Geoff Burnstock, purinergic signalling, and chemosensory control of breathing.

This article is the authors' contribution to the tribute issue in honour of Geoffrey Burnstock, the founder of this journal and the field of purinergic signalling. We give a brief account of the results of experimental studies which at the beginning received valuable input from Geoff, who both directly and indirectly influenced our research undertaken over the last two decades. Research into the mechanisms controlling breathing identified ATP as the common mediator of the central and peripheral chemosensory transduction. Studies of the sources and mechanisms of chemosensory ATP release in the CNS suggested that this signalling pathway is universally engaged in conditions of increased metabolic demand by brain glial cells - astrocytes. Astrocytes appear to function as versatile CNS metabolic sensors that detect changes in brain tissue pH, CO2, oxygen, and cerebral perfusion pressure. Experimental studies on various aspects of astrocyte biology generated data indicating that the function of these omnipresent glial cells and communication between astrocytes and neurons are governed by purinergic signalling, - first discovered by Geoff Burnstock in the 70's and researched through his entire scientific career

Cardiac vagal preganglionic neurones: An update

The autonomic nervous system controls the heart by dynamic recruitment and withdrawal of cardiac parasympathetic and sympathetic activities. These activities are generated by groups of sympathoexcitatory and vagal preganglionic neurones residing in a close proximity to each other within well-defined structures of the brainstem. This short essay provides a general overview and an update on the latest developments in our understanding of the central nervous origins and functional significance of cardiac vagal tone. Significant experimental evidence suggests that distinct groups of cardiac vagal preganglionic neurones with different patterns of activity control nodal tissue (controlling the heart rate and atrioventricular conductance) and the ventricular myocardium (modulating its contractility and excitability)

Cardioprotection evoked by remote ischaemic preconditioning is critically dependent on the activity of vagal pre-ganglionic neurones

AIMS: Innate mechanisms of inter-organ protection underlie the phenomenon of remote ischaemic preconditioning (RPc) in which episode(s) of ischaemia and reperfusion in tissues remote from the heart reduce myocardial ischaemia/reperfusion injury. The uncertainty surrounding the mechanism(s) underlying RPc centres on whether humoral factor(s) produced during ischaemia/reperfusion of remote tissue and released into the systemic circulation mediate RPc, or whether a neural signal is required. While these two hypotheses may not be incompatible, one approach to clarify the potential role of a neural pathway requires targeted disruption or activation of discrete central nervous substrate(s). METHODS AND RESULTS: Using a rat model of myocardial ischaemia/reperfusion injury in combination with viral gene transfer, pharmaco-, and optogenetics, we tested the hypothesis that RPc cardioprotection depends on the activity of vagal pre-ganglionic neurones and consequently an intact parasympathetic drive. For cell-specific silencing or activation, neurones of the brainstem dorsal motor nucleus of the vagus nerve (DVMN) were targeted using viral vectors to express a Drosophila allatostatin receptor (AlstR) or light-sensitive fast channelrhodopsin variant (ChIEF), respectively. RPc cardioprotection, elicited by ischaemia/reperfusion of the limbs, was abolished when DVMN neurones transduced to express AlstR were silenced by selective ligand allatostatin or in conditions of systemic muscarinic receptor blockade with atropine. In the absence of remote ischaemia/reperfusion, optogenetic activation of DVMN neurones transduced to express ChIEF reduced infarct size, mimicking the effect of RPc. CONCLUSION: These data indicate a crucial dependence of RPc cardioprotection against ischaemia/reperfusion injury upon the activity of a distinct population of vagal pre-ganglionic neurones

Brain Stem Death as the Vital Determinant for Resumption of Spontaneous Circulation after Cardiac Arrest in Rats

BACKGROUND:Spontaneous circulation returns to less than half of adult cardiac arrest victims who received in-hospital resuscitation. One clue for this disheartening outcome arises from the prognosis that asystole invariably takes place, after a time lag, on diagnosis of brain stem death. The designation of brain stem death as the point of no return further suggests that permanent impairment of the brain stem cardiovascular regulatory machinery precedes death. It follows that a crucial determinant for successful revival of an arrested heart is that spontaneous circulation must resume before brain stem death commences. Here, we evaluated the hypothesis that maintained functional integrity of the rostral ventrolateral medulla (RVLM), a neural substrate that is intimately related to brain stem death and central circulatory regulation, holds the key to the vital time-window between cardiac arrest and resumption of spontaneous circulation. METHODOLOGY/PRINCIPAL FINDINGS:An animal model of brain stem death employing the pesticide mevinphos as the experimental insult in Sprague-Dawley rats was used. Intravenous administration of lethal doses of mevinphos elicited an abrupt cardiac arrest, accompanied by elevated systemic arterial pressure and anoxia, augmented neuronal excitability and enhanced microvascular perfusion in RVLM. This period represents the vital time-window between cardiac arrest and resumption of spontaneous circulation in our experimental model. Animals with restored spontaneous circulation exhibited maintained neuronal functionality in RVLM beyond this critical time-window, alongside resumption of baseline tissue oxygen and enhancement of local blood flow. Intriguingly, animals that subsequently died manifested sustained anoxia, diminished local blood flow, depressed mitochondrial electron transport activities and reduced ATP production, leading to necrotic cell death in RVLM. That amelioration of mitochondrial dysfunction and bioenergetic failure in RVLM by coenzyme Q10, the mobile electron carrier in mitochondrial respiratory chain, or oxygenation restored spontaneous circulation further established a causal relationship between functionality of RVLM and resumed spontaneous circulation after cardiac arrest. CONCLUSIONS/SIGNIFICANCE:We conclude that whereas necrotic cell death because of bioenergetic failure triggered by anoxia in RVLM, which precipitates brain stem death, negates resuscitation of an arrested heart, maintained functional integrity of this neural substrate holds the key to resumption of spontaneous circulation after cardiac arrest in rats

A2 Noradrenergic Lesions Prevent Renal Sympathoinhibition Induced by Hypernatremia in Rats

Renal vasodilation and sympathoinhibition are recognized responses induced by hypernatremia, but the central neural pathways underlying such responses are not yet entirely understood. Several findings suggest that A2 noradrenergic neurons, which are found in the nucleus of the solitary tract (NTS), play a role in the pathways that contribute to body fluid homeostasis and cardiovascular regulation. The purpose of this study was to determine the effects of selective lesions of A2 neurons on the renal vasodilation and sympathoinhibition induced by hypertonic saline (HS) infusion. Male Wistar rats (280–350 g) received an injection into the NTS of anti-dopamine-beta-hydroxylase-saporin (A2 lesion; 6.3 ng in 60 nl; n = 6) or free saporin (sham; 1.3 ng in 60 nl; n = 7). Two weeks later, the rats were anesthetized (urethane 1.2 g⋅kg−1 b.wt., i.v.) and the blood pressure, renal blood flow (RBF), renal vascular conductance (RVC) and renal sympathetic nerve activity (RSNA) were recorded. In sham rats, the HS infusion (3 M NaCl, 1.8 ml⋅kg−1 b.wt., i.v.) induced transient hypertension (peak at 10 min after HS; 9±2.7 mmHg) and increases in the RBF and RVC (141±7.9% and 140±7.9% of baseline at 60 min after HS, respectively). HS infusion also decreased the RSNA (−45±5.0% at 10 min after HS) throughout the experimental period. In the A2-lesioned rats, the HS infusion induced transient hypertension (6±1.4 mmHg at 10 min after HS), as well as increased RBF and RVC (133±5.2% and 134±6.9% of baseline at 60 min after HS, respectively). However, in these rats, the HS failed to reduce the RSNA (115±3.1% at 10 min after HS). The extent of the catecholaminergic lesions was confirmed by immunocytochemistry. These results suggest that A2 noradrenergic neurons are components of the neural pathways regulating the composition of the extracellular fluid compartment and are selectively involved in hypernatremia-induced sympathoinhibition

Sumoylation of Hypoxia-Inducible Factor-1α Ameliorates Failure of Brain Stem Cardiovascular Regulation in Experimental Brain Death

One aspect of brain death is cardiovascular deregulation because asystole invariably occurs shortly after its diagnosis. A suitable neural substrate for mechanistic delineation of this aspect of brain death resides in the rostral ventrolateral medulla (RVLM). RVLM is the origin of a life-and-death signal that our laboratory detected from blood pressure of comatose patients that disappears before brain death ensues. At the same time, transcriptional upregulation of heme oxygenase-1 in RVLM by hypoxia-inducible factor-1α (HIF-1α) plays a pro-life role in experimental brain death, and HIF-1α is subject to sumoylation activated by transient cerebral ischemia. It follows that sumoylation of HIF-1α in RVLM in response to hypoxia may play a modulatory role on brain stem cardiovascular regulation during experimental brain death.A clinically relevant animal model that employed mevinphos as the experimental insult in Sprague-Dawley rat was used. Biochemical changes in RVLM during distinct phenotypes in systemic arterial pressure spectrum that reflect maintained or defunct brain stem cardiovascular regulation were studied. Western blot analysis, EMSA, ELISA, confocal microscopy and immunoprecipitation demonstrated that drastic tissue hypoxia, elevated levels of proteins conjugated by small ubiquitin-related modifier-1 (SUMO-1), Ubc9 (the only known conjugating enzyme for the sumoylation pathway) or HIF-1α, augmented sumoylation of HIF-1α, nucleus-bound translocation and enhanced transcriptional activity of HIF-1α in RVLM neurons took place preferentially during the pro-life phase of experimental brain death. Furthermore, loss-of-function manipulations by immunoneutralization of SUMO-1, Ubc9 or HIF-1α in RVLM blunted the upregulated nitric oxide synthase I/protein kinase G signaling cascade, which sustains the brain stem cardiovascular regulatory machinery during the pro-life phase.We conclude that sumoylation of HIF-1α in RVLM ameliorates brain stem cardiovascular regulatory failure during experimental brain death via upregulation of nitric oxide synthase I/protein kinase G signaling. This information should offer new therapeutic initiatives against this fatal eventuality

A Single Gene Target of an ETS-Family Transcription Factor Determines Neuronal CO2-Chemosensitivity

Many animals possess neurons specialized for the detection of carbon dioxide (CO2), which acts as a cue to elicit behavioral responses and is also an internally generated product of respiration that regulates animal physiology. In many organisms how such neurons detect CO2 is poorly understood. We report here a mechanism that endows C. elegans neurons with the ability to detect CO2. The ETS-5 transcription factor is necessary for the specification of CO2-sensing BAG neurons. Expression of a single ETS-5 target gene, gcy-9, which encodes a receptor-type guanylate cyclase, is sufficient to bypass a requirement for ets-5 in CO2-detection and transforms neurons into CO2-sensing neurons. Because ETS-5 and GCY-9 are members of gene families that are conserved between nematodes and vertebrates, a similar mechanism might act in the specification of CO2-sensing neurons in other phyla

Pregnancy and Infant Outcomes among HIV-Infected Women Taking Long-Term ART with and without Tenofovir in the DART Trial

Diana Gibb and colleagues investigate the effect of in utero tenofovir exposure by analyzing the pregnancy and infant outcomes of HIV-infected women enrolled in the DART trial