GIRK2-containing channels are enriched near excitatory synapses15, 39, and some overlap between GIRK2 and PSD-95 was reported in cultured hippocampal neurons35. As the PDZ interaction motif found in GIRK2c may promote the association of GIRK channels with excitatory synapses, we next compared the overlap between endogenous PSD-95 and either GIRK2a or GIRK2c in Girk2 −/− pyramidal neurons (Fig. 4A,B). No significant difference in the extent of overlap between PSD-95 and either GIRK2a or GIRK2c was detected at baseline (Fig. 4C, control). We also explored the possibility that incubation with morphine, a treatment previously shown to increase the overlap between GIRK2 and PSD-95 in cultured hippocampal neurons35, might reveal an isoform-dependent difference. Consistent with published observations, we found that morphine treatment increased total GIRK2 protein level by approximately 2-fold relative to vehicle-treated controls (Supplementary Fig. S4A,B), while having no apparent effect on PSD-95 labeling intensity (Supplementary Fig. S4C). Morphine treatment increased the overlap between PSD-95 and both GIRK2 isoforms relative to control conditions, with GIRK2c showing a small but significantly higher degree of overlap with PSD-95 than did GIRK2a following treatment (Fig. 4C, morphine; Supplementary Fig. S4D,E).
