Chandelier cells (ChCs) that are present only in cortical structures selectively form synapses on the axon initial segments (AISs) of principal neurons (PNs) – the site of action potential generation. Therefore, these GABAergic cells are in the position to control the spiking of their postsynaptic partners in the most powerful way. Here, we compared anatomical and electrophysiological features of ChCs sampled in the medial prefrontal cortex (mPFC) of the Nkx2.1-Cre and PVeGFP mouse lines.By performing whole-cell recordings from ChCs in acute prefrontal cortical slices, we found that ChCs labeled with the two approaches differed both in their passive and active membrane properties. Our paired recordings revealed that ChCs of the Nkx2.1-cre line evoke unitary inhibitory postsynaptic currents (uIPSCs) in PNs with significantly slower kinetic properties, but the short-term dynamics of these synapses are similar to those recorded from the PVeGFP mouse line. Post hoc analysis of ChC morphological properties uncovered that the distribution of axons and dendrites of ChCs sampled in the two transgenic mouse lines showed some differences between layers, indicating potentially distinct input and output features. Additionally, we observed that the length of innervated AISs or the percentage of AISs targeted by ChCs are comparable, but ChCs labeled in the Nkx2.1-cre line establish fewer boutons on individual AISs.Taken together, these results show that ChCs in the mPFC show anatomical and electrophysiological differences in their inhibitory connections. The variability of ChC properties in prefrontal cortical microcircuits might be an important factor in the control of local network operation.