Purinergic modulation of preBötzinger complex inspiratory rhythm in rodents: the interaction between ATP and adenosine

Abstract

Non-technical summary Hypoxia causes an increase in breathing followed by a secondary depression that is most pronounced, and potentially life-threatening, in premature infants. Adenosine triphosphate (ATP) is released in brainstem respiratory networks during hypoxia, where it attenuates the secondary respiratory depression. Mechanisms are unknown but likely to be complex because ATP is degraded by enzymes into ADP, which is excitatory, and adenosine (ADO), which is inhibitory. We demonstrate in mouse, like rat, that ATP in the preBötzinger complex (preBötC), a site critical for inspiratory rhythm generation, increases frequency. Unlike rat, this increase is only observed in mouse if ADO receptors are blocked. Differential ATP sensitivity is likely to reflect that ADO is only inhibitory in mouse, and that mouse preBötC enzymes favour ADO production. Thus, purinergic signalling in preBötC networks appears balanced to favour inhibition in mouse but excitation in rat. Knowledge of purinergic signalling increases our understanding of processes underlying respiratory responses to hypoxia.Abstract ATP signalling in the CNS is mediated by a three-part system comprising the actions of ATP (and ADP) at P2 receptors (P2Rs), adenosine (ADO) at P1 receptors (P1Rs), and ectonucleotidases that degrade ATP into ADO. ATP excites preBötzinger complex (preBötC) inspiratory rhythm-generating networks where its release attenuates the hypoxic depression of breathing. Its metabolite, ADO, inhibits breathing through unknown mechanisms that may involve the preBötC. Our objective is to understand the dynamics of this signalling system and its influence on preBötC networks. We show that the preBötC of mouse and rat is sensitive to P2Y 1 purinoceptor (P2Y 1 R) activation, responding with a >2-fold increase in frequency. Remarkably, the mouse preBötC is insensitive to ATP. Only after block of A 1 ADORs is the ATP-evoked, P2Y 1 R-mediated frequency increase observed. This demonstrates that ATP is rapidly degraded to ADO, which activates inhibitory A 1 Rs, counteracting the P2Y 1 R-mediated excitation. ADO sensitivity of mouse preBötC was confirmed by a frequency decrease that was absent in rat. Differential ectonucleotidase activities are likely to contribute to the negligible ATP sensitivity of mouse preBötC. Real-time PCR analysis of ectonucleotidase isoforms in preBötC punches revealed TNAP (degrades ATP to ADO) or ENTPDase2 (favours production of excitatory ADP) as the primary constituent in mouse and rat, respectively. These data further establish the sensitivity of this vital network to P2Y 1 R-mediated excitation, emphasizing that individual components of the three-part signalling system dramatically alter network responses to ATP. Data also suggest therapeutic potential may derive from methods that alter the ATP-ADO balance to favour the excitatory actions of ATP.

A1 adenosine receptor modulation of chemically and electrically evoked lumbar locomotor network activity in isolated newborn rat spinal cords

“…Ruangkittisakul and Ballanyi, 2010;Zwicker et al, 2011). The latter findings indicate a species difference for ADO effects for both rhythmogenic cranial inspiratory and spinal locomotor CPGs because ADO (modestly) inhibits both systems in newborn mice in vitro (Zwicker et al, 2011;Witts et al, 2012). In the following, we discuss our findings regarding the literature for interstitial ADO levels and putative mechanisms involved in the A 1 receptor-mediated ADO effects on newborn rat lumbar CPGs revealed here.…”

“…For quantification of these Results, see Table 4. Ruangkittisakul and Ballanyi, 2010;Zwicker et al, 2011). The latter findings indicate a species difference for ADO effects for both rhythmogenic cranial inspiratory and spinal locomotor CPGs because ADO (modestly) inhibits both systems in newborn mice in vitro (Zwicker et al, 2011;Witts et al, 2012).…”

“…Similarly, slowing of fictive locomotion in isolated newborn mouse cords seems to be due to CPG inhibition resulting from A 1 receptor activation by glial-derived ADO (Witts et al, 2012). Correspondingly, focal injection of ATP into newborn mouse brainstem slices does not accelerate pre-Bo¨tzinger complex (preBo¨tC) inspiratory center bursting that is normally mediated by P2 receptors because of the rapid degradation of this typically excitatory neuromodulator to ADO which then activates A 1 receptors (Zwicker et al, 2011). Consequently, (modest) slowing of inspiratory-related rhythm was seen in the latter study upon ADO injection into the preBo¨tC of 0306-4522/12 $36.00 Ó 2012 IBRO.…”

Purinergic Signaling in the Airways

“…Recent data indicated that ionotropic glutamate and P2X receptors in the rostral ventrolateral medulla of the Bötzinger complex play roles in opposite directions in the control of respiratory responses to chemoreflex activation in awake rats (Moreaes et al, 2011). It has been shown in rodents that the ATP-adenosine balance is a determining factor in the purinergic modulation of the PBC inspiratory rhythm and that the underlying pathways seem to differ between mice and rats (Zwicker et al, 2011). Another article showed that the chemosensitive regions of the ventrolateral medulla were immunoreactive for the P2X2 receptor, which mediates ATP potentiation of respiratory discharge frequency (Lorier et al, 2004).…”

C1 neurons: the body's EMTs

“…This response is independent of the carotid bodies and is largely mediated by the activation of the barosensitive neurons of the RVLM including the C1 cells (47, 48, 67, 180). Their depolarization, like that of the pre-Bötzinger complex, could be partly a cell-autonomous response, for example, an increase in persistent sodium current or calcium permeability (69,136) or could result from the release of gliotransmitters such as ATP (65,115,169,181,202). Reis and collaborators championed the idea that the C1 cells are oxygen sensors, implying that these neurons are activated by small "physiologically relevant" reductions in brain PO 2 and then trigger countervailing responses including an increase in AP and a neurogenic rise in cerebral blood flow without concomitant change in brain metabolism (180).…”