BackgroundIn many cells, depletion of intracellular calcium (Ca2+) reservoirs triggers Ca2+ entry through store-operated Ca2+ channels in the plasma membrane. However, the mechanisms of agonist-induced calcium entry (ACE) in keratinocytes are not fully understood.ObjectivesThis study was designed to determine if pharmacological inhibition of calcium-independent phospholipase A (iPLA2) impairs ACE in normal human epidermal keratinocytes.MethodsConfocal laser scanning microscopy was used to monitor the dynamics of Ca2+ signalling in keratinocytes loaded with the calcium-sensitive dye Fluo-4. Cells were stimulated with extracellular nucleotides [adenosine triphosphate (ATP) or uridine triphosphate (UTP)] or with lysophosphatidic acid (LPA), a bioactive lipid that regulates keratinocyte proliferation and differentiation.ResultsBoth ATP and UTP induced Ca2+ release in primary human keratinocytes. This was not followed by robust Ca2+ influx when the experiments were performed in low Ca2+ (70 μmol L−1) medium. Upon elevation of extracellular Ca2+ to 1·2 mmol L−1, however, a biphasic response consisting of an initial Ca2+ peak followed by an elevated plateau was observed. The plateau phase was inhibited when cells were treated with bromoenol lactone, a specific pharmacological inhibitor of iPLA2. These findings indicate that iPLA2 activity is required for ACE in keratinocytes. LPA also evoked Ca2+ release in keratinocytes but failed to induce sustained Ca2+ entry even when extracellular Ca2+ was elevated to 1·2 mmol L−1.ConclusionOur results demonstrate for the first time an important role for iPLA2 in regulating ACE in primary human keratinocytes.