Differential effects of divalent cations on spontaneous and evoked glycine release from spinal interneurons

The effects of Ca2+, Sr2+, and Ba2+ on spontaneous and evoked glycinergic inhibitory postsynaptic currents (mIPSCs and eIPSCs) were studied using the "synaptic bouton" preparation of rat spinal neurons and conventional whole cell recording under voltage-clamp conditions. In response to application of Ca2+-free solution, the frequency of mIPSC initially rapidly decreased to 40 approximately 50% of control followed by a gradual further decline in mIPSC frequency to approximately 30% of control. Once mIPSC frequency had significantly decreased in Ca2+-free solution, application of Ca2+, Sr2+, or Ba2+ increased mIPSC frequency. The rank order of effect in restoring mIPSCs was Ba2+>>Ca2+>Sr2+. Moreover, the application of excess external [K+]o solution (30 mM) containing Sr2+ or Ba2+ after 2 h in Ca2+-free solution also increased mIPSC frequency in the order Sr2+>or==Ba2+>Ca2+. The mean mIPSC amplitude was not affected at all. In contrast, eIPSCs produced by focal stimulation of single boutons were completely abolished in Ca2+-free solution or when Ca2+ was replaced by Sr2+ or Ba2+ (2 mM each). However, eIPSCs were restored in increased concentrations of Sr2+ or Ba2+ (5 mM each). The results show that these divalent cations affect mIPSC and eIPSCs differently and indicate that the mechanisms underlying transmitter release that generates eIPSCs and mIPSC in presynaptic nerve terminals are different. The different mechanisms might be explained by the different sensitivity of synaptotagmin isoforms to Ca2+, Sr2+, and Ba2+.