The effects on acetylcholine-induced membrane currents (ACh currents), produced by agents known to modify the activity of intracellular messengers, were studied in the neurons of the guinea-pig ileum submucous plexus (SMP) using a... more
The effects on acetylcholine-induced membrane currents (ACh currents), produced by agents known to modify the activity of intracellular messengers, were studied in the neurons of the guinea-pig ileum submucous plexus (SMP) using a whole-cell patch clamp recording method. The ACh currents were not affected by forskolin, the adenylate cyclase activator, regardless of whether or not ATP and GTP were present in the intracellular solution, and by phorbol 12-myristate 13-acetate, the protein kinase C activator. The ACh currents were strongly suppressed by thapsigargin, the microsomal calcium ATPase inhibitor, and genistein, the tyrosine protein kinase inhibitor. They were also suppressed by 3-isobutyl-1-methylxanthine, the cyclic-AMP phosphodiesterase inhibitor, regardless of the presence of forskolin in the extracellular solution and ATP and GTP in the intracellular solution. In addition, the currents were suppressed by activation of P2 purinoceptors with ATP, which could not be explaine...
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ABSTRACT Effects of exogenous adenosine 5′-triphosphate (ATP) on dissociated guinea pig ileum submucous neurons were studied using a conventional whole-cell patch-clamp technique. With the holding potential of −50 mV, application of... more
ABSTRACT Effects of exogenous adenosine 5′-triphosphate (ATP) on dissociated guinea pig ileum submucous neurons were studied using a conventional whole-cell patch-clamp technique. With the holding potential of −50 mV, application of 50–1,000 μM ATP evoked an inward current (ATP-induced current) in most (90%) of the tested neurons (n-35). ATP-induced currents were observed regardless of whether or not guanosine 5′-triphosphate (GTP, 0.2 mM) and ATP (2 mM) were present in the intracellular solution, or GTP was replaced with equimolar concentration of guanosine 5′-O-3-thiotriphosphate (n-5). In 26 of 29 neurons studied, which responded to ATP, applications of 50–1,000 μM ATP induced slowly declining currents. ATP receptors did not appear to be completely desensitized during a long pulse (up to 4 min) of 200 μM ATP. Suramin (200 μM) accelerated an increase to peak of the current induced by 200 μM ATP without affecting the maximum response amplitude (n−4_. In about 10% of the neuronsn−3), 50 μM ATP evoked rapidly declining (about 1 sec) currents. Application of 100 μM α,β-Me-ATP to these neurons evoked similar responses. The above results suggest that submucous neurons express two specific subtypes of ionotropic P2x-purinoceptors, which might be involved in distinct excitatory processes in these neurons.
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ATP-induced membrane durrents in the submucous neurons of the guinea pig small intestine were studied using the whole-cell patch-clamp recording technique. Being applied at −50 mV. ATP activated an inward non-selective cationic current in... more
ATP-induced membrane durrents in the submucous neurons of the guinea pig small intestine were studied using the whole-cell patch-clamp recording technique. Being applied at −50 mV. ATP activated an inward non-selective cationic current in 68.3% of the investigated neurons. An increase in ATP concentration within the 1–1,000 µM range resulted in the s-like increase in the amplitude of ATP-induced current. The EC50 was 150.0±18.5 µM, while the Hill number was 1.6. The current was selectively activated by ATP and was not blocked by P2 purinoreceptor antagonist suramin (50–300 µM).α,β-Methylene-ATP (100–200 µM) andβ,γ-methylene-ATP (100–200µM), which are P2-purinoreceptor agonists, as well as adenosine (100–300 µM), exerted no effects. Reactive blue 2, if applied up to 4 min, enhanced ATP-induced current, while its longer application partially suppressed this current. In most submucous neurons, acetylcholine (ACh) likewise activated an inward cationic current. The amplitude of ACh-induced current was lower if ACh was applied during a long-lasting application of ATP than if ACh only was applied. Hexamethonium (50 µM), d-tubocurarine (20–40 µM), and trimethaphan (30 µM) completely and reversibly blocked ACh-induced currents, regardless of the presence of ATP, and did not affect ATP-induced currents. The results suggest that ATP-induced currents in submucous neurons are due to activation of a unique type of P2 purinoreceptors, which function in connection with nicotinic ACh receptors.