Possibility of multiquantal transmission at single inhibitory synapse in cultured rat hippocampal neurons

Abstract

Miniature, spontaneous and evoked inhibitory postsynaptic currents were studied using the whole-cell patch-clamp technique on synaptically connected cultured hippocampal neurons, at a holding potential of −75 mV. All experiments were done in tetrodotoxin-containing solution to exclude an action potential generation. Spontaneous miniature inhibitory postsynaptic currents were observed in Ca 2+-free solution. The distribution of miniature inhibitory postsynaptic currents was skewed to larger current amplitudes and could be fitted reliably by one Gaussian with the mean at 10.0±1.2 pA ( n=7). Spontaneously occurring whole-cell spontaneous inhibitory postsynaptic currents were recorded in physiological solution (Ca 2+ 2 mM). The average amplitude of spontaneously occurring currents depended on membrane potential and reversed at −18±5 mV ( n=5). The amplitude distribution of spontaneous inhibitory postsynaptic currents had one peak clearly detectable with the mean of 20.0±2.0 pA ( n=6) or 10.0±2.0 pA ( n=2). Inhibitory postsynaptic stimulus-evoked currents arose in responses to gradual activation of neurotransmitter release by direct extracellular electrical stimulation of a single presynaptic bouton by short depolarizing pulses. The current–voltage relation of the averaged amplitudes of evoked inhibitory postsynaptic currents was linear and reversed at potential predicted by the Nernst equation for corresponding intra- and extracellular Cl − concentrations. The time-course of decay of miniature, spontaneous and evoked inhibitory postsynaptic currents was fitted by a sum of two exponents and their time-constants were the same in the range of standard deviation. The stimulus-evoked inhibitory postsynaptic currents fluctuated with regard to the discrete aliquot values of their peak amplitudes in all the investigated synapses from a measurable minimum of about 8 pA to 200 pA. The evoked inhibitory postsynaptic currents were assumed as superimposition of statistically independent quantal events. Fitting amplitude histograms of evoked inhibitory postsynaptic currents with several Gaussian curves resulted in peaks that were equidistant with the mean space of 20±3 pA ( n=10), which was assumed as one quantum (quantum size) to construct the Poisson's distribution. A possibility of simultaneous multiquantal release at single inhibitory synapses of rat hippocampal neurons was discussed.