Gonadotropin-releasing hormone stimulates mass changes in phosphoinositides and diacylglycerol accumulation in purified gonadotrope cell cultures.

ABSTRACT

GnRH stimulates rapid and specific hydrolysis of the phosphoinositides and their conversion to diacylglycerols (DAGs) in enriched gonadotrope cultures. In short term labeling studies, we observed stimulation of 32P incorporation into inositol phospholipids; no corresponding change was seen in other major phospholipids. The observation that this response continues in the presence of the Ca2+ channel antagonist D600 suggests that, unlike LH release, stimulation of phosphatidylinositol (PI) production is independent of mobilization of extracellular Ca2+. Agents that substitute for endogenous DAGs (phorbol myristate acetate) or mobilize Ca2+ directly (ionophore A23187) did not stimulate 32P incorporation into PI. Relative mass changes in PIs in response to GnRH were measured in a 32P/33P protocol. These showed a fall (40%) in the level of the inositol phospholipids within 45 sec, with a new steady state achieved at a lower level by 5 min. Phosphatidic acid formation was stimulated by GnRH within 2 min, with a 150% increase above control values at the highest dose of GnRH tested (10(-7) M). In cells prelabeled with [3H]arachidonic acid, GnRH caused a transient increase in DAG formation (twice the control value) within 1 min (with or without D600) which declined thereafter. This effect of GnRH was not seen in cells stimulated to release LH with ionophore A23187 or phorbol myristate acetate, suggesting that these may act distally to inositol phospholipid turnover. The data suggest that GnRH stimulates the hydrolysis of newly synthesized PIs, leading to the formation of DAGs. This pathway appears to be regulated independently of mobilization of extracellular Ca2+.