Calcitonin gene-related peptide potentiates nicotinic acetylcholine receptor-operated slow Ca²⁺ mobilization at mouse muscle endplates

1. The involvement of calcitonin gene-related peptide (CGRP) in the non-contractile slow Ca²⁺ mobilization induced by prolonged nicotinic stimulation was investigated by measurement of [Ca²⁺](i) levels in mouse single muscle cells (flexor digitorum brevis; FDB) loaded with a Ca²⁺ indicator fluo-3 using confocal laser scanning microscopy. 2. CGRP (3-30 nM) potentiated acetylcholine (ACh, 1 μM)-elicited slow Ca²⁺ mobilization in a concentration-dependent manner. 3. The potentiation by CGRP of the slow Ca²⁺ component was greatly depressed by a competitive nicotinic antagonist (+)-tubocurarine (5 μM). The Ca²⁺ channel blocker nitrendipine (1 μM) affected neither ACh responses nor the CGRP potentiation. 4. The slow Ca²⁺ component was completely abolished by reducing [Ca²⁺](o) from 2.5 to 0.25 mM whereas the fast component was not affected. The CGRP-induced potentiation of slow Ca²⁺ signal was also depressed by decreasing [Ca²⁺](o). 5. Isoproterenol (30 μM) and 8-bromo-adenosine 3',5'-cyclic monophosphate (1 mM) potentiated the ACh-elicited slow Ca²⁺ response. The potentiation by CGRP of the slow Ca²⁺ component was completely abolished by a protein kinase-A inhibitor H-89 (1 μM). 6. These findings indicate that CGRP potentiates the nicotinic ACh receptor-operated slow Ca²⁺ signal via the activation of protein kinase-A system at the skeletal muscle endplates.