Phorbol ester and diacylglycerol mimic growth factors in raising cytoplasmic pH

Abstract

There is now good evidence that cytoplasmic pH (pH_i) may have an important role in the metabolic activation of quiescent cells^1,2. In particular, growth stimulation of mammalian fibroblasts leads to a rapid increase in pH_i (refs 3–6), due to activation of a Na⁺/H⁺ exchanger in the plasma membrane⁴, and this alkalinization is necessary for the initiation of DNA synthesis⁷. However, the mechanism by which mitogens activate the Na⁺/H⁺ exchanger to raise pH_i is not known, although an increase in cytoplasmic free Ca²⁺ ([Ca²⁺]_i) has been postulated as the primary trigger^8–10. We now present data suggesting that the Na⁺/H⁺ exchanger is set in motion through protein kinase C, a phospholipid- and Ca²⁺-dependent enzyme normally activated by diacylglycerol produced from inositol phospholipids in response to external stimuli^11,12. Using newly developed pH microelectrodes and fluorimetric techniques, we show that a tumour promoting phorbol ester and synthetic diacylglycerol, both potent activators of kinase C (refs 12–15), mimic the action of mitogens in rapidly elevating pH_i in different cell types. Furthermore, we demonstrate that, contrary to previous views, an early rise in [Ca²⁺]_i is not essential for the activation of Na+/H+ exchange and the resultant increase in pH_i. Finally, we suggest that an alkaline pH_i shift, mediated by Na+/H+ exchange, may be a common signal in the action of those hormones which elicit the breakdown of inositol phospholipids.