(A) Schematic illustrating the receptor composition of presynaptic inhibition across rod (RBC) and cone bipolar cell (CBC) axon terminals in wildtype adult littermate control (WT) and KO retina; T6 CBC refers to Type 6 ON CBC (B) α1-subunit-containing GABAA receptor (GABAA α1) immunolabeling (yellow) and ON BC labeling (tdTomato; red) in outer and inner plexiform layers (OPL and IPL respectively) of WT retina and Ai9/Grm6Cre/Gabrg2 (KO) retina. In the KO, GABAAα1 immunofluorescence is present in the OFF lamina of the IPL but not in the ON lamina. (C) (Left) GABAAα1 receptor (yellow; signal within terminals) and protein kinase C (PKC; red) immunolabeling of RBC terminals in WT retina. The merged panel consists of the PKC signal and the receptor signal within PKC positive RBC terminals. (Right) Image of KO retina shows reduced GABAAα1 receptor immunofluorescence within RBC terminals. (Far right) Quantifications of receptor expression confirmed a significant reduction of GABAAα1 expression in the KO (mean ± sem = 0.8 ± 0.1) retina relative to WT (mean ± sem = 4 ± 0.1). (D) (Left) Immunolabeling of RBC terminals (PKC;red) in WT retina with antibodies against the ρ-subunit-containing GABAC receptor (GABACρ; yellow – signal within RBC terminals). (Right) Image shows GABACρ immunoreactivity within RBC terminals in the KO retina. (Far right) Quantification of RBC terminal GABACρ receptor expression in KO (mean ± sem = 7.9 ± 0.6) retina relative to WT (mean ± sem = 8.4 ± 0.6). (E) Exemplar traces of evoked responses of an RBC after GABA puff application at its axon terminal. WT (Left, black trace); KO (Right, red traces). TPMPA (GABAC receptor antagonist) and GABAzine (GABAA receptor antagonist) were used to pharmacologically isolate GABAA and GABAC receptor-mediated components of the evoked responses. The GABAA component is revealed after application of TPMPA (labeled +TPMPA) and is eliminated upon the addition of GABAzine (TPMPA + GABAzine; labeled +GABAzine). Note the reduction of the GABAA receptor-mediated component in the KO relative to the WT. (F) Bar graph quantifying the GABAA and GABAC-mediated component of RBC evoked responses in WT (black) and KO (red) retina. The mean ± sem peak amplitudes of GABAA + GABAC currents were 202.5 ± 5.5 pA in WT retina and 112.3 ± 5.6 pA in KO retina. The mean ± sem peak amplitudes of GABAA currents were 96.8 ± 5.2 pA in WT retina and 15.2 ± 1.5 pA in KO retina. The mean ± sem peak amplitudes of GABAC currents were 105.8 ± 1.8 pA in WT retina and 97 ± 4.3 pA in KO retina. Note that the significant reduction in the total response (GABAA + GABAC) in the KO can be attributed to the reduction in the GABAA-mediated component. In all figures, error bars indicate sem and ‘n’ refers to the number of cells analyzed except 1C, 1D, and Figure 1—figure supplement 1E occupancy quantifications where ‘n’ refers to the number of retinas analyzed.