Hippocampal neurons maintain a large PtdIns(4)P pool that results in faster PtdIns(4,5)P(2) synthesis

PtdIns(4,5)P(2) is a signaling lipid central to the regulation of multiple cellular functions. It remains unknown how PtdIns(4,5)P(2) fulfills various functions in different cell types, such as regulating neuronal excitability, synaptic release, and astrocytic function. Here, we compared the dynamics of PtdIns(4,5)P(2) synthesis in hippocampal neurons and astrocytes with the kidney-derived tsA201 cell line. The experimental approach was to (1) measure the abundance and rate of PtdIns(4,5)P(2) synthesis and precursors using specific biosensors, (2) measure the levels of PtdIns(4,5)P(2) and its precursors using mass spectrometry, and (3) use a mathematical model to compare the metabolism of PtdIns(4,5)P(2) in cell types with different proportions of phosphoinositides. The rate of PtdIns(4,5)P(2) resynthesis in hippocampal neurons after depletion by cholinergic or glutamatergic stimulation was three times faster than for tsA201 cells. In tsA201 cells, resynthesis of PtdIns(4,5)P(2) was dependent on the enzyme PI4K. In contrast, in hippocampal neurons, the resynthesis rate of PtdIns(4,5)P(2) was insensitive to the inhibition of PI4K, indicating that it does not require de novo synthesis of the precursor PtdIns(4)P. Measurement of phosphoinositide abundance indicated a larger pool of PtdIns(4)P, suggesting that hippocampal neurons maintain sufficient precursor to restore PtdIns(4,5)P(2) levels. Quantitative modeling indicates that the measured differences in PtdIns(4)P pool size and higher activity of PI4K can account for the experimental findings and indicates that high PI4K activity prevents depletion of PtdIns(4)P. We further show that the resynthesis of PtdIns(4,5)P(2) is faster in neurons than astrocytes, providing context to the relevance of cell type–specific mechanisms to sustain PtdIns(4,5)P(2) levels.