MUSCARINIC AUTOINHIBITION RELEASE IN MOUSE ATRIA IS NOT TRANSDUCED THROUGH CYCLIC-AMP OR PROTEIN-KINASE-C

1 The present study investigated the second messenger pathways that ma y mediate muscarinic receptor autoinhibition of acetylcholine release in mouse atria. The stimulation-induced (S-I) outflow of radioactivity from mouse isolated atria incubated with [H-3]-choline was Ca2+ depen dent and tetrodotoxin-sensitive and was used as an index of neuronal a cetylcholine release. 2 The cell permeable analogue of cyclic AMP, 8-b romocyclic AMP (1 x 10(-3)M) enhanced the S-I outflow of radioactivity (33%), lower concentrations having no effect. Similarly, the adenylat e cyclase activator forskolin (1x10(-5)M) had a small facilitatory eff ect on acetylcholine release. On the other hand the phosphodiesterase inhibitor 3-isobutylmethylxanthine (1 x 10(-4)M) had no effect on the S-I outflow of radioactivity. Together these results suggest that the adenylate cyclase/cyclic AMP system does not have an appreciable role in the modulation of acetylcholine release. 3 The protein kinase C act ivator phorbol dibutyrate (0.1-3 x 10(-6)M) enhanced the S-I acetylcho line release (maximally by 45%). The effects of phorbol dibutyrate wer e attenuated by the protein kinase inhibitor staurosporine (1 x 10(-7) M), which by itself had no effect on the S-I outflow of radioactivity. This latter result suggests chat there is no tonic activation of prot ein kinase C during acetylcholine release. 4 Atropine (1 x 10(-7)M) ma rkedly enhanced (232%) the S-I outflow of radioactivity, presumably by preventing feedback inhibition on acetylcholine release through preju nctional muscarinic receptors. This effect is unlikely to involve aden ylate cyclase or protein kinase C since it was far greater than the ef fects of activation of either system with forskolin and phorbol dibuty rate, respectively. Furthermore, the facilitatory effect of atropine w as not attenuated by staurosporine, which although a protein kinase C inhibitor, is also an effective inhibitor of cyclic AMP dependent prot ein kinase (protein kinase A).