Modulation of the kinetics of evoked quantal release at mouse neuromuscular junctions by calcium and strontium Ellya A. Bukharaeva,* ,  Dmitry Samigullin,* Eugeny E. Nikolsky* ,  and Lev G. Magazanikà *Institute of Biochemistry and Biophysics, Russian Academy of Sciences, Kazan, Russia  Kazan State Medical University, Kazan, Russia àI.M. Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, St-Petersburg, Russia Abstract The effects of calcium and strontium on the quantal content of nerve-evoked endplate currents and on the kinetic parameters of quantal release (minimal synaptic delay, value of main mode of synaptic delay histogram, and variability of synaptic delay) were studied at the mouse neuromuscular synapse. At low calcium ion concentrations (0.2–0.6 mmol/L), evoked signals with long synaptic delays (several times longer than the value of main mode) were observed. Their number decreased substantially when [Ca 2+ ] o was increased (i.e. the release of transmitter became more synchronous). By con- trast, the early phase of secretion, characterized by minimal synaptic delay and accounting for the main peak of the syn- aptic delay histogram, did not change significantly with increasing [Ca 2+ ] o . Hence, extracellular calcium affected mainly the late, ‘asynchronous’, portion of phasic release. The average quantal content grew exponentially from 0.09 ± 0.01 to 1.04 ± 0.07 with the increase in [Ca 2+ ] o without reaching saturation. Similar results were obtained when calcium was replaced by strontium, but the asynchronous portion of phasic release was more pronounced and higher strontium con- centrations (to 1.2–1.4 mmol/L) were required to abolish responses with long delays. Treatment of preparations with 1,2-bis(2-aminophenoxy)ethane-N,N,N¢,N¢-tetraacetic acid tetrakis acetoxymethyl ester (BAPTA-AM) (25 lmol/L), but not with ethylene glycol-bis(2-aminoethylether)-N,N,N¢,N¢- tetraacetic acid acetoxymethyl ester (EGTA-AM) (25 lmol/L), abolished the responses with long delays. The dependence of quantal content and synchrony of quantal release on calcium and strontium concentrations have quite different slopes, suggesting that they are governed by different mechanisms. Keywords: BAPTA-AM, calcium, EGTA-AM, quantal release, strontium, time course of the evoked quanta release. J. Neurochem. (2007) 100, 939–949. At the neuromuscular junction, transmitter quanta are released with variable delay following an action potential in the motor nerve terminal. An accurate estimate of synaptic delay can be defined using extracellular recording as the interval between the peak of the pre-synaptic action potential and the onset of a post-synaptic response (Katz and Miledi 1965). The duration of this delay reflects the rates of depolarization–release coupling (Katz and Miledi 1965; Lin and Faber 2002). The initial phasic quantal release is fast but not absolutely synchronous. At room temperature, it usually begins 0.2– 0.5 ms after the peak of action potential (minimal synaptic delay), rapidly reaches a maximum, and then decays during several milliseconds (Katz and Miledi 1965; Barrett and Stevens 1972a; Hochner et al. 1991). Analysis of the distri- bution of delays of individual quantal events gives insight into the intraterminal processes determining the time course of release probability (Barrett and Stevens 1972a; Van der Kloot 1988; Schneggenburger and Neher 2000; Lin and Faber 2002). The dependence of the quantal content of release on calcium is well known (Katz 1969; Van der Kloot and Molgo 1994; Zucker 1996; Augustine 2001, for review), but evidence concerning the influence of calcium on the latency of release is still contradictory. A clear dependence of synaptic delay on calcium concentration at release sites was shown using a calcium ion uncaging technique at the Received July 14, 2006; revised manuscript received September 4, 2006; accepted September 18, 2006. Address correspondence and reprint requests to Lev G. Magazanik, I.M. Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, Thorez pr., 44, St-Petersburg 194223, Russia. E-mail: magazanik@mail.ru Abbreviations used: BAPTA-AM, 1,2-bis(2-aminophenoxy)ethane- N,N,N¢,N¢-tetraacetic acid tetrakis acetoxymethyl ester; EGTA-AM, ethylene glycol-bis(2-aminoethylether)-N,N,N¢,N¢-tetraacetic acid acet- oxymethyl ester; EPC, endplate current; NAP, nerve action potential; SA, stimulus artifact. Journal of Neurochemistry , 2007, 100, 939–949 doi:10.1111/j.1471-4159.2006.04282.x Ó 2006 The Authors Journal Compilation Ó 2006 International Society for Neurochemistry, J. Neurochem. (2007) 100, 939–949 939