The mossy fiber synapse between dentate granule cells and CA3 pyramidal cells in the guinea pig hippocampus shows a robust short-term synaptic enhancement. We have simultaneously measured presynaptic residual free calcium ([Ca2+]i) and postsynaptic field potentials at this synapse to examine the role of [Ca2+]i in this enhancement. Single action potentials produced an increase in [Ca2+]i of 10–50 nM that decayed to resting levels with a time constant of about 1 sec. Trains of action potentials produced larger [Ca2+]i increases that returned more slowly to resting levels. Following the onset of moderate frequency stimulus trains (0.1–5 Hz), synaptic transmission and [Ca2+]i both increased and eventually plateaued. During the steady-state phase a linear relationship between [Ca2+]i and synaptic enhancement was observed. During the initial buildup, however, [Ca2+]i rose more rapidly than synaptic enhancement. Similarly, during the decay phase immediately following termination of a stimulus train, [Ca2+]i returned to prestimulus levels faster than synaptic enhancement. High concentrations of the calcium buffer EGTA in the presynaptic terminal slowed the buildup and decay of both [Ca2+]i and synaptic enhancement produced by stimulus trains. Under these conditions, the time course of [Ca2+]i and synaptic enhancement were well matched. This suggests that, despite the differences in kinetic rates observed for normal buffering conditions, increases in [Ca2+]i play a causal role in short-term enhancement. An increase in [Ca2+]i of 10–30 nM produced a twofold enhancement. We propose a simple kinetic model to explain these results. The model assumes that synaptic enhancement is controlled by a Ca-dependent first-order reaction. According to this scheme, a change in [Ca2+]i alters neurotransmitter release, but the slow kinetics of the underlying reaction introduces a temporal filter, producing a delay in the change in synaptic enhancement.