INTRODUCTION: Chemokines, together with neurotransmitters and hormones, are signaling molecules that play a key role in the maintenance of the neuro‑immune‑endocrine system homeostasis. Accumulating evidence shows that they can modulate the activity of neurons through different mechanisms. One of their members, CX3CL1, and its cognate receptor, CX3CR1, play a crucial role in neuronal‑microglia signaling. AIM(S): As the amygdala is a relevant structure for integrating stress signaling as well as inflammatory responses from the periphery, this study aimed to elucidate the role of the CX3CL1/CX3CR1 axis on circuits within the amygdala. METHOD(S): We used whole-cell patch-clamp and immunohistochemistry and focused on two nuclei of the amygdala: the basolateral (BLA, main input structure), and central (CeA, output structure) nuclei. Electrophysiological recordings were performed using acute brain slices (300 μm) containing the BLA and CeA. Recordings of both spontaneous inhibitory and excitatory currents (sIPSC/sEPSC), as well as, basal membrane properties of recorded cells were collected during baseline and after CX3CL1 (2nM) application. The specificity of observed effects was investigated using the same experimental protocol with additional incubation in a CX3CR1 antibody. Additionally, to specify the cell types that express CX3CR1 and CX3CL1, appropriate immunostainings were performed. RESULTS: Our results revealed that CX3CL1 was mostly expressed within the BLA and it significantly hyperpolarized resting membrane potential of most of the recorded principal cells (70%) and decreased their excitability; however, CX3CL1 did not alter their membrane resistance. CONCLUSIONS: Our data show that CX3CL1 has a profound effect on synaptic activity in the rat amygdala, indicating that this protein can be an active modulator of neuronal activity in the fear-related response circuitry, which may have significant scientific and therapeutic implications. FINANCIAL SUPPORT: Supported by National Science Centre, grant 2016/21/N/NZ4/03621.