Simultaneous measurement of intracellular Ca²⁺ and asynchronous transmitter release from the same crayfish bouton

1. A technique has been developed to monitor neurotransmitter release simultaneously with intracellular Ca²⁺ concentration ([Ca²⁺](i)) in single release boutons whose diameters range from 3 to 5 μm. 2. Using this technique, we have found a highly non-linear relationship between the rate of asynchronous release and [Ca²⁺](i). The Hill coefficient lies between 3 and 4. 3. The affinity (K(d)) of the putative release-related Ca²⁺ receptor for asynchronous release was calculated to be in the range of 2-4 μM. 4. The same range of values of Hill coefficient and K(d) were obtained when [Ca²⁺](i) was elevated both by bath application of ionomycin and by repetitive stimulation at high frequency 5. Our results show that the Ca²⁺ receptor(s) associated with asynchronous release exhibits high affinity for Ca²⁺.