Abstract

[³⁵S]Guanosine-5′-O-(3-thio)triphosphate ([³⁵S]GTPγS) binding to G proteins was measured byin vitro autoradiography in guinea pig and rat brain sections after activation by 5-hydroxytryptamine (5-HT) receptor agonists. 5-Carboxamidotryptamine stimulated binding strongly in hippocampus and lateral septum and weakly in substantia nigra. This effect was blocked in the substantia nigra by the 5-HT_1B/1Dreceptor antagonist GR-127,935 and in the former two regions by the 5-HT_1A antagonist NAN-190. 5-HT_1B/1D receptor agonists stimulated binding in substantia nigra and in areas containing 5-HT_1A receptors. In guinea pig substantia nigra, 5-(nonyloxy)-tryptamine maximally stimulated [³⁵S]GTPγS binding by 54%, with an EC₅₀ value of 62 nm; at 100 μm, this agonist increased binding by ∼200% in hippocampus (with a 2-fold weaker EC₅₀ value). The distribution of [³H]8-OH-DPAT binding sites was identical to that of the [³⁵S]GTPγS labeling stimulated by the 5-HT_1A agonist (R)-8-hydroxy-2-dipropylaminotetralin [(R)-8-OH-DPAT)]. (R)-8-OH-DPAT, (S)-8-OH-DPAT, and buspirone stimulated [³⁵S]GTPγS binding in hippocampus by 340%, 140%, and 78%, with EC₅₀ values of 71, 51, and 132 nm. Enhanced [³⁵S]GTPγS binding was not detected in the presence of 5-HT_1F, 5-HT₂, 5-HT₄, and 5-HT₇ receptor agonists. Because activation of μ-opioid, muscarinic M₂, histamine H₃, and cannabinoid receptors was also visualized successfully, these data suggest that only receptors coupled to pertussis toxin-sensitive G proteins can be seen by [³⁵S]GTPγS binding autoradiography. This study also shows that different 5-HT receptors coupled to these proteins can show a wide range of [³⁵S]GTPγS binding stimulation. Although the functional significance of these variations is unclear, this technique offers advantages over receptor autoradiography because it does not require high affinity radioligands and provides a measure of agonist efficacies in various brain regions.