Subtypes of α1-adrenoceptors in DDT 1 MF-2 and BC3H-1 clonal cell lines

Abstract

We examined which subtype(s) of α 1-adrenoceptors are expressed in the widely used DDT 1 MF-2 and BC3H-1 cell lines. Pretreatment with chloroethylclonidine (CEC) inactivated 76–85% of the specific [ 125I]BE 2254 binding sites in membrane preparations from both cell lines. Competition with subtype-selective competitive antagonists showed primarily the α 1B subtype in both cell lines. However, in BC3H-1 cells 5-methyl-urapidil showed complex behavior suggesting that about half of the binding sites had a lower affinity. Chloroethylclonidine pretreatment eliminated [ 3H]inositol phosphate responses to norepinephrine in both cell lines. Measurement of intracellular Ca 2+ with fura-2 in DDT 1 MF-2 cells showed that norepinephrine induced a complex response involving both transient and sustained components. Chloroethylclonidine pretreatment blocked both responses, while chelation of extracellular Ca 2+ left the transient response intact but eliminated the sustained component. These results support previous work that these cell lines contain α 1B-adrenoceptors linked to inositol phosphate formation and mobilization of intracellular Ca 2+. However, these results show that α 1B-adrenoceptors can be linked to Ca 2+ influx as well intracellular mobilization, and support the existence of pharmacologically distinct α 1B variants.