Stimulation of Phosphoinositide Hydrolysis by Serotonin in C6 Glioma Cells

Abstract

Abstract: 5‐Hydroxytryptamine (serotonin or 5‐HT) stimulated the incorporation of 32 P i into phosphatidylinositol (PI) but not into polyphosphoinositides in C6 glioma cells with an EC 50 of 1.2 ± 10 ‐7 M . The phosphoinositide response was blocked by the 5‐HT 2 antagonists ketanserin and spiperone but inhibited only partly by methysergide and mianserin. Atropine, prazosin, and yohimbine did not block the response, whereas fluphenazine and haloperidol did so partially but also inhibited basal incorporation by ˜30%. The 5‐HT 1A agonist 8‐hydroxy‐2 (di‐ n ‐propylamino)tetralin did not cause stimulation. Incubation with 5‐HT (1 μ M ) for 1 h increased the incorporation of [2‐ 3 H] myo ‐inositol into all phosphoinositides but not into inositol phosphates (IPs). Li + alone at 10 m M increased labeling in inositol bisphosphate (IP 2 ) and trisphosphate (IP 3 ), whereas labeling in IP and phosphoinositides remained unaltered. Addition of 5‐HT had no effect on this increase. Mn 2+ at 1 m M enhanced labeling in PI, PI‐4‐phosphate, lyso‐PI, glycerophosphoinositol, and IP, but the presence of 5‐HT again did not cause further stimulation. 5‐HT also stimulated the release of IPs in cells prelabeled with [2‐ 3 H] myo ‐inositol, incubated with LiCl (10 m M ) and inositol (10 m M ), and then exposed to 5‐HT (1 μ M ). Radioactivity in IP 2 and IP 3 was very low, was stimulated ˜50% as early as 30 s, and remained elevated for at least 20 min. Radioactivity in IP was at least 10 times as high as in IP 3 but was increased only from 3 min on with a peak at 20 min, when the elevation was ˜40 times that in IP 3 . The EC 50 value of 1.8 ± 10 ‐7 M was comparable with that obtained from 32 P i studies measuring labeled PI. Ketanserin and spiperone inhibited 5‐HT‐stimulated [2‐ 3 H]IP release with IC 50 values of 3.1 ± 10 ‐9 and 1.8 ± 10, ‐8 M , respectively. This study demonstrates that phosphoinositide hydrolysis is enhanced by 5‐HT in C6 glioma cells and that this phenomenon is linked to 5‐HT 2 ‐like binding sites.