Signal processing at mammalian carotid body chemoreceptors.

Mammalian carotid bodies are richly vascularized chemosensory organs that sense blood levels of O(2), CO(2)/H(+), and glucose and maintain homeostatic regulation of these levels via the reflex control of ventilation. Carotid bodies consist of innervated clusters of type I (or glomus) cells in intimate association with glial-like type II cells. Carotid bodies make afferent connections with fibers from sensory neurons in the petrosal ganglia and receive efferent inhibitory innervation from parasympathetic neurons located in the carotid sinus and glossopharyngeal nerves. There are synapses between type I (chemosensory) cells and petrosal afferent terminals, as well as between neighboring type I cells. There is a broad array of neurotransmitters and neuromodulators and their ionotropic and metabotropic receptors in the carotid body. This allows for complex processing of sensory stimuli (e.g., hypoxia and acid hypercapnia) involving both autocrine and paracrine signaling pathways. This review summarizes and evaluates current knowledge of these pathways and presents an integrated working model on information processing in carotid bodies. Included in this model is a novel hypothesis for a potential role of type II cells as an amplifier for the release of a key excitatory carotid body neurotransmitter, ATP, via P2Y purinoceptors and pannexin-1 channels.

Expanding role of ATP as a versatile messenger at carotid and aortic body chemoreceptors.

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) ( ), 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.

Developmental regulation of glucosensing in rat adrenomedullary chromaffin cells: potential role of the K(ATP) channel.

During birth, when the maternal supply of glucose is occluded, there is a drastic fall in blood glucose in the newborn. This stimulus triggers the non-neurogenic release of catecholamines from adrenomedullary chromaffin cells, which restores blood glucose homeostasis. In this report we present preliminary data showing that glucosensing is present in neonatal chromaffin cells from adrenal slices but absent in chromaffin cells from juvenile slices. Moreover, we show that the aglycemia-evoked rise in intracellular Ca2+ is robust in neonatal chromaffin cells but blunted in juvenile chromaffin cells. Lastly, we show that the Kir6.2 subunit of the KATP channel, is upregulated in the adrenal medulla in juvenile animals providing a potential mechanism for the developmental regulation of glucosensing.

P2Y2 receptor activation opens pannexin-1 channels in rat carotid body type II cells: potential role in amplifying the neurotransmitter ATP.

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.

Effects of chemostimuli on [Ca2+]i responses of rat aortic body type I cells and endogenous local neurons: comparison with carotid body cells.

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 Ca2+ 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 [Ca2+] in AB type I cells. The [Ca2+]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 Ca2+ channel antagonist, 2mM Ni2+. 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 [Ca2+]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.

Confocal immunofluorescence study of rat aortic body chemoreceptors and associated neurons in situ and in vitro.

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.

Glucosensing in an immortalized adrenomedullary chromaffin cell line: role of ATP-sensitive K+ channels.

Using an immortalized adrenal chromaffin cell line (MAH cells), we investigated the cellular mechanisms underlying sensitivity to glucose-free solution (aglycemia) using ratiometric Ca2+ imaging and whole-cell recording. Though few cells (< 15%) responded to aglycemia with an increase in intracellular-free Ca2+ concentration ([Ca2+]i), in most cells (approximately 75%), aglycemia caused > 50% suppression of the Delta[Ca2+]i induced by the depolarizing stimulus, high (10 mM) K+. Moreover, in normal K+, the average aglycemia-induced rise in Cai2+ as well as the proportion of aglycemia-responsive cells increased in the presence of the K(ATP) channel blocker, glibenclamide. During membrane potential (Vm) measurements, aglycemia induced either hyperpolarization (6/20), depolarization (4/20) or no change in Vm. RT-PCR and Western blotting confirmed the presence of K(ATP) channel subunits Kir6.2 and SUR1 in MAH cells. These findings suggest a dual inhibitory and excitatory action of aglycemia in MAH cells, where activation of K(ATP) channels effectively inhibits or blunts the Delta[Ca2+]i due to the excitatory effect. Thus, this cell line appears as an attractive model for studying the molecular mechanisms of glucosensing.