Genistein inhibits Na+/Ca2+ exchange activity in primary rat cortical neuron culture

We have examined the possible regulatory effect of tyrosine kinase activity on Ca2+ transport observed in the cultured rat cortical neurons. Na+/Ca2+ exchange was studied using cells cultured for various time periods. A nearly two fold increase in Ca2+ uptake was seen when comparing 3 day and 9 day cultures. Western blot analysis also showed a two fold increase in Na+/Ca2+ exchanger (NCX1) protein levels as cells matured in culture. To study the effect of genistein (a specific tyrosine kinase inhibitor) cells were incubated with 100 microM genistein (in 1% DMSO) for 1 hour before the assay of Na+/Ca2+ exchange activity. There was a significant decrease of Ca2+ uptake in genistein treated neurons (control: 4.596+/-0.205 nmol/mg protein/15 min, n=12; genistein: 1.420+/-0.131 nmol/mg protein/15 min, n=12, mean+/-S.E. P<0.001). Daidzein, an inactive analog of genistein and phorbol myristate acetate (PMA), a PKC activator were without effect. The results suggest that as cells mature in culture, Na+/Ca2+ exchange capacity increases, as a result of greater protein expression. Exposure to genistein inhibited Ca2+ uptake suggesting that the exchanger may be modulated by tyrosine phosphorylation.