The molecule responsible for the enzyme activity plasma membrane (PM) aminophospholipid translocase (APLT), which catalyzes phosphatidylserine (PS) translocation from the outer to the inner leaflet of the plasma membrane, is unknown in mammals. A Caenorhabditis elegans study has shown that ablation of transbilayer amphipath transporter-1 (TAT-1), which is an ortholog of a mammalian P-type ATPase, Atp8a1, causes PS externalization in the germ cells. We demonstrate here that the hippocampal cells of the dentate gyrus, and Cornu Ammonis (CA1, CA3) in mice lacking Atp8a1 exhibit a dramatic increase in PS externalization. Although their hippocampi showed no abnormal morphology or heightened apoptosis, these mice displayed increased activity and a marked deficiency in hippocampus-dependent learning, but no hyper-anxiety. Such observations indicate that Atp8a1 plays a crucial role in PM-APLT activity in the neuronal cells. In corroboration, ectopic expression of Atp8a1 but not its close homolog, Atp8a2, caused an increase in the population (V max) of PM-APLT without any change in its signature parameter K m in the neuronal N18 cells. Conversely, expression of a P-type phosphorylation-site mutant of Atp8a1 (Atp8a1*) caused a decrease in V max of PM-APLT without significantly altering its K m. The Atp8a1*-expressing N18 cells also exhibited PS externalization without apoptosis. Together, our data strongly indicate that Atp8a1 plays a central role in the PM-APLT activity of some mammalian cells, such as the neuronal N18 and hippocampal cells. A decade-long search for the enzyme (termed 'PM-APLT') that moves the lipid molecule phosphatidylserine (PS) from the outer to the inner face of the cell membrane has yielded the energy-producing protein Atp8a1. Atp8a1 deficiency is associated with PS exposure in nerve cells and functional impairment in the nerve-filled brain region hippocampus. Atp8a1 is most likely the PM-APLT of nerve cells. © 2011 The Authors.