[3H]LY334370, a novel radioligand for the 5-HT1F receptor. I. In vitro characterization of binding properties

[3H]LY334370 was developed as a radioligand to study the characteristics of this compound’s interaction with the 5-HT1F receptor. Monovalent or divalent cations did not enhance the binding of [3H]LY334370 to the cloned human 5-HT1F receptor. In the presence of MgCl2, the time to reach equilibrium was approximately 2 h, while in its absence equilibrium was reached in less than 1 h. [3H]LY334370 had high affinity for the cloned human 5-HT1F receptor (Kd=0.446 nM) and the 5-HT1F receptor in rat brain (Kd=0.388 nM). The expression density of 5-HT1F receptors, as determined by binding to homogenates of cortical regions from rat, was low (Bmax=79.1 fmol/mg protein). There was a statistically significant correlation between the apparent pKi for inhibition of [3H]LY334370 binding and the pEC50 for stimulation of [35S]GTPγS binding to homogenates of cells expressing the cloned human 5-HT1F receptor. In addition, there was a statistically significant correlation between the apparent pKi for inhibition of [3H]LY334370 binding to the cloned human 5-HT1F receptor and the pID50 for inhibition of trigeminal nerve stimulated dural plasma protein extravasation in the guinea pig. The conclusion from these studies is that [3H]LY334370 is a high affinity radioligand which can be used for the study of the 5-HT1F receptor in rat brain or in cells transformed with the human 5-HT1F receptor.