Vasoactive Intestinal Peptide is not the Dominant Mediator of Coronary Blood Flow
The chemoreflex is an autonomic reflex which maintains oxygen supply to vital organs during hypoxia.
Cardiac and Translational Physiology | Research | Amy Olley
This involves the heart via an increase in coronary blood flow, which is mediated by parasympathetic activity through unknown mechanisms. This project investigates the role of parasympathetic neurotransmitter, vasoactive intestinal peptide, in the increase of coronary blood flow during the chemoreflex.
We stimulated the chemoreflex and then compared the coronary blood flow response with and without VIP antagonism. The VIP antagonist had no effect on the coronary blood flow chemoreflex response. This indicates that VIP does not increase COBF during the chemoreflex.
Rationale
I began my summer research project with a literature review, where I was able to delve deeper into cardiac physiology than provided during my undergraduate degree. From this, I became interested in the autonomic control of the heart, particularly during the chemoreflex.
The chemoreflex is an autonomic reflex that is activated by hypoxia and is often upregulated and mechanistically distorted during heart failure [1- 2]. Peripheral chemoreceptors include the carotid body (CB) in the neck and the aortic bodies (AB) in the aortic arch [2-3]. These sense changes in blood oxygen content, sending signals to the brain via the carotid sinus and vagal nerves, respectively [4]. Sympathetic and parasympathetic neural outputs then trigger bradycardia and increased respiration, blood pressure (BP), and coronary blood flow (COBF) [3-4]. These responses maintain the blood and oxygen supply to vital organs [3-4]. The coronary arteries stem from the aorta which supplies oxygenated blood to cardiac tissue [5]. COBF is determined by the drive of blood out of the left ventricle (LV) and into the aorta and the resistance of the coronary vessels against blood flow [5]. The level of resistance is manipulated by vessel diameter, with a smaller diameter imposing a greater resistance [5]. Autonomic innervation controls this alongside local metabolic factors [3-5]. A visual representation of these relationships is outlined in figure 1.
Figure 1. A simplified illustration of the heart’s anatomy, including the coronary vasculature and the approximate major autonomic branches [6], [7]. Note that the black represents parasympathetic innervation, and green the sympathetic. This image was adapted by the author of this article.
Not only are both arms of the autonomic nervous systems activated during the chemoreflex, but stimulation of each increases COBF [1-3]. This has made it difficult to determine their distinct roles in increasing COBF during the chemoreflex. The neurotransmitters responsible for these processes are still being investigated.
The AB and CB chemoreflexes may use different neurotransmitters. Investigations into sympathetic modulation have found that propranolol had no effect on the COBF response during both AB and CB stimulation [2-3]. This suggests that neither the AB nor CB chemoreflex utilises the beta-adrenergic system to increase COBF. Vagally, atropine attenuated the COBF response with AB stimulation, but had no effect during CB stimulation [2-3]. This indicates that the CB chemoreflex does not use acetylcholine to increase COBF. In contrast, the AB chemoreflex does use acetylcholine. However, the persistence of the COBF response suggests acetylcholine is not the sole parasympathetic neurotransmitter involved [2].
My summer research project investigated the role of vasoactive-intestinal peptide (VIP) in increasing COBF during the CB and AB chemoreflexes. It is known that electrical stimulation of cardiac vagal branches causes the release of VIP into cardiac tissue, leading to an increase in COBF [8-9]. Furthermore, direct administration of VIP into the coronary arteries dilates the vessels to increase COBF [9]. This is despite a muscarinic and betaadrenergic blockade, indicating that VIP increases COBF via binding to its own VIP-specific receptors [9]. Based on this, I created the following aims:
1. Investigate the role of VIP in the COBF chemoreflex response using a VIP antagonist.
2. Document any differences between the role of VIP in the AB and CB chemoreflex.
I hypothesised that the VIP antagonist would attenuate the COBF increase during the CB and/or AB chemoreflex. I also expected to observe a difference between the CB and AB chemoreflex COBF response upon VIP antagonisation, although I was unsure what that would be.
Methods
My project stimulated the AB and CB chemoreflexes using potassium cyanide (KCN). In a conscious and anaesthetised large animal model (University of Auckland, AEC #2268), doses of 10µg/kg, 20µg/kg and 30µg/ kg of KCN were used. KCN was administered through the carotid artery in the neck to stimulate the CB chemoreflex and into the LV to stimulate the AB chemoreflex. A VIP antagonist was infused to block any VIP effects. I then analysed the data using SPIKE2 and Excel. The baseline was defined as the 15s period prior to KCN administration.
Research outcomes
In Figure 2A, the KCN30 dose produced the greatest increase in COBF of approx. +13mL/ min from baseline as expected [1, 3]. In Figure 2B, the VIP antagonist was infused, yet COBF reached a value similar to the control of approx. +16mL/min from baseline. There is no statistically significant difference between the control and VIP antagonist COBF CB chemoreflex response. This indicates that VIP does not increase COBF during the CB chemoreflex.
Due to the small number of trials performed [3], it would be interesting to repeat this project with a larger sample size. Furthermore, this project did not investigate whether the VIP antagonist was effectively binding to the VIP receptors. If these larger sample investigations are consistent with this project’s findings, then these results prompt future research into whether VIP is released during the chemoreflex, as it requires high-frequency vagal nerve activity [8-9]. Additionally, this project is unaware of any direct vagal nerve recordings during the chemoreflex, and as atropine has not attenuated this response, it may be possible that parasympathetic activity does not modulate the CB chemoreflex like it does the AB chemoreflex [1-3]. From this, research could be directed towards sympathetic transmitters.
Figure 2. The effect of intracarotid KCN on COBF where the KCN dose was administered at 0s. Image A displays the control and image B with VIP antagonism.
Figure 3. The effect of LV KCN30 to activate the ABs on COBF. This was observed with and without (control) VIP antagonism and atropine. KCN30 was administered at 0s, and HR was paced at 85bpm.
In Figure 3, the control reached a peak COBF of approximately 48mL/min above baseline after the LV KCN30 dose. Compared to Figure 2A, this was much higher than the COBF response to CB stimulation despite the HR pacing. Like other studies, this project indicates that the AB may have a more cardiac-focused response to hypoxia, whereas the CB may have a more peripheral-vasculature-focused response [1- 3]. Any neurotransmission differences between them are thus vital to understanding how this differential control occurs.
In Figure 3, the VIP antagonist appeared to increase the COBF response to LV KCN30 compared to the control, reaching approx. 58mL/min above baseline. This result does not agree with prior hypotheses and research where cardiac administration of a VIP antagonist abolished the increase in coronary blood flow resulting from vagal stimulation [2, 8-9]. As only one trial was conducted, it could be assumed that there would be no statistically significant difference between the control and the VIP antagonisation. If future trials agree with this outcome, it is likely due to similar reasons as explained for the CB chemoreflex.
In Figure 3, infusion of atropine attenuated the COBF response to LV KCN30 compared to control, reaching approx. 35mL/min above baseline. This agrees with Pen et al. [2] and Hackett et al. [10]. Thus, this project agrees that the AB chemoreflex utilises acetylcholine to increase COBF. However, because the COBF response was not abolished, other parasympathetic co-transmitters should be investigated, particularly Substance P [2, 8-9].
My summer research project improved my laboratory and data analysis skills, fortified my interest in cardiac physiology, and improved my confidence as an independent researcher. Although this project cannot conclude a role for VIP in the COBF response during the AB and CB chemoreflexes, it cannot rule out the need for investigating this role further, particularly in heart failure models. Already, it has been revealed that the chemoreflex has altered neurotransmission in hypertension and heart failure [1-2]. Thus, whether there is a difference in VIP’s role between healthy and diseased models is critical in improving the knowledge and treatment of heart failure.
I would like to thank the Heart Foundation for funding this project. I would also like to thank Dr. Julia Shanks for their mentorship, feedback, and support, as well as the Cardiac Physiology lab group’s PhD students and fellows for their patience and teaching.
Acknowledgements
Glossary
Sympathetic: One arm of the autonomic nervous system. Traditionally thought of as ‘fight or flight’.
Parasympathetic: The other arm of the autonomic nervous system. Traditionally thought of as ‘rest and digest’.
Neurotransmitter: A signalling molecule typically released from a nerve axon that binds a receptor on another neuron or tissue.
COBF: Coronary blood flow
Autonomic innervation: Where a tissue is supplied with nerves, enabling the nerve to manipulate cellular function via the release of neurotransmitters
Propranolol: A beta-adrenergic antagonist. Atropine: An acetylcholine antagonist.
Acetylcholine: The ‘main’ parasympathetic neurotransmitter.
Control: No VIP antagonist administered.
Antagonist: Binds a specific receptor type without activating them. This blocks other molecules from binding and exerting an effect.
Beta-adrenergic: A sympathetic receptor involved in the relaxation of blood vessels and airways. Bound by adrenaline or noradrenaline.
KCN: Inhibits aerobic metabolism via acting on mitochondrial cytochrome C oxidase. This inhibits the electron transport chain – responsible for producing large amounts of ATP. As a result, cells must rely on anaerobic metabolism, producing large amounts of lactate and thus hypoxia.
HR pacing: Maintains a stable HR and energy demand. This reduces the number of variables that could alter COBF.
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Amy has just completed her undergraduate Bachelor of Science degree, majoring in physiology. Over the years, Amy has developed a specific interest in cardiac physiology, particularly as a consequence of disease.