The identification of a novel inositol lipid, phosphatidylinositol trisphosphate (PIP3), in rat cerebrum using in vivo techniques

Rats received intraventricular injections of 20 uCi of 3H]-myo-inositol, and were sacrificed 24 hrs later by high-power head-focused microwave fixation. Two inositol lipid extraction methods were compared: The Hauser and Eichberg method yielded higher recovery of inositol lipids, but a lower inositol phosphate content. The Schacht method yielded reduced radiolabel in the lipid fractions, but increased water soluble phosphates. Both methods extracted a novel inositol lipid (PIP3) which contained inositol tetrakisphosphate (IP4) as its polar head group. This was determined by alkaline hydrolysis and analyzed by high performance liquid chromatography with authentic IP4 standard. Furthermore, preliminary studies of the fatty acid composition indicated a similarity with other inositol lipids. The radiolabel ratio of PIP2:PIP3 was 5:1. In summary, we have isolated a novel inositol phospholipid in rat brain, PIP3, the parent compound for inositol tetrakisphosphate (IP4).