Rapidly-labelled, acidic phospholipids of the goldfish brain.

Homogenates and particulate fractions of goldfish brain incorporated radioactivity from y-[B*P]ATP selectively into acidic phospholipids during brief periods of incubation. Phosphatidate and lysophosphatidate became strongly labelled and activity was also found in phosphatidyl inositol phosphate and in phosphatidyl inositol diphosphate. When tetraphenylborate (a K+-complexing agent) was added, a selective stimulation of incorporation of 3*P into phosphatidate occurred. The addition of perchlorate (also known to bind K+) did not produce a similar stimulation, nor did the addition of K+ block the stimulation by tetraphenylborate. The stimulation of the labelling of phospholipids by tetraphenylborate appeared to be the result of multiple actions. Besides the evidence that it acted by stimulating the phosphoinositide phosphodiesterase of brain, data were obtained suggesting that it stimulated diglyceride kinase and blocked endogenous destruction of ATP as well. The stimulation by tetraphenylborate was blocked by addition of atropine but not of arecoline. THE PHOSPHOINOSITIDES, although ubiquitous in animal tissues, frequently exist in trace amounts demonstrable only by tracer methods. The greater content of phosphoinositides in excitable tissues (FOLCH-PI, 1949; DAWSON and DITTMER, 1961 ; BROCKERHOFF and BALLOU, 1962) and the evidence from labelling experiments of rapid turnover (DAWSON, 1954; HOKIN and HOKIN, 1958; ANSELL and SPANNER, 1959; LEBARON, KISTLER and HAUSER, 1960; HOLZL and WAGNER, 1964; KFOURY and KERR, 1964; SEIFFERT and AGRANOFF, 1965) have suggested an important physiological role for these compounds in excitable tissues. Alterations of the labelling of phosphatidyl inositol (PhI) and phosphatidic acid (PhA) have been described in a variety of neural and secretory tissues in response to the appropriate neurohumor or secretagogue (cf. reviews by: HAWTHORNE, 1960; HOKIN and HOKIN, 1960; CumBERT, 1967). In sympathetic ganglia increased labelling has been correlated with synaptic transmission (LARRABEE and LEICHT, 1965; LARRABEE, 1968). Recently DURELL, GARLAND and FRIEDEL (1969) have suggested that such alterations are the result of activation of phosphatidyl inositol phosphodiesterase releasing diglyceride and inositol phosphate, but the detailed mechanism of the effect of acetylcholine remains speculative. The present experiments were undertaken to clarify the nature of these alterations in phospholipid labelling. The goldfish was selected because of the current interest in this laboratory in goldfish behaviour and its metabolic consequences. Various aspects of the labelling patterns of phospholipids in the goldflsh brain, the effects of acetylcholine in goldfish brain, and the stimulation by tetraphenylborate of phospholipid labelling comprise the present report. 1 Supported by grant NB3101 of the United States Public Health Service. A preliminary communication of some of these results has been reported (HOLLANDER, HALLENBECK and AGRANOFF, 1969). * Special Fellow of the NINDS, 2F l lNB 1900. Present address: Department of Neurology, University of Rochester, Rochester General Hospital, Rochester, New York 14621. Abbreoiutions used: PhA, phosphatidic acid; PhI, phosphatidyl inositol; PhIP, phosphatidyl inositol phosphate; PhIP,, phosphatidyl inositol diphosphate; TPB, sodium tetraphenylborate.