In slices of adult rat somatomotor cortex, paired intracellular recordings determined the properties of a novel class of excitatory connection, that of presynaptic pyramidal axon collaterals onto burst firing, spiny inhibitory interneurons. Single axon excitatory postsynaptic potentials were brief in time course and displayed conventional voltage relations, increasing in amplitude with membrane hyperpolarization with no change in time course. Excitatory postsynaptic potential amplitude distributions were not skewed. Paired pulse facilitation was profound at interspike intervals < 50 ms, but not altered by raising extracellular [Ca2+] from 2.5 to 5 mM, despite an apparent increase in release probability. Raising presynaptic firing frequency did however produce an increase in excitatory postsynaptic potentials elicited by first spikes that was associated with a decline in excitatory postsynaptic potentials elicited by second and third spikes in brief trains of presynaptic spikes. That this pattern of synaptic activity may result from low probabilities of transmitter release is discussed. It is proposed that while raising Ca2+ and increasing presynaptic firing both increase release probability, repetitive presynaptic firing raises probability more effectively than does raising extracellular [Ca2+]. However, concomitant exhaustion of readily releasable transmitter at higher firing rates may partially obscure this effect. It is concluded that the major differences in the firing rate- and firing pattern-dependent properties of pyramid-pyramid and pyramid-interneuron connections are due to the typically lower release probability at synapses onto interneurons. The accompanying paper describes the morphology of these connections.