Cortical fast-spiking (FS) neurons generate high-frequency action potentials (APs) without apparent frequency accommodation, thus providing fast and precise inhibition. However, the maximal firing frequency that they can reach, particularly in primate neocortex, remains unclear. Here, by recording in human, monkey, and mouse neocortical slices, we revealed that FS neurons in human association cortices (mostly temporal) could generate APs at a maximal mean frequency (Fmean) of 338 Hz and a maximal instantaneous frequency (Finst) of 453 Hz, and they increase with age. The maximal firing frequency of FS neurons in the association cortices (frontal and temporal) of monkey was even higher (Fmean 450 Hz, Finst 611 Hz), whereas in the association cortex (entorhinal) of mouse it was much lower (Fmean 215 Hz, Finst 342 Hz). Moreover, FS neurons in mouse primary visual cortex (V1) could fire at higher frequencies (Fmean 415 Hz, Finst 582 Hz) than those in association cortex. We further validated our in vitro data by examining spikes of putative FS neurons in behaving monkey and mouse. Together, our results demonstrate that the maximal firing frequency of FS neurons varies between species and cortical areas.