Animals are capable of evaluating sensory cues for possible threats and adapting their behaviours accordingly. Fear learning is an evolutionarily conserved behaviour crucial for animal survival, during which sensory percepts with a negative reinforcing quality, also called unconditioned stimuli (US), influence the valence of initially neutral sensory cues (conditioned stimulus, CS). The lateral amygdala (LA) is thought to integrate information about sensory cues like tones (the CS), and information about primary reinforcers like painful stimuli (the US). Nevertheless, synaptic afferents to the LA and upstream brain areas remain understudied in fear learning. The posterior insular cortex (pInsCx) has been shown to project to the LA and is known to be a multimodal sensory area which processes auditory, somatosensory-nociceptive, and interoceptive information, making it an interesting brain area with a putative role in fear learning. My PhD thesis therefore aimed to investigate a possible connection from the pInsCx to the LA, and the role of this pathway in the fear learning circuitry. I first used anatomical anterograde and retrograde tracing strategies in the mouse to study the afferents to the LA. Besides previously known inputs, I found that the granular (GI) and the dysgranular (DI) areas of the pInsCx strongly project to the anterior LA. With optogenetically-assisted circuit mapping (OACM), I found a glutamatergic monosynaptic connection from the pInsCx to LA principal neurons that, in agreement with the anatomical findings, was localized more to the anterior LA. In comparison, a well-studied auditory thalamic (MGm/PIN) input was localized more to the posterior LA. Despite this regional input specificity, ~30% LA principal neurons received projections from both areas. Thus, in the first part, using anatomical tracing and OACM, I have unambiguously shown a robust functional excitatory connection from the pInsCx (GI/DI) to the LA. In the second part of my thesis, I aimed to characterize the role of the pInsCx and the pInsCx-LA pathway in fear learning. Using archaerhodopsin-mediated optogenetic inhibition, I found that silencing of footshock-driven activity of pInsCx neurons in-vivo, partially blocks auditory-cued fear memory. These results were also supported by chemogenetic silencing of the pInsCx during the training day, which led to a modest impairment of fear retrieval a day later. Next, to investigate the role of the pInsCx-LA projection more specifically, I used a retrograde expression approach to silence LA-projectors during the footshock, which again, suppressed fear memory one day later. In further ex-vivo experiments, I found that EPSCs at the pInsCx-LA connection exhibited an increased AMPA/NMDA ratio following fear learning, indicating that plasticity occurs at this pathway during fear learning. Finally, miniature microscopic Ca2+ imaging of LA neurons combined with halorhodopsin-mediated axon inhibition, showed that the pInsCx-LA excitatory synapse contributes to transmitting footshock information to a sub-population of LA neurons. This study unravels the relationship between two serial nodes, the pInsCx and the LA, in the fear learning network. Thus, the pInsCx-LA connection is a novel pathway in auditory-cued fear memory that is strengthened with learning and that partly provides aversive information to the amygdala.