Julian Francis Richmond Paton

Exercise intolerance in heart failure with preserved ejection fraction (HFpEF) is linked to impaired vascular regulation and exaggerated autonomic responses. Although sensory afferent signalling via P ₂ X ₃ receptors (P ₂ X ₃ Rs) contributes to exaggerated cardiovascular reflex activity in hypertension, its role in mediating haemodynamic limitation during HFpEF is unknown. We tested whether selective P ₂ X ₃ R antagonism improved blood pressure and exercise haemodynamics in a conscious ovine model of hypertensive HFpEF. Female hypertensive HFpEF sheep and age‐matched sham controls were instrumented for chronic measurement of arterial pressure, cardiac output and blood flow to the hindlimbs, kidney and coronary vasculature. Haemodynamic responses were assessed at rest and during graded treadmill exercise before and after systemic selective P ₂ X ₃ R blockade. Postexercise heart‑rate recovery was assessed as an index of physical fitness. L4 dorsal root ganglia (DRG) were analysed for gene expression related to neuronal excitability. P ₂ X ₃ R antagonism lowered resting mean arterial pressure (from 116 ± 10.2 to 110 ± 7.8 mmHg, P =  0.014), increased hindlimb blood flow at peak exercise (vehicle 4.4 ± 2.6 vs . P ₂ X ₃ R blockade 5.2 ± 2.9 L/min, P = 0.009) and improved postexercise heart rate recovery in HFpEF sheep, despite no difference in P ₂ X ₃ R mRNA expression compared with sham controls. These findings demonstrate that P ₂ X ₃ R signalling contributes to resting hypertension and impaired exercise haemodynamics in hypertensive HFpEF. The improvement in peak exercise haemodynamics in the absence of altered gene expression suggests receptor hypersensitivity or posttranslational gain of function in HFpEF. These findings identify P ₂ X ₃ R signalling as a potential mechanistic contributor to exercise limitation in HFpEF and support further investigation of this pathway as a possible therapeutic strategy. image Key points: Exercise intolerance is an important symptom in heart failure with preserved ejection fraction (HFpEF) and is linked to impaired autonomic responses. Although purinergic receptors (P ₂ X ₃ subtype) have been linked to autonomic reflex control, the role of these receptors in mediating the haemodynamic limitation during HFpEF is unknown. We used a preclinical large‐animal model of hypertensive HFpEF to show that P ₂ X ₃ R signalling contributes to resting hypertension. Blockade of these receptors improved hindlimb blood flow during peak exercise and improved heart rate recovery. Our findings identify P ₂ X ₃ R signalling as a potential mechanistic contributor to exercise limitation in HFpEF.

Arterial hypertension is a prevalent condition and a major risk factor for cardiovascular and neurological diseases. Notably, medical treatment fails to control blood pressure in a large proportion of diagnosed subjects, highlighting gaps in our understanding of hypertension's pathophysiology. Using a mathematical model of the human cardiovascular system, we quantify the contributions of various cardiovascular compartments to the hypertensive state. The impact of uncertainty in modelling assumptions is assessed through stochastic modelling. Our results show that reduced venous vascular compliance significantly increases arterial pressure, making it the second most influential factor in determining hypertension, after increased peripheral resistance. image Key points: We used a closed‐loop mathematical model of the full human circulation to simulate adaptation to hypertension, including stochastic variation in parameters. The model quantifies the relative importance of different cardiovascular parameters in determining arterial pressure. Venous compliance emerged as the second most influential determinant of arterial pressure, after systemic resistance. Reduced venous compliance alone was sufficient to cause a hypertensive state in the model. These findings highlight the potential of the venous system as a novel therapeutic target for blood pressure regulation.

Aims: The carotid body (CB) is a multimodal chemosensory organ, yet how it encodes differential external stimuli to drive distinct peripheral chemoreflex responses remains unclear. This study aimed to define differences in intrinsic cellular mechanisms underlying CB signalling and to investigate how these processes are altered in pre-hypertension. Methods and results: We used potassium cyanide (KCN) as a hypoxia mimetic to activate the CB. K+ channels implicated in hypoxia sensing were pharmacologically inhibited, with TWIK-related acid-sensitive K+ (TASK) channels targeted using Ba2+ or ML365 and BK/Kv channels targeted using TEA + 4AP or iberiotoxin. We combined in vitro Ca2+ imaging of dissociated glomus cells, ex vivo carotid sinus nerve (CSN) recordings, and an in situ double-perfused working heart-brainstem preparation to investigate chemosensory signalling in juvenile Wistar and age-matched pre-hypertensive spontaneously hypertensive rats (SHRs). KCN application to the CB ex vivo evoked a biphasic CSN response, consisting of two temporally distinct components. This biphasic response was exaggerated in SHRs, with disproportionate changes in the magnitude of each component. Ca2+ imaging revealed that these components reflect distinct glomus cell subpopulations, characterized by transient vs. prolonged KCN-evoked Ca2+ events, with their relative proportions shifted in hypertension. Pharmacological dissection suggested that differential inactivation of K+ channels contributes to these divergent Ca2+ dynamics. In situ, we confirmed that localized Ba2+ applied to the CB facilitated stronger tachypnoeic and bradycardic chemoreflex evoked responses than TEA + 4AP, consistent with functional heterogeneity among glomus cell populations. Compared with Wistar rats, in vitro TASK and BK/Kv channel expression and activity were differentially altered in SHRs, accompanied by corresponding changes in in situ chemoreflex characteristics. Conclusion: We identify, for the first time, distinct glomus cell subpopulations defined by their K+ channel expression that differentially contribute to CB signalling patterns and distinct chemoreflex pathways. These findings provide mechanistic support for the recently proposed ‘ribbon cable’ hypothesis, whereby discrete glomus cell populations couple to specific chemoreflex outputs. These findings offer new insight into CB signalling complexity and its role in autonomic dysfunction during the early stages of hypertension.

Cardiorespiratory homeostasis is maintained by carotid body (CB) peripheral chemoreceptors that monitor blood oxygen and stimulate breathing. While proportionality between peripheral chemoreflex sensitivity and respiratory response is relative to hypoxia severity, the fundamental regulatory mechanisms remain unknown. We hypothesized that intercellular amino acid transmission between glomus cells modulates CB sensitivity. We identified a full complement of glutamate and gamma‐aminobutyric acid (GABA) synaptic signalling machinery in the CB. During hypoxia stimulation, glutamate acting via NMDA receptors (NMDA‐R) stimulated CB afferents while releasing GABA, which acted on GABA _A receptors to rapidly attenuate afferent drive mediated by purinergic P2X receptors. We propose that this intrinsic ‘accelerator–brake’ mechanism regulates CB sensitivity to hypoxia. Further, glutamate and NMDA‐R were essential for long‐term facilitation (LTF) of respiratory chemoreflex responses induced by repeated hypoxia or glutamate exposures. The accelerator–brake mechanism establishes the set‐point of chemoreflex gain and provides plasticity for modulation, explaining ventilatory LTF to repeated bouts of hypoxia. image Key points: An intra‐carotid body interaction between glutamate and GABA transmission is crucial for the generation of the biphasic carotid body afferent nerve response when stimulated with low doses of sodium cyanide. Glutamate stimulates the carotid body chemoreflex to increase inspiratory drive without affecting sympathetic activity, heart rate or perfusion pressure, indicating selective activation of a carotid body respiratory reflex arc. Repetitive application of glutamate to the carotid body sensitizes chemoreflex phrenic nerve responses producing long‐term facilitation.

ATP acting on P2X3 receptors contributes to carotid body (CB) hyperexcitability in spontaneously hypertensive rats (SHRs). We investigated whether this reflects altered ATP release and/or impaired ATP metabolism. Using in vitro CB preparations, we quantified ATP release in Wistar rats and SHRs ( n = 10 each) and found significantly greater ATP release in SHRs ( P = 0.00062). Intracellular ATP depletion in dissociated glomus cells was similar between strains ( n = 4 each). Morphometric analysis revealed a disproportionate increase in glomus tissue, measured as tyrosine hydroxylase‐positive area, in juvenile SHRs compared with Wistar rats ( n = 8 each; P = 5.44 × 10 ^{− 6} ). This enlargement was associated with upregulated Epas1 mRNA ( n = 8 each), encoding HIF‑2α, a driver of CB hyperplasia. Transcripts for ATP‑degrading enzymes Enpp1, Enpp3 and Entpd2 were also downregulated in SHRs ( n = 8 each), suggesting reduced extracellular ATP breakdown. Electrophysiological recordings in vitro and reflex testing in situ showed that topical ATP or α,β‑methylene‐ATP applied to the CB evoked tachypnoea and sympathoexcitation in both strains, but SHRs displayed a greater sympathetic reflex ( n = 4 each; P = 0.037), indicating increased sensitivity to purinergic stimulation. In contrast, testing adenosine transmission revealed no difference between strains; adenosine did not contribute to CB hyperexcitability in SHRs ( n = 11 each; P = 0.53). Overall, increased ATP release in SHRs likely reflects CB enlargement from glomus cell expansion, compounded by reduced ATP‑degrading enzyme expression, thereby enhancing purinergic drive and chemoreflex sensitivity. image Key points: The carotid bodies (CBs) of spontaneously hypertensive rats (SHRs) release more ATP than those of Wistar rats. Glomus cell expansion underpins CB hyperplasia and contributes to increased ATP release in juvenile SHRs. Upregulated Epas1 (i.e. HIF‐2α) gene expression is likely responsible for CB hyperplasia in SHRs. ATP transmission is linked to exacerbated chemoreflex sympathoexcitation. Adenosine transmission does not contribute to CB hyperexcitability in SHRs.

General anesthetics enable invasive experimentation but can affect cardiovascular and respiratory physiology, biasing preclinical outcomes. We compared five anesthetic regimens in adult male Wistar rats, tribromoethanol (TBE, 250 mg/kg i.p.), chloral hydrate (CH, 400 mg/kg i.p.), ketamine–xylazine (KX, 80/10 mg/kg i.p.), thiopental (TP, 80 mg/kg i.p.), and isoflurane (ISO, 4% induction, 2% maintenance), to investigate integrated cardiorespiratory and biochemical markers. Femoral arterial catheterization allowed continuous blood pressure (BP) and derived heart rate (HR) recordings, while ventilation was assessed through pletysmography at baseline (awake), during induction, and recovery phases of anesthesia. Variability was evaluated in the time and frequency domains, including HR, systolic blood pressure (SBP), and spontaneous baroreflex sensitivity. In an independent cohort of rats, butyrylcholinesterase (BChE), CK-MB, cTnI, and LDH were measured. Baseline BP was unchanged by TBE and TP, whereas all anesthetics affected HR. Minute ventilation and breathing frequency were reduced with all agents, while tidal volume decreased with KX and TBE only. LDH and cTnI were unaffected, BChE was reduced by KX, TBE, and ISO, and CK-MB increased with CH and KX. Variability analysis showed that all anesthetics depressed pulse-interval and SBP variability and shifted spectral power toward higher frequencies, while baroreflex sensitivity and effectiveness were consistently reduced. During recovery, KX and TP restored most variability indices, whereas CH, TBE, and ISO showed persistent suppression. These findings highlight distinct profiles of cardiovascular depression and biomarker responses across anesthetics and underscore the importance of accounting for autonomic variability when selecting different anesthetics in experimental protocols. Highlights: Five anesthetic regimens were tested in rats. All anesthetics reduced ventilation, and KX and TBE also reduced tidal volume. CH and KX increased CKMB, while KX, TBE and ISO reduced BChE. All anesthetics reduced blood pressure variability and baroreflex sensitivity. Variability recovered with TP and KX, whereas CH, TBE and ISO showed persistent suppression.

BACKGROUND: Hypertension is the single most important risk factor for cardiovascular diseases and remains poorly controlled. Currently, 40% of treated patients remain hypertensive. We propose that this is in part due to excessive sympathetic activity. We hypothesized that neuronal expiratory oscillations, in the medullary lateral parafacial (pFL) region, become activated in conditions of neurogenic hypertension and that silencing their activity would be antihypertensive. METHODS: pFL neurons were manipulated by viral transfection. Using optogenetic and pharmacogenetic modulation, pFL neurons were either excited or inhibited in rats while recording sympathetic neurons from the rostral ventrolateral medulla (RVLM) and pontine noradrenergic A5 region, sympathetic activity, respiratory motor outflows, and arterial pressure from normotensive and hypertensive rats in vitro, in situ, and in vivo. Rats were made neurogenically hypertensive using chronic intermittent hypoxia. RESULTS: Optogenetic activation of pFL neurons triggered active expiration and positively modulated sympathetic activity during expiration, causing blood pressure to rise. pFL neurons projected to the RVLM and A5 presympathetic neurons and excited them during expiration, but in hypertension, only the pFL-RVLM synaptic transmission was enhanced. Pharmacogenetic inhibition of pFL neurons eliminated the expiratory-related sympatho-excitation and normalized arterial pressure in hypertensive rats. CONCLUSIONS: The heightened sympathetic activity inducing hypertension triggered by chronic intermittent hypoxia is caused, in the most part, by the emergence of pFL expiratory oscillations driving RVLM and A5 sympathetic vasomotor neurons and active expiration simultaneously. We propose that suppressing pFL neurons would have therapeutic potential.

Aims: ATP acting on P2X3 receptors (P2X3R) within carotid bodies (CBs) underpins chemoreflex-mediated sympathetic overactivity in spontaneously hypertensive rats (SHR). Pyridoxal 5′ phosphate (PLP), the active form of vitamin B6, has been reported to act as a non-selective P2X receptor blocker. Hence, we hypothesised that PLP antagonism of P2X3R in the CB would treat hypertension. Methods and results: Herein, we employed a multipronged approach to investigate PLP’s capability to attenuate CB hyperexcitability in hypertension. First, PLP inhibited Ca2+ responses evoked by α, β-methylene ATP in cell lines expressing human (h) P2X3R with an IC50 of 8.7 µM. Next, in-silico data predicted that PLP binds to the same site of Gefapixant, supporting an allosteric antagonism. Using an isolated perfused carotid artery bifurcation-CB preparation, arterial infusion of PLP (50 µM; 15 min) attenuated CBs sensory firing in SHR (P = 0.012). Using the in situ working-heart brainstem preparation, carotid artery injections of PLP (1–5 mM) attenuated the chemoreflex-evoked sympathetic (P = 0.023) but not phrenic (P = 0.62) responses; the CB was stimulated with potassium cyanide (KCN,50 µL; 0.04%). In awake telemetered SHR (n = 6), intravenous infusion of PLP (48 mg/Kg/h; 30 min) attenuated KCN-evoked chemoreflex responses and reduced systolic, diastolic, and mean blood pressures (ΔMBP = −15.6 mmHg; P = 0.025). Translating our results, we performed a small double-blind, randomised clinical trial. In volunteers with hypertension (n = 14), oral supplementation with pyridoxine hydrochloride (600 mg) attenuated the hypoxic ventilatory response only in patients with high peripheral chemoreflex sensitivity (P = 0.021). Conclusion: Our findings suggest that PLP binds to and antagonises P2X3R and is a viable candidate for larger clinical trials to treat CB dysregulation in cardiovascular diseases.