In synaptic transmission, the fast, precise, and synchronous release of neurotransmitters relies on a tight coupling between the synaptic release machinery, the synaptic vesicles, and the voltage-gated calcium channels (P/Q-type). Scaffolding molecules, including Rab-Interacting molecules (RIM), RIM-Binding Protein (RBP), ELKS, Munc13, Bassoon/ Piccolo, and Liprin-α are the main known regulators of this coupling. Specifically, RIM and RBP have been shown to uniquely regulate synaptic vesicle docking and fusion, since no synaptic activity remains in RIM/RBP-deficient neurons (Acuna et al, 2016). The purpose of our study is to investigate the involvement of the scaffold protein RBP in voltage-gated Ca2+-channel function using an autaptic model of mouse hippocampal glutamatergic neurons with conditional genetic deletions of RIM1α, RIM1β, RBP1, and RBP2. Synaptic vesicles can be artificially brought into the vicinity of Ca2+-channels by expressing the fusion protein Zn-β4 composed of the RIM1α Zinc-finger domain and the β4 subunit of Ca2+-channels (Tan et al, 2020). When expressed in RIM/RBP-deficient neurons, Zn-β4 is insufficient to restore Ca2+-dependent neurotransmitter release, as measured electrophysiologically. Using calcium imaging of the GCAMP6f signal, we show that the influx of Ca2+ occurring during cell depolarisation is also significantly impaired. Immunocytochemistry reveals that both Munc13 and P/Q-Ca2+-channel expression is reduced in RIM/RBP-deficient neurons, whereas the levels are restored upon the Zn-β4 expression. Most recently, we found that co-expressing the Zn-β4 fusion protein with RBP in RIM/RBP-deficient neurons rescues Ca2+-evoked response to wild-type levels, strongly suggesting a critical function of RBP in presynaptic Ca2+-channel gating.