Key points Post‐tetanic potentiation (PTP) is attributed mainly to an increase in release probability (Pr) and/or readily‐releasable pool (RRP) in many synapses, but the role of endocytosis in PTP is unknown. Using the calyx of Held synapse from tissue‐specific dynamin‐1 knockout (cKO) mice (P16–20), we report that cKO synapses show enhanced PTP compared to control. We found significant increases in both spontaneous excitatory postsynaptic current (spEPSC) amplitude and RRP size (estimated by a train of 30 APs at 100 Hz) in cKO over control during PTP. Actin depolymerization blocks the increase in spEPSC amplitude in both control and cKO, and it abolishes the enhancement of PTP in cKO. PTP is sensitive to the PKC inhibitor GF109203X in both control and cKO. We conclude that an activity‐dependent quantal size increase contributes to the enhancement of PTP in cKO over control and an altered endocytosis affects short‐term plasticity through quantal size changes. Abstract High‐frequency stimulation leads to post‐tetanic potentiation (PTP) at many types of synapses. Previous studies suggest that PTP results primarily from a protein kinase C (PKC)‐dependent increase in release probability (Pr) and/or readily‐releasable pool (RRP) of synaptic vesicles (SVs), but the role of SV endocytosis in PTP is unknown. Using the mature calyx of Held (P16–20), we report that tissue‐specific ablation of dynamin‐1 (cKO), an endocytic protein crucial for SV regeneration, enhances PTP in cKO over control. To explore the mechanism of this enhancement, we estimated the changes in paired‐pulse ratios (PPRs) and RRP size during PTP. RRP was estimated by the back‐extrapolation of cumulative EPSC amplitudes during a train of 30 action potentials at 100 Hz (termed RRPtrain). We found an increase in RRPtrain during PTP in both control and cKO, but no significant changes in the PPR. Moreover, the amplitude and frequency of spontaneous excitatory postsynaptic currents (spEPSCs) increased during PTP in both control and cKO; however, the spEPSC amplitude in cKO during PTP was significantly larger than in control. Actin depolymerization reagent latrunculin‐B (Lat‐B) abolished the activity‐dependent increase in spEPSC amplitude in both control and cKO, but selectively blocked the enhancement of PTP in cKO, without affecting PTP in control. PKC inhibitor GF109203X nearly abolished PTP in both control and cKO. These data suggest that the quantal size increase contributes to the enhancement of PTP in dynamin‐1 cKO, and this change depends on strong synaptic activity and actin polymerization.