Effects of ruthenium red (RuR) on adenylates, plasma membrane potential (delta psi p) and cytosolic free calcium concentration ([Ca2+]c) in cortical synaptosomes from guinea-pig were investigated. Ten micromoles of RuR did not affect either energy levels as indicated by ATP/ADP ratio or the basal delta psi p. The resting [Ca2+]c in the presence of RuR was unchanged, but above 5 microM it inhibited by more than 50% of the voltage-activated increase in [Ca2+]c by K+-depolarization. In another experiment the potencies of 10 microM RuR and 100 microM verapamil to inhibit high K+-induced increase in [Ca2+]c were compared. It was found that either produced 59% inhibition and this inhibition was not potentiated by the substances together (65% inhibition). The extent of depolarisation of delta psi p by high external K+ was independent of the presence of RuR. RuR blocked only 20% of the increase in [Ca2+]c by veratridine treatment, indicating that Ca2+ accumulation into synaptosomal cytoplasm by veratridine involves some additional mechanisms other than depolarisation of delta psi p. The mechanism of inhibition of evoked release of neurotransmitters by RuR is discussed.