Reactive oxygen species contribute to the presynaptic action of extracellular ATP at the frog neuromuscular junction

Abstract

During normal cell metabolism the production of intracellular ATP is associated with the generation of reactive oxygen species (ROS), which appear to be important signalling molecules. Both ATP and ROS can be released extracellularly by skeletal muscle during intense activity. Using voltage clamp recording combined with imaging and biochemical assay of ROS, we tested the hypothesis that at the neuromuscular junction extracellular ATP generates ROS to inhibit transmitter release from motor nerve endings. We found that ATP produced the presynaptic inhibitory action on multiquantal end-plate currents. The inhibitory action of ATP (but not that of adenosine) was significantly reduced by several antioxidants or extracellular catalase, which breaks down H 2 O 2 . Consistent with these data, the depressant effect of ATP was dramatically potentiated by the pro-oxidant Fe 2+ . Exogenous H 2 O 2 reproduced the depressant effects of ATP and showed similar sensitivity to anti-and pro-oxidants. While NO also inhibited synaptic transmission, inhibitors of the NO-producing cascade did not prevent the depressant action of ATP. The ferrous oxidation in xylenol orange assay showed the increase of ROS production by ATP and 2-MeSADP but not by adenosine. Suramin, a non-selective antagonist of P2 receptors, and pertussis toxin prevented the action of ATP on ROS production. Likewise, imaging with the ROS-sensitive dye carboxy-2 ,7 -dichlorodihydrofluorescein revealed increased production of ROS in the muscle treated with ATP or ADP while UTP or adenosine had no effect. Thus, generation of ROS contributed to the ATP-mediated negative feedback mechanism controlling quantal secretion of ACh from the motor nerve endings.

Inactivation of GABAA receptor is related to heat shock stress response in organism model Caenorhabditis elegans

“…Sigmoid movement depends on acetylcholine (ACh) neurotransmission as well but in our experiments appears unaffected by heat shock treatment. These results concurred with reports that ROS actions do not deactivate nicotinic acetylcholine receptors (nAChR) at the neuromuscular junction (Ben-Haim et al 1973;Giniatullin et al 2005) On the contrary, ROS and heat shock inhibit or deactivate enzyme acetylcholinesterase (Eichler and Silman 1995), keeping the ACh action longer in the synaptic cleft.…”

Reactive Oxygen Species Mediate the Potentiating Effects of ATP on GABAergic Synaptic Transmission in the Immature Hippocampus

“…To test whether the potentiating effect of ATP on GABAergic SPSCs was mediated by ROS, we used the antioxidant NAC, which is a widely used ROS scavenger (5,27). The action of ATP on SPSCs was prevented by NAC ( Fig.…”

“…ROS Imaging-The above mentioned experiments clearly indicated that ATP is able to induce calcium transient in astrocytes via P2Y 1 receptors. To directly test whether ATP triggers ROS generation in astrocytes, we performed imaging of ROS in brain slices using the fluorescent dye DCF (5,10,21).…”

“…Although at the periphery, ROS contribute to the inhibitory effect of ATP on quantal acetylcholine release from motor nerve endings (5), in the central nervous system, ROS produce both enhancement and depression of synaptic transmission (6 -9). In pathological conditions, increased amounts of ROS are involved in apoptosis and neurodegenerative disorders such as Parkinson and Alzheimer disease (3,4,10).…”

Purinergic signalling in the musculoskeletal system

“…To date, much of the evidence for purinergic involvement in skeletal neuromuscular transmission has come from studies of the fish electric organ, frog and chick neuromuscular junctions (see [13] and [14][15][16][17][18][19][20][21]). …”