Inhibition of spontaneous acetylcholine secretion by 2-chloroadenosine as revealed by a protein kinase inhibitor at the mouse neuromuscular junction

1. Previous studies have reported discrepancies in the potencies of A₁ adenosine receptor agonists at mouse motor nerve terminals. In addition, conflicting results on the role of protein kinase A (PKA) in mediating the inhibitory effects of A₁ receptor agonists have been published. We thus decided to investigate the possibility of endogenous control of adenosine receptor sensitivity by protein kinases, using a variety of protein kinase inhibitors in conjunction with the adenosine receptor agonist 2-chloroadenosine (CADO). 2. CADO, at the concentration employed previously to study spontaneous ACh release in the mouse (1 μM), did not inhibit spontaneous ACh release in our experiments. However, a higher concentration of CADO (10 μM) produced highly statistically-significant reductions in spontaneous ACh release. 3. In the presence of the non-selective protein kinase inhibitor, H7 (50 μM), the potency of CADO was increased such that 1 μM CADO now reduced spontaneous quantal ACh release to approximately 63% of control. 4. Both H7, and the selective PKA inhibitor, KT5720 (500 nM) prevented increases in ACh release produced by CPT cyclic AMP (250 μM), suggesting these kinase inhibitors were blocking PKA. In contrast to H7, however, KT5720, did not reveal an inhibitory effect of 1 μM CADO. A number of other non-selective PKA inhibitors also failed to increase the potency of CADO. 5. The results suggest that an endogenous H7-sensitive process modulates the sensitivity of the mouse A₁ adenosine receptor and that the inhibitory effects of CADO are independent of cyclic AMP accumulation or PKA inhibition.