Guiding G protein signaling by target enhancement of GPCRs

Activation of G protein coupled receptors coupling to the Gi/o pathway leads to the activation of G protein-activated inward rectifier potassium channels (GIRK) in a fast membrane-delimited manner in excitable cells. Activation of GIRK causes the hyperpolarization of the cell membrane, where hyperpolarization is dependent on te availability of Gi/o coupled GPCRs and GIRK. In particular, in optogenetic and chemogenetic experiments neuronal silencing depends on downstream targets of Gi/o-coupled GPCRs. To selectively enhance Gi/o mediated GIRK currents, we created expression cassettes consisting of a homomer forming GIRK subunit and various light-activated Gi/o-coupled GPCRs (Melanopsin, Long-wave-sensitive opsin 1, Parapinopsin or Opsin 7b). We demonstrate that light-activation of the GIRK/GPCR constructs induces robust GIRK currents in human embryonic kidney 293 cells, cardiomyocytes and cerebellar Purkinje cells and changes the net effect of G protein signaling of the promiscuous Opn4L from a Gq/11 mediated excitation towards an Gi/o mediated inhibition. Thus, our tools enhance target selectivity and improve optogenetic control of the Gi/o pathway by light in excitable cells.