ENHANCEMENT OF THE MUSCARINIC SYNAPTOSOMAL PHOSPHOLIPID LABELING EFFECT BY THE IONOPHORE-A23187

  • 1 January 1981
    • journal article
    • research article
    • Vol. 37  (4) , 968-977
Abstract
Addition of either acetylcholine (ACh) or the ionophore A23187 [calcimycin] to synaptosomes resulted in a selective stimulation of 32Pi incorporation into phosphatidate (PhA) and phosphatidylinositol (PhI), while the labeling of phosphatidylinositol phosphate (PhIP) and phosphatidylinositol diphosphate (PhIP2) was reduced. The inclusion of both ACh and A23187 resulted in a synergistic increase in PhA and PhI labeling, and a synergistic decrease in the labeling of the polyphosphoinositides. Added Ca was not required, although inclusion of EGTA [ethyleneglycol-bis(.beta.-aminoethyl ether)N,N,N'',N''-tetraacetic acid] prevented the alterations in lipid labeling. The enhanced labeling of PhA and PhI by ACh or A23187 was not the result of either an increase in the radioactivity of the precursor [32P]ATP pool, or increased de novo synthesis of these lipids as judged from the incorporation of [3H]glycerol. [3H]glucose or [3H]myo-inositol. The synergistic alterations in PhA, PhI and polyphosphoinositide labeling were observed with ionophore only in the presence of selected muscarinic agonists, and with the inclusion of atropine or scopolamine the labeling reverted to a value which approximated that seen with the ionophore alone. Synergistic effects on phospholipid labeling with muscarinic agonists were also obtained with the calcium ionophore, ionomycin, but not with X537A [lasalocid] monensin or valinomycin. Neither the apparent number of muscarinic receptors present, nor their affinity for the ligand were altered by the presence of A23187. In prelabeling experiments, A23187 accelerated the loss of [32P]label from PhIP and PhIP2, and the rate of loss was further augmented by the addition of Ach. Neither agent produced comparable effects on the breakdown of prelabeled PhA or PhI. Phosphodiesteratic cleavage of the polyphosphoinosides might account for both the decrease in labeled PhIP and PhIP2 and increased labeling of PhA and PhI via the availability of resultant diglyceride. The results demonstrate that the turnover of polyphosphoinositides, in addition to that of PhA and PhI, is linked to the activation of muscarinic receptors.