Key points• Carotid body (CB) chemoreceptor complexes consist of receptor type I cells, intimately associated with glia-like type II cells whose function is poorly understood.• We show that type II cells in the rat CB express gap junction-like proteins, pannexin-1 (Panx-1) channels, which form non-selective pores permeable to ions and large molecules such as ATP, a key CB neurotransmitter.• Activation of purinergic P2Y2 receptors on type II cells led to a rise in intracellular Ca 2+ , and a prolonged membrane depolarization due to opening of Panx-1 channels.• In a CB co-culture model, where purinergic P2X2/3-expressing petrosal neurones served as a reporter or biosensor of ATP release, we show that selective activation of P2Y2 receptors on type II cells can lead to ATP release via Panx-1 channels.• We propose that type II cells may function as amplifiers of the neurotransmitter ATP during chemotransduction, via the mechanism of ATP-induced ATP release.Abstract Signal processing in the carotid body (CB) is initiated at receptor glomus (or type I) cells which depolarize and release the excitatory neurotransmitter ATP during chemoexcitation by hypoxia and acid hypercapnia. Glomus cell clusters (GCs) occur in intimate association with glia-like type II cells which express purinergic P2Y2 receptors (P2Y2Rs) but their function is unclear. Here we immunolocalize the gap junction-like protein channel pannexin-1 (Panx-1) in type II cells and show Panx-1 mRNA expression in the rat CB. As expected, type II cell activation within or near isolated GCs by P2Y2R agonists, ATP and UTP (100 μM), induced a rise in intracellular [Ca 2+ ]. Moreover in perforated-patch whole cell recordings from type II cells, these agonists caused a prolonged depolarization and a concentration-dependent, delayed opening of non-selective ion channels that was prevented by Panx-1 blockers, carbenoxolone (5 μM) and 4,4 -diisothiocyano-2,2 -stilbenedisulfonic acid (DIDS; 10 μM). Because Panx-1 channels serve as conduits for ATP release, we hypothesized that paracrine, type II cell P2Y2R activation leads to ATP-induced ATP release. In proof-of-principle experiments we used co-cultured chemoafferent petrosal neurones (PNs), which express P2X2/3 purinoceptors, as sensitive biosensors of ATP released from type II cells. In several cases, UTP activation of type II cells within or near GCs led to depolarization or increased firing in nearby PNs, and the effect was reversibly abolished by the selective P2X2/3 receptor blocker, pyridoxalphosphate-6-azophenyl-2 ,4 -disulphonic acid (PPADS; 10 μM). We propose that CB type II cells may function as ATP amplifiers during chemotransduction via paracrine activation of P2Y2Rs and Panx-1 channels. Abbreviations BGM, basic growth medium; CB, carotid body; GC, glomus cell cluster; P2Y2R, P2Y2 receptor; PN, petrosal neurone.

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Key points• Mammalian aortic bodies (ABs) are putative peripheral chemoreceptor cells presumed to monitor the oxygen content of arterial blood, although their direct chemosensitivity has never been previously demonstrated at the cellular level.• We used Ca 2+ imaging to show for the first time that a variety of stimuli, including hypoxia, isohydric and acidic hypercapnia, and isocapnic acidosis, caused increases in cytosolic [Ca 2+ ] in AB chemoreceptor cells.• We also showed that some local neurons, known to be uniquely associated with these AB paraganglia in situ, generated robust [Ca 2+ ] i responses to these chemostimuli, suggesting that they may subserve a sensory function.• These results will help us better understand how AB cells sense the composition of the blood in their local environment near the heart, and how they communicate with sensory neurons to initiate homeostatic reflexes during situations of low oxygen supply, like anaemia.Abstract Mammalian aortic bodies (ABs) are putative peripheral arterial chemoreceptors whose function remains controversial, partly because information on their cellular physiology is lacking. In this study, we used ratiometric Ca 2+ imaging to investigate for the first time chemosensitivity in short-term cultures of dissociated cells of juvenile rat ABs, located near the junction of the left vagus and recurrent laryngeal nerves. Among the surviving cell population were glomus or type I cell clusters, endogenous local neurons and glia-like cells. A variety of chemostimuli, including hypoxia, isohydric or acidic hypercapnia, and isocapnic acidosis, caused a rise in intracellular [Ca 2+ ] in AB type I cells. The [Ca 2+ ] i responses were indistinguishable from those in carotid body (CB) type I cells grown in parallel cultures from the same animals, and responses to acidic hypercapnia were prevented by the non-specific voltage-gated Ca 2+ channel antagonist, 2 mM Ni 2+ . Furthermore, we identified a subpopulation (∼40%) of glia-like cells in AB cultures that resembled CB type II cells based on their approximately equal sensitivity to ATP and UTP, consistent with the expression of purinergic P2Y2 receptors. Finally, we showed that some local neurons, known to be uniquely associated with these AB paraganglia in situ, generated robust [Ca 2+ ] i responses to these chemostimuli. Thus, these AB type I cells and associated putative type II cells resemble those from the well-studied CB. Unlike the CB, however, they also associate with a special group of endogenous neurons which we propose may subserve a sensory function in local cardiovascular reflexes. Abbreviations AB, aortic body; BBS, bicarbonate-buffered solution; CB, carotid body; GFAP, glial-fibrillary acidic protein; nNOS, neuronal nitric oxide synthase; RLN, recurrent laryngeal nerve; TH, tyrosine hydroxylase; VAChT, vesicular acetylcholine transporter.

Aortic bodies (ABs) are putative peripheral arterial chemoreceptors, distributed near the aortic arch. Though presumed to be analogous to the well-studied carotid bodies (CBs), their anatomical organization, innervation, and function are poorly understood. By using multilabel confocal immunofluorescence, we investigated the cellular organization, innervation, and neurochemistry of ABs in whole mounts of juvenile rat vagus and recurrent laryngeal (V-RL) nerves and in dissociated cell culture. Clusters of tyrosine hydroxylase-immunoreactive (TH-IR) glomus cells were routinely identified within these nerves. Unlike the CB, many neuronal cell bodies and processes, identified by peripherin (PR) and neurofilament/growth-associated protein (NF70/GAP-43) immunoreactivity, were closely associated with AB glomus clusters, especially near the V-RL bifurcation. Some neuronal cell bodies were immunopositive for P2X2 and P2X3 purinoceptor subunits, which were also found in nerve terminals surrounding glomus cells. Immunoreactivity against the vesicular acetylcholine transporter (VAChT) was detected in local neurons, glomus cells, and apposed nerve terminals. Few neurons were immunopositive for TH or neuronal nitric oxide synthase. A similar pattern of purinoceptor immunoreactivity was observed in tissue sections of adult rat V-RL nerves, except that glomus cells were weakly P2X3-IR. Dissociated monolayer cultures of juvenile rat V-RL nerves yielded TH-IR glomus clusters in intimate association with PR- or NF70/GAP-43-IR neurons and their processes, and glial fibrillary acidic protein-IR type II (sustentacular) cells. Cocultures survived for several days, wherein neurons expressed voltage-activated ionic currents and generated action potentials. Thus, this coculture model is attractive for investigating the role of glomus cells and local neurons in AB function.

In mammals, peripheral arterial chemoreceptors monitor blood chemicals (e.g. O 2 , CO 2 , H + , glucose) and maintain homeostasis via initiation of respiratory and cardiovascular reflexes. Whereas chemoreceptors in the carotid bodies (CBs), located bilaterally at the carotid bifurcation, control primarily respiratory functions, those in the more diffusely distributed aortic bodies (ABs) are thought to regulate mainly cardiovascular functions. Functionally, CBs sense partial pressure of O 2 (P O 2 ), whereas ABs are considered sensors of O 2 content. How these organs, with essentially a similar complement of chemoreceptor cells, differentially process these two different types of signals remains enigmatic. Here, we review evidence that implicates ATP as a central mediator during information processing in the CB. Recent data allow an integrative view concerning its interactions at purinergic P2X and P2Y receptors within the chemosensory complex that contains elements of a 'quadripartite synapse' . We also discuss recent studies on the cellular physiology of ABs located near the aortic arch, as well as immunohistochemical evidence suggesting the presence of pathways for P2X receptor signalling. Finally, we present a hypothetical 'quadripartite model' to explain how ATP, released from red blood cells during hypoxia, could contribute to the ability of ABs to sense O 2 content.