MAM (meprin/A5 protein/receptor protein tyrosine phosphatase mu) domain glycosylphosphatidylinositol anchor 1 (MDGA1), a unique cell surface glycoprotein, is similar to Ig-containing cell adhesion molecules that influence neuronal migration and process outgrowth. We show in postnatal mice that MDGA1 is expressed by layer 2/3 neurons throughout the neocortex. During development, MDGA1 is expressed in patterns consistent with its expression by migrating layer 2/3 neurons, suggesting a role for MDGA1 in controlling their migration and settling in the superficial cortical plate. To test this hypothesis, we performed loss-of-function studies using RNA interference (RNAi) targeting different sequences of mouse MDGA1. RNAi or empty vectors were coelectroporated with an enhanced green fluorescent protein reporter in utero into the lateral ventricle at embryonic day 15.5 to transfect progenitors of superficial layer neurons; the distributions of transfected neurons were analyzed late on postnatal day 0. We found a direct correlation between effectiveness of an RNAi in suppressing MDGA1 expression and disrupting migration of superficial layer neurons. An RNAi with no effect on MDGA1 expression has no effect on the migration. In contrast, an RNAi that suppresses MDGA1 expression also blocks proper migration of transfected superficial layer neurons, with essentially all transfected cells found deep in the cortical plate or beneath it. This migration defect is rescued by cotransfection of a rat MDGA1 expression construct along with the effective RNAi, confirming that the RNAi effect is specific to diminishing mouse MDGA1 expression. RNAi transfections of deep layer neurons that do not express MDGA1 do not significantly affect their migration. We conclude that MDGA1 acts cell autonomously to control the migration of MDGA1-expressing superficial layer cortical neurons.