in our studies.⁸

Enzyme-linked immunosorbent assay analysis of neuropeptide Y

Blood samples were collected from the jugular vein. The rat hearts were dissected after ventricular electrophysiology detection. The samples were stored at −80°C until use. The NPY levels in both the plasma and heart samples were determined using a general NPY ELISA kit (Nanjing Jiancheng Bioengineering Institute, Nanjing, China) following the manufacturer’s instructions. The absorbance was measured using a spectrophotometer at a wavelength of 450 nm, and a standard curve was constructed for each plate. The NPY concentrations were analysed in duplicate, and the intra-assay variation was less than 9%. The NPY values were expressed as pg/mL, as described in previous studies.⁷

Western blot of neuropeptide Y 1 receptor

The NPY1R protein expression was analysed by immunoblotting. Left ventricular tissues were collected and immediately frozen in liquid nitrogen, and then stored at −80°C. The membranes were incubated overnight at 4°C with anti-NPY1R antibody (diluted 1:1000, from Servicebio Biotechnology Co., Ltd, Wuhan, China) and then incubated with secondary antibodies (from Servicebio Biotechnology Co., Ltd, Wuhan, China) for 1 h at room temperature in the dark. An anti-GAPDH antibody (mouse polyclonal, 1:1000 dilution; Abcam, Cambridge, UK) was used as an internal control. Chemiluminescence was used to detect the signals, which were then quantified using video densitometry. The band intensities were normalized to GAPDH expression.

Quantitative real-time reverse transcription-polymerase chain reaction

The RNA extraction protocol used TRIzol reagent, and SYBR Green was used for real-time polymerase chain reaction (PCR) analysis to determine NPY levels with GAPDH as an internal control on the QuantStudio™ 6 Flex Real-Time PCR System (ABI, USA). A segment of the rat NPY1R cDNA sequence was targeted with the forward primer 5′-ACGTTCGCTTGAAAGGAGA-3′ and the reverse primer 5′- CATGACGTTGATTCGTTTGG-3′. A segment of the rat GAPDH sequence was amplified with the upstream primer 5′-TGGTATCGTGGAAGGACTCAT-3′ and the downstream primer 5′-GTGGGTCGCTGTTGAAGTC-3′. The target mRNA quantity was obtained using the 2-ΔΔCT method and normalized to an endogenous reference, with values presented relative to a calibrator.

Statistical analysis

All continuous variables are presented as the mean ± standard deviation (SD). The comparison between data from two groups that obey a normal distribution measurement was conducted by Student's t-test. The multi-group comparisons among groups were analysed using one-way analysis. The incidence of VF between two groups was compared using Fisher’s exact test. Statistical analysis was performed using SPSS 22.0 software (IBM Corp, New York, USA). P < 0.05 was considered statistically significant.

Results

All rats in every group were subjected to the entire experimental protocol.

Effects of subarachnoid haemorrhage on delayed repolarization over time

The results show that delayed repolarizations were present in the acute phase of SAH from Day 1 to Day 7, with a peak at Day 3 (Figure 1). The QT intervals of Day 1, Day 3 and Day 7 in the SAH groups were significantly prolonged compared to Day 0 (all P < 0.05). However, the QT intervals in Day 14 and Day 28 were not significantly different from those of Day 0 in the SAH groups. Additionally, the QT intervals of Day 1, Day 3 and Day 7 in the SAH groups were significantly prolonged compared to the respective Control groups (all P < 0.05). The QTc interval (calculated as QT interval divided by the square root of RR interval) in Day 1, Day 3 and Day 7 was significantly prolonged in the SAH groups (all P < 0.05), but there was no significant difference in the QTc interval among Day 14 and Day 28, and Day 0 groups. Similarly, the QTc of Day 1, Day 3 and Day 7 in the SAH groups were significantly prolonged compared to the respective subgroups in the Control groups (all P < 0.05). There were no significant differences in RR intervals at different time points or between Control and SAH groups. It indicated that the prolonged QT intervals and QTc intervals caused by SAH returned to baseline at Day 14.

Figure 1.

Delayed repolarization in subarachnoid haemorrhage (SAH) rats. (A) Representative electrocardiogram showing QT intervals in Day 0 and Day 3 rats in the SAH groups. (B) QT intervals were significantly prolonged in SAH rats during the acute phase period (at Day 1, Day 3 and Day 7) but not changed in the late phase period (at Day 14 and Day 28) compared to Day 0. Additionally, the QT intervals of Day 1, Day 3 and Day 7 in the SAH groups were significantly prolonged compared to the respective Control groups. (C) QTc interval showed similar trend. (D) There were no significant differences in RR intervals at different time points or between Control and SAH groups. SAH, subarachnoid haemorrhage.

Effects of subarachnoid haemorrhage on the ventricular effective refractory period over time

Figure 2 shows that ventricular ERP was significantly shortened in the acute phase of SAH from Days 1 to 7, with a peak at Day 3. The ventricular ERPs at various sites in Day 1, Day 3 and Day 7 were significantly shortened compared to Day 0 in the SAH groups (all P < 0.05). However, the ventricular ERPs in Day 14 and Day 28 were not significantly different from Day 0 in the SAH groups. Additionally, the ERPs of Day 1, Day 3 and Day 7 in the SAH groups were significantly shortened compared to the respective Control groups (all P < 0.05).

Figure 2.

Effects of subarachnoid haemorrhage (SAH) on ventricular effective refractory period (ERP) and ERP dispersion at four sites of ventricular surface in SAH rats. (A) Representative electrocardiogram showing and ventricular ERP stimulating programme. (B) Effective refractory period at left ventricular base were significantly shortened in SAH rats during the acute phase period (at Day 1, Day 3 and Day 7) but not changed in the late phase period (at Day 14 and Day 28) compared to Day 0. Additionally, the ERPs of Day 1, Day 3 and Day 7 in the SAH groups were significantly shortened compared to the respective Control groups. (C) ERP at left ventricular apex showed similar trend. (D) Effective refractory period at right ventricular base showed similar trend. (E) Effective refractory period at right ventricular apex showed similar trend. (F) Effective refractory period dispersion were significantly increased in SAH rats during the acute phase period (at Day 1, Day 3 and Day 7) but not changed in the late phase period (at Day 14 and Day 28) compared to Day 0. Similarly, the ERPs of Day 1, Day 3 and Day 7 in the SAH groups were significantly increased compared to the respective Control groups. ECG, electrocardiogram; ERP, effective refractory period; LVB, left ventricular base, RVA, right ventricular apex, RVB, right ventricular base, SAH, subarachnoid haemorrhage.

The ERP dispersion reflects the susceptibility of VAs. The ventricular ERP dispersions in Day 1, Day 3 and Day 7 were significantly increased compared to Day 0 in the SAH groups (all P < 0.05). However, ventricular ERP dispersions in Day 14 and Day 28 were not significantly different from Day 0 in the SAH groups. Similarly, the ERPs of Day 1, Day 3 and Day 7 in the SAH groups were significantly increased compared to the respective Control groups (all P < 0.05). These results suggest that the shortened ERPs and increased ERP dispersions caused by SAH returned to baseline at Day 14.

Effects of subarachnoid haemorrhage on the ventricular fibrillation threshold over time

The representative images (Figure 3A) show VT/VF induced by burst stimulation. The incidence of VT/VF was evident from Day 1 to 7 compared to Day 0 in the SAH group (all P < 0.05). However, there were no significant differences in the incidence of VT/VF during the late phase of SAH (Day 14 and 28) compared to Day 0. Furthermore, the incidence of VT/VF on Day 1, Day 3 and Day 7 was significantly higher in the SAH groups than in their respective Control groups (all P < 0.05). The VFT was significantly shortened in the acute phase of SAH from Day 1 to 7. The VFT was significantly lower in the SAH groups on Day 1, Day 3 and Day 7 compared to Day 0 (all P < 0.05), but there were no significant differences on Day 14 and Day 28. Similarly, VFT on Day 1, Day 3 and Day 7 in the SAH groups was significantly lower than in their respective Control groups (all P < 0.05). These results suggest that SAH increases the susceptibility to VF in the acute phase, but this effect returns to baseline by Day 14.

Figure 3.

Effects of subarachnoid haemorrhage (SAH) on ventricular fibrillation threshold (VFT) variations in SAH rats. (A) Representative electrocardiogram with burst stimulating on heart. (B) Ventricular tachycardia/ventricular fibrillation (VT/VF) incidence were significantly increased in SAH rats during the acute phase period (at Day 1, Day 3 and Day 7) but not changed in the late phase period (at Day 14 and Day 28) compared to Day 0. Furthermore, the incidence of VT/VF on Day 1, Day 3 and Day 7 was significantly higher in the SAH groups than in their respective Control groups. (C) Ventricular fibrillation threshold was significantly decreased in SAH rats during the acute phase period (at Day 1, Day 3 and Day 7) but not changed in the late phase period (at Day 14 and Day 28) compared to Day 0. Similarly, VFT on Day 1, Day 3, and Day 7 in the SAH groups was significantly lower than in their respective Control groups. VT, ventricular tachycardia; VF, ventricular fibrillation.

Effects of subarachnoid haemorrhage on left stellate ganglion neural activity over time

Figure 4A shows representative examples of LSG neural activity recordings at different time points in subgroups of both Control and SAH groups. The frequency and amplitude of LSG neural activity were significantly increased in Day 1, Day 3 and Day 7 compared to Day 0 in the SAH groups (all P < 0.05). However, in Day 14 and Day 28, the frequency and amplitude of LSG neural activity were not significantly different from Day 0 in the SAH groups. Moreover, the frequency and amplitude of LSG neural activity in Day 1, Day 3 and Day 7 in the SAH groups were significantly increased than the respective Control groups (all P < 0.05). These results indicate that SAH increases cardiac sympathetic nervous activity in the acute phase, but this effect returns to baseline by Day 14.

Figure 4.

Effects of subarachnoid haemorrhage (SAH) on left stellate ganglion (LSG) activity measurements. (A) Representative LSG activity among six groups. (B) Frequency of LSG activity were significantly increased in SAH rats during the acute phase period (at Day 1, Day 3 and Day 7) but not changed in the late phase period (at Day 14 and Day 28) compared to Day 0. Additionally, the frequency and amplitude of LSG neural activity in Day 1, Day 3 and Day 7 in the SAH groups were significantly increased than the respective Control groups. (C) Amplitude of LSG activity showed similar trend. SAH, subarachnoid haemorrhage.

Time course of neuropeptide Y levels in plasma and heart after subarachnoid haemorrhage

Figure 5 illustrates that in the acute phase of SAH, from Days 1 to 7, there was a significant increase in NPY levels in both plasma and heart tissues, peaking at Day 3. The NPY levels in the SAH groups on Day 1, Day 3 and Day 7 were significantly higher compared to Day 0 (all P < 0.05). However, there were no significant differences in NPY levels in Day 14 and Day 28 groups compared to Day 0 in the SAH groups. Additionally, the NPY levels in plasma and heart on Day 1, Day 3 and Day 7 in the SAH groups were significantly higher compared to the respective Control groups (all P < 0.05). These findings suggest that SAH increases NPY expression in the acute phase, but this effect returns to baseline by Day 14.

Figure 5.

Effects of subarachnoid haemorrhage (SAH) on plasma and heart detected neuropeptide Y (NPY) concentrations. (A) Plasma NPY concentrations were significantly increased in SAH rats during the acute phase period (at Day 1, Day 3 and Day 7) but not changed in the late phase period (at Day 14 and Day 28) compared to Day 0. Additionally, the NPY levels in plasma and heart on Day 1, Day 3 and Day 7 in the SAH groups were significantly higher compared to the respective Control groups. (B) Myocardial NPY concentrations showed similar trend. NPY, neuropeptide Y.

Time course of ventricular neuropeptide Y receptor level after subarachnoid haemorrhage

Figure 6 depicts that there was a significant increase in NPY1R protein and mRNA expression in heart tissues during the acute phase of SAH (Days 1–7), with the peak expression observed on Day 3. The NPY1R protein and mRNA expression in the SAH groups on Day 1, Day 3 and Day 7 were significantly higher compared to Day 0 (all P < 0.05). However, there were no significant differences in NPY1R protein and mRNA expression on Day 14 and Day 28 compared to Day 0 in the SAH groups. Moreover, the NPY1R protein and mRNA expression in heart tissues on Day 1, Day 3 and Day 7 in the SAH groups were significantly higher compared to the respective Control groups (all P < 0.05). These results suggest that SAH increases NPY1R protein and mRNA expression in ventricular tissues during the acute phase, but this effect returns to baseline by Day 14.

Figure 6.

Effects of subarachnoid haemorrhage (SAH) on myocardial neuropeptide Y 1 receptor (NPY1R) expression. (A) Representative western blot of heart NPY1R in Control and SAH groups. (B) Neuropeptide Y 1 receptor protein expression by western blot analysis were significantly increased in SAH rats during the acute phase period (at Day 1, Day 3 and Day 7) but not changed in the late phase period (at Day 14 and Day 28) compared to Day 0. (C) mRNA expression of NPY1R showed similar trend. Moreover, the NPY1R protein and mRNA expression in heart tissues on Day 1, Day 3 and Day 7 in the SAH groups were significantly higher compared to the respective Control groups. NPY1R, Neuropeptide Y 1 receptor; SAH, subarachnoid haemorrhage.

Discussion

Major findings

In this study, we investigated the impact of SAH on ventricular electrophysiological properties and its potential mechanisms over time. The major findings included that (i) SAH altered ventricular electrophysiological properties in the acute phase, and peaked at Day 3; (ii) SAH significantly overactivated cardiac sympathetic nervous activity in acute phase, recovering at Day 14; and (iii) the mechanisms of SAH on ventricular electrophysiology potentially occur via the NPY/NPY1R axis. These results provide important insights for clinicians on the optimal timing and strategies for preventing VAs in patients with SAH. Furthermore, they suggest a potential therapeutic target for treating SAH-induced VAs in the acute phase.

Brain and heart interaction: cardiac electrophysiological properties

Cardiac damage and arrhythmias are common occurrences in acute stroke patients, even in the absence of primary heart disease, and are grouped clinically under the term ‘stroke-heart syndrome’.^9,10 Aneurysmal SAH is a well-known cause of sudden cardiac arrest,¹¹ and a Danish study found that SAH was responsible for 11.1% of neurogenic cardiac deaths and 0.6% of overall SCDs.¹² Subarachnoid haemorrhage can also lead to out-of-hospital cardiac arrest in 4% of patients. Therefore, identifying the cause of cardiac arrest is crucial for guiding appropriate management. Recently, Lanzino et al. reported that 91% of patients with nontraumatic SAH had evidence of cardiac abnormalities (atrial and VAs) observed on ECG.¹³ Subarachnoid haemorrhage serves as an excellent model to study heart–brain interactions due to the high incidence of arrhythmias. Stroke can cause prolongation of the QT interval on an ECG. Prolonged QT interval can cause Torsade de Pointes, a type of ventricular tachycardia that can degenerate into VF and cause SCD.^14,15 In this study, we established a direct link between the brain and heart and found that SAH induced QT interval prolongation and shortened VFT in the acute phase. These findings suggest that SAH in the acute phase increases the instability of ventricular electrophysiology and predisposes patients to develop VAs.

The potential mechanisms of subarachnoid haemorrhage on ventricular electrophysiological property changes

After a stroke, several potential mechanisms, including sympathetic overactivation, alterations in the hypothalamic–pituitary–adrenal (HPA) axis, and a catecholamine surge, may mediate cardiac dysfunction.¹⁶ Brain injury can activate sympathetic tone and increase the release of catecholamines,¹⁷ which contribute to myocardial damage and the development of arrhythmias in the acute phase of SAH. In this study, we directly observed that cardiac sympathetic activity was significantly increased during the acute phase of SAH.

Activation of the HPA axis can cause a surge in catecholamines, which act on β receptors, activate G protein, increase cytosolic cAMP, regulate protein kinase A subunits and ultimately influence membrane potentials.¹⁸ The β-cAMP-PKA pathway mediates the activation of L-type Ca²⁺ channels, increases Ca²⁺ influx and triggers the release of Ca²⁺ from the sarcoplasmic reticulum. Dysfunction in Ca²⁺ handling can lead to afterdepolarizations and electrical instability. However, in addition to the effects of catecholamines, NPY released from sympathetic nerves may also increase rates of VAs. Kalla et al. recently reported that NPY acts on Y1 receptors and predisposes to VAs in ST-elevation myocardial infarction, despite beta-blocker treatment.¹⁹ The NPY shortens the duration of the calcium transient and lowers the VFT. Similarly, Hoang et al. demonstrated that high-frequency stimulation of sympathetic nerves increases NPY release and shortens ventricular action potential duration.²⁰ Both studies provide evidence that NPY, released from sympathetic terminals, contributes to VAs.

In this study, we observed an increase in cardiac sympathetic tone and elevated plasma, and heart NPY concentrations during the acute phase of SAH. We also observed up-regulation of the NPY1R in left ventricular tissues during this phase, which gradually disappeared in the late phase of SAH. This result suggests that SAH alters ventricular electrophysiological properties, potentially by increasing cardiac sympathetic nervous system activity and the NPY/NPY1R axis.

Time course of arrhythmia onset during the acute phase of subarachnoid haemorrhage

Previous studies have shown that significant VAs, including SCD, often occur within the first 24 h in patients with acute stroke.²¹ However, Klingelhofer et al. demonstrated that cardiac autonomic dysfunction after stroke appears with temporal latency and indicates cardiac arrhythmias beyond the first 24 h.⁴ Furthermore, both clinical data and experimental studies on the time profile of arrhythmia onset during the acute or late phase are extremely scarce. Prosser et al. reported that the peak of serious cardiac adverse events occurred between Days 2 and 3.¹¹ However, currently, there was no study to explore the time course of SAH on VAs.

In the present study, we observed a transient change in ventricular ERP and VFT shortening in the early phase of SAH, which was most prominent at Day 3 after SAH. We also found that significant VAs and VFT decreases occurred during the early phase of SAH, and the greatest risk of this adverse event occurred on Day 3. Interestingly, all of these effects caused by SAH returned to baseline in chronic phase. These data therefore provide evidence that cardiac monitoring should be extended to the early phase of SAH to screen for VAs. Naredi et al. showed a dramatic elevation in sympathetic nervous activity that persisted for at least 7 days.²² The marked sympathetic activation may contribute to cardiac disturbances. In the present study, we found that SAH increased cardiac sympathetic nervous activity during the acute phase, but not in the late phase.

Clinical application

The findings of this study have important clinical implications for the management of patients with SAH. Given the transient change in ventricular electrophysiological properties observed in this study, it is recommended that cardiac monitoring be extended beyond the first 24 h of SAH to screen for VAs, which may not be detected during routine clinical assessment. This can help to improve early detection and management of cardiac complications in patients with SAH.

Furthermore, the observed increase in cardiac sympathetic activity and serum levels suggests that interventions targeting the sympathetic nervous system, such as LSG block or NPY antagonism, may represent potential therapeutic strategies for the prevention and treatment of malignant VAs in patients with SAH. Further studies are needed to investigate the safety and efficacy of these interventions in this patient population.

Overall, the findings of this study highlight the importance of close cardiac monitoring and consideration of targeted interventions to reduce the risk of adverse cardiac events in patients with SAH.

Translational perspective

This study provides valuable insights into the exact time course of VAs and the underlying mechanisms in patients with SAH. These findings have important clinical implications. Understanding the temporal changes in ventricular electrophysiology after SAH can aid in the development of appropriate monitoring and management strategies during the acute phase, particularly around Day 3, when VAs are most likely to occur. Monitoring QTc intervals, autonomic activity and NPY levels could help identify patients at higher risk of developing VAs in this critical period. Moreover, the study highlights the involvement of sympathetic activity and NPY1R in the mechanisms underlying VAs after SAH. Targeting these pathways could offer potential therapeutic avenues for preventing or mitigating arrhythmic events in SAH patients. Modulating sympathetic activity or NPY1R expression may hold promise for improving outcomes and reducing the risk of SCD in this patient population. These findings pave the way for future research and clinical interventions aimed at improving risk stratification, monitoring and treatment strategies for patients with SAH-associated cardiac complications.

Study limitations

The present study has some limitations that should be acknowledged. Firstly, all experimental animals were under anaesthesia, which may have influenced cardiac autonomic activity. Nonetheless, all animal experiments were performed under controlled anaesthetic conditions. Secondly, although we observed that SAH caused temporary changes in ventricular electrophysiological properties, the exact underlying mechanisms require further investigation. Thirdly, previous studies have suggested that NPY, a co-transmitter released by the sympathetic terminal, plays a critical role in the occurrence of VAs. While our study supports the notion that NPY may contribute to the changes in ventricular electrophysiological properties, we did not use patch-clamp electrophysiology to measure the electrical properties of myocytes induced by NPY.

Conclusions

In conclusion, our study demonstrates that SAH leads to transient changes in ventricular electrophysiological properties and increases the risk of malignant VAs. This effect may be due to the activation of the LSG and overexpression of NPY. These findings shed light on the underlying mechanisms of brain–heart interaction in the regulation of ventricular electrophysiology. Further studies are needed to explore the clinical relevance of our findings and the potential benefits of targeted therapies for patients with SAH.

Funding

Financial support was obtained from the National Natural Science Foundation of China (grant No. 81800302), Provincial Natural Science Foundation of Hunan (grant No. 2019JJ50871).

Data availability

The data sets analysed in this study are available from the corresponding author upon reasonable request.