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 explained by a direct effect of ATP on nicotinic acetylcholine receptors (nAChRs). Reactive blue 2, the P2y purinoceptor antagonist, did not abolish inhibition of the ACh current by ATP. Alpha,beta-Imido-ATP and adenosine caused no membrane current responses and did not influence the ACh currents. These results suggest that the activity of the nAChRs in the SMP neurons is strongly suppressed by raised intracellular Ca2+ level, without involvement of protein kinases A and C, and may involve the participation of tyrosine kinase. The activity of nAChRs is also influenced by the activity of P2 purinoceptors; the mechanisms responsible for this influence are not yet clear. So, the activity of the SMP neuronal nAChRs is relatively independent on the intracellular signaling known to influence many other groups of transmitter-gated receptors of neuronal membrane.