ITCs have high intrinsic connectivity within the Im2226 and it is possible endogenously released opioids could also inhibit these GABAergic synapses. To study Im-Im neuron synapses exclusively, excluding GABAergic inputs from other regions such as the CeA, we performed paired recordings between synaptically coupled Im neurons (Fig. 6a). We found that exogenous ME (10 μM) strongly inhibited paired IPSCs (Fig. 6b,c) and in three of the four synaptic pairs, ME also increased synaptic failure rate (CTL: 3.75±0.1% failure, ME: 45.6±0.4% failure, for example, Fig. 6b). This inhibition was reversed by either CTAP (Fig. 6b) or wash of ME and likely resulted from ME acting pre-synaptically to reduce GABA release as ME significantly increased the PPR (Fig. 6d). Unfortunately, the success rate for obtaining viable synaptically coupled pairs of Im neurons was low (2.4%) and as a result, we were unable to test the actions of endogenously released opioids using this approach. Instead, we recorded local Im synaptic activity by electrically stimulating within the Im to evoke eIPSCs (Fig. 6e). Exogenous ME (10–30 μM) robustly inhibited eIPSCs (Fig. 6f,g) and this was reversed by the MOR antagonist CTAP, but not by DOR or KOR antagonists (Fig. 6h). Surprisingly however, we did not find a consistent change in PPR (Fig. 6i), which was in distinct contrast to opioid regulation of glutamatergic inputs (Fig. 2e) and paired Im neuron recordings (Fig. 6a–d). Although unexpected, we reasoned that since ME almost completely eliminated paired Im-Im inputs (Fig. 6b,c), this may allow MOR insensitive GABAergic inputs with different synaptic characteristics to dominate the remaining eIPSC. Indeed, this is supported by the change in eIPSC kinetics before and after ME treatment, in which both the 10–90% rise time (baseline: 1.7±0.2 ms; ME: 1.3±0.1 ms, P<0.01, paired t-test) and the weighted decay time constant (τw; baseline: 34.3±4.8 ms; ME: 17.8±2.8 ms, P<0.01, paired t-test) showed a significant decrease.