Recent findings suggest that COMP360 (Compass Pathways’ investigational proprietary formulation of synthetic psilocybin) modulates negative affective biases in a rat model. Based on our current behavioural data, the mechanism is hypothesised to depend on psilocin, the active metabolite of psilocybin, acting as a partial agonist at both 5-HT2A and 5-HT1A receptors and altering neuronal activity, particularly in circuits which regulate affect and include the medial prefrontal cortex (mPFC). In the present study, we investigated effects of acute psilocin on stimulus-evoked excitatory postsynaptic potentials on L5 pyramidal neurons in prelimbic cortex of rat brain slices. We show that psilocin selectively depressed glutamatergic transmission at synapses onto cortico-amygdalar neurons, labelled with a retrograde fluorescent tracer, while facilitating synaptic transmission onto non-labelled L5 pyramidal cells. These effects of psilocin on cortico-amygdalar neurons could be blocked by antagonists of either the 5-HT2A or 5-HT1A receptor.24 hours after a single low dose (0.3 mg/kg) of COMP360 psilocybin given to rats in vivo, a significant increase in the amplitude of miniature inhibitory post-synaptic currents was observed in ex vivo brain slices, only in cortico-amygdalar neurons, and not cortico-cortical or non-labelled cells. These findings suggest that COMP360 psilocybin’s ability to modulate affective biases may involve modification of synapses on cortico-amygdalar cells in the mPFC, and long-term changes at synapses in this affect-associated circuit implicate alterations to cortical inhibition. The precise contributions of changes to excitatory and inhibitory synapses onto cortico-amygdalar neurons, and how these alterations contribute to behaviour are areas that warrant further research.