G protein-coupled receptor kinase-2 confers isoform-specific calcium sensitivity to dopamine D2 receptor desensitization

The dopamine D2 receptor (D2R) functions as an autoreceptor on dopaminergic cell bodies and terminals and as a postsynaptic receptor on a variety of neurons in the central nervous system. As a result of alternative splicing, the D2R is expressed as two isoforms: long (D2LR) and short (D2SR) differing by a stretch of 29 residues in the third intracellular loop, with D2SR being the predominant presynaptic isoform. Recent reports described a Ca2+ sensitivity of the desensitization time course of potassium currents elicited via D2SR, but not via D2LR, when either isoform was selectively expressed in dopaminergic neurons. Here, we aimed to study the mechanism behind this subtype-specific Ca2+ sensitivity. Thus, we measured the desensitization of potassium channel responses evoked by D2LR and D2SR using two-electrode voltage clamp in Xenopus oocytes in the absence and presence of different amounts of β-arrestin2 and G protein-coupled receptor kinase-2 (GRK2), both of which are known to play important roles in D2R desensitization in native cells. We found that co-expression of both GRK2 and β-arrestin2 was necessary for reconstitution of the Ca2+ sensitivity of D2SR desensitization, while D2LR did not display Ca2+ sensitivity under these conditions. The effect of Ca2+ chelation by BAPTA-AM to slow the rate of D2SR desensitization was mimicked by the GRK2 inhibitor, Cmpd101, and by the kinase-inactivating GRK2 mutation, K220R, but not by the PKC inhibitor, Gö6976, nor by the calmodulin antagonist, KN-93. Thus, Ca2+-sensitive desensitization of D2SR appears to be mediated via a GRK2 phosphorylation-dependent mechanism.