The Journal of Neurophysiology Vol. 85 No. 1 January 2001, pp. 287-294
Copyright ©2001 by the American Physiological Society

Intraterminal Ca²⁺ and Spontaneous Transmitter Release at the Frog Neuromuscular Junction

 ¹Department of Biological Sciences, University of Denver, Denver 80208; and  ²Department of Physiology and Biophysics, University of Colorado Medical School, Denver, Colorado 80262

Angleson, J. K. and W. J. Betz. Intraterminal Ca²⁺ and Spontaneous Transmitter Release at the Frog Neuromuscular Junction. J. Neurophysiol. 85: 287-294, 2001. We investigated the relationship between intraterminal Ca²⁺ concentration ([Ca²⁺]_i) and the frequency of miniature end plate potentials (MEPPs) at the frog neuromuscular junction by use of ratiometric imaging of fura-2-loaded nerve terminals and intracellular recording of MEPPs. Elevation of extracellular [KCl] over the range of 2-20 mM resulted in increases in [Ca²⁺]_i and MEPP frequency. Loading terminals with the fast and slow Ca²⁺-buffers bis-(o-aminophenoxy)-N,N,N',N'-tetraacetic acid-acetoxymethyl (BAPTA-AM) and EGTA-AM resulted in equivalent reductions in the KCl-dependent increases in MEPP frequency. The [Ca²⁺]_i dependence of MEPP frequency determined by elevation of [Ca²⁺]_i due to application of 0.1-10 µM ionomycin was similar to that determined when [Ca²⁺]_i was raised by increasing extracellular KCl. Measurements in 10 mM extracellular KCl revealed that application of the phorbol ester phorbol 12 myristate 13-acetetate (PMA) caused an increase in MEPP frequency while the inactive analogue, 4-PMA, did not. PMA application also caused an increase in [Ca²⁺]_i. The relationship between [Ca²⁺]_i and MEPP frequency in PMA was the same as was determined by the other methods of raising [Ca²⁺]_i. Under all conditions tested, our data revealed a low [Ca²⁺]_i threshold for activation of transmitter release and are consistent with a K_d for [Ca²⁺]_i on the order of 1 µM.