DIFFERENTIATION ALTERS THE EXPRESSION OF THE 2 SPLICE VARIANTS OF THESEROTONIN-5-HT3 RECEPTOR-A MESSENGER-RNA IN NG108-15 CELLS

The serotonin 5-HT3-A receptor (5-HT(3)R-A) mRNA has been shown recent ly to be expressed as two forms (5-HT(3)R-A(L) and 5-HT(3)R-A(S)) vary ing by the presence or the absence of a sequence of 18 bases in the re gion corresponding to the second cytoplasmic domain of the receptor, a nd generated by alternative splicing at the level of the 3' acceptor s ite of exon 9. As the long form of the receptor exhibits a potential p hosphorylation site that is disrupted by the alternative splicing, the hypothesis of functional identity and stochastic expression of these two variants was questioned. In the present study, we used quantitativ e reverse transcriptase-polymerase chain reaction to examine the possi ble influence of culture conditions on the expression and the alternat ive splicing of 5-HT(3)R-A mRNA in NG108-15 clonal cells. Cell differe ntiation induced by dibutyryl cyclic AMP or theophyllin plus prostagla ndin E(1) in the presence of 10% serum reduced by threefold the expres sion of total 5-HT(3)R-A mRNA, and favored the short form of the messa ge as the ratio S/L (5-HT(3)R-A(S) mRNA/5-HT(3)R-A(L) mRNA) shifted fr om 2.23 to 7.33 after 9 days of treatment. Culture with 0.3% serum (in stead of 10%) lowered by 10-fold the level of expression of total 5-HT (3)R-A mRNA, but only slightly reduced the S/L ratio. However, this ra tio fell to 0.06 in the presence of 0.3% serum plus 10 ng/ml basic fib roblast growth factor. These results demonstrate that external factors can influence the differential expression of the two variants of the 5-HT(3)R-A in NG108-15 cells, Appropriate culture conditions for the a lmost exclusive expression of 5-HT(3)R-A(S) mRNA or 5-HT(3)R-A(L) mRNA in NG108-15 cells should allow the identification of possible differe nces in the respective functional properties of each of these two form s of the native 5-HT3 receptor.