Differential release of β‐NAD⁺ and ATP upon activation of enteric motor neurons in primate and murine colons

Abstract

Background The purinergic component of enteric inhibitory neurotransmission is important for normal motility in the gastrointestinal (GI) tract. Controversies exist about the purine(s) responsible for inhibitory responses in GI muscles: adenosine 5′-triphosphate (ATP) has been assumed to be the purinergic neurotransmitter released from enteric inhibitory motor neurons, however recent studies demonstrate that β-nicotinamide adenine dinucleotide (β-NAD+) and ADP-ribose mimic the inhibitory neurotransmitter better than ATP in primate and murine colons. The study was designed to clarify the sources of purines in colons of Cynomolgus monkeys and C57BL/6 mice. Methods HPLC with fluorescence detection was used to analyze purines released by stimulation of nicotinic acetylcholine receptors (nAChR) and serotonergic 5-HT3 receptors (5-HT3R), known to be present on cell bodies and dendrites of neurons within the myenteric plexus. Key Results nAChR or 5-HT3R agonists increased overflow of ATP and β-NAD+ from tunica muscularis of monkey and murine colon. The agonists did not release purines from circular muscles of monkey colon lacking myenteric ganglia. Agonist-evoked overflow of β-NAD+, but not ATP, was inhibited by tetrodotoxin (0.5 μM) or ω-conotoxin GVIA (50 nM), suggesting that β-NAD+ release requires nerve action potentials and junctional mechanisms known to be critical for neurotransmission. ATP was likely released from nerve cell bodies in myenteric ganglia and not from nerve terminals of motor neurons. Conclusions & Inferences These results support the conclusion that ATP is not a motor neurotransmitter in the colon and are consistent with the hypothesis that β-NAD+, or its metabolites, serve as the purinergic inhibitory neurotransmitter.

Uridine adenosine tetraphosphate is a novel neurogenic P2Y1 receptor activator in the gut

“…ATP (20), NAD + (14,15,21), and adenosine 5′-diphosphate ribose (ADPR) (17) activate P2Y1R and SK channels and might be inhibitory neurotransmitters in the colon (22). Because Up4A appears to stimulate P2Y1Rs in endothelium, and P2Y1Rs are important for purinergic signaling in the colon, we investigated whether Up4A is released in colonic muscle, whether Up4A affects membrane potentials and contractions of colonic muscles, whether Up4A is an agonist for P2Y1R, whether cells expressing PDGF receptor α (PDGFRα) are targets of Up4A, and how Up4A is metabolized in colons of humans and mice.…”

The Enteric Nervous System and Gastrointestinal Innervation: Integrated Local and Central Control

“…The inhibitory neurons have multiple transmitters, including nitric oxide (NO), VIP and ATP-like transmitters [27,82]. The primary transmitter of the neurons appears to be NO, and deficits in transmission are observed if NO synthase is knocked out [83].…”

Purinergic signalling in the enteric nervous system (An overview of current perspectives)

“…For intact enteric plexuses (in situ), the process of ATP release is more complex than for isolated neuronal varicosities. Either field electrical stimulation or neuronal stimulants cause the release of both ATP and β-NAD + , yet only the release of the latter is significantly inhibited by the NaV inhibitor, TTX, or the VDCC inhibitor, ω-CTX (Mutafova-Yambolieva et al, 2007;Durnin et al, 2013). In similar experiments, field electrical stimulation enhanced the release of Up4A, but neuronal stimulants failed to do so .…”

“…This was shown first for nicotine and DMPP (at nACHRs) (Gillespie and MacKenna, 1960), thereafter for GABA and muscimol (at GABAA) (for example, see: Boeckxstaens et al, 1991;Bayer et al, 2002) and subsequently, for 5-HT and 2-methyl-5-HT (at 5-HT3Rs) (Zhou and Galligan, 1999;Neal and Bornstein, 2007;Dickson et al, 2010). In each of these cases, the actions of neuronal stimulants are blocked either by tetrodotoxin (TTX; an inhibitor of voltage-gated sodium ion channels (NaV)) or by selective antagonists of the above receptor subtypes, to inhibit the output of purines (Durnin et al, 2013). ATP and α,β-meATP-gated ion channels can join this list of neuronal stimulants, but they may act in a different way and at different locations (Boeckxstaens et al, 1991;De Man et al, 2003;Van Crombruggen et al, 2007;King and Townsend-Nicholson, 2008).…”

“…This related substance may be beta-nicotinamide adenine dinucleotide (β-NAD + ), or its CD38-generated bioactive metabolite, adenosine 5′-diphosphate ribose (ADPr) (Mutafova-Yambolieva et al, 2007;Durnin et al, 2012Durnin et al, , 2013. Both β-NAD + and ADPr are agonists of recombinant P2Y1 isoforms, although weaker agonists compared to the potency of ADP/ATP (Gustafsson et al, 2011;Mutafova-Yambolieva et al, 2007).…”