Response of SCC-12F, a human squamous cell carcinoma cell line, to complement attack

Serotonin 5-HT1A receptors belong to the superfamily of G-protein-coupled receptors. Receptor activation of G-proteins can be determined by agonist-stimulated 35S]GTPgammaS binding in the presence of excess GDP, and in vitro autoradiographic adaptation of this technique allows visualization of receptor-activated G-proteins in tissue sections. The present study was performed to examine 5-HT1A receptor activation of G-proteins using 8-OH-DPAT-stimulated 35S]GTPgammaS binding in membranes and brain sections. In hippocampal membranes, 8-OH-DPAT stimulated 35S]GTPgammaS binding by twofold, with an ED50 value of 25 nM. 5-HT1 antagonists, but not 5-HT2 antagonists, increased the ED50 of 8-OH-DPAT in a manner consistent with competitive antagonists. Scatchard analysis of 35S]GTPgammaS binding showed that 8-OH-DPAT induced the formation of high affinity 35S]GTPgammaS binding sites with a KD for GTPgammaS of 3.2 nM. 35S]GTPgammaS autoradiography, performed in brain sections with the 5-HT1A agonist 8-OH-DPAT, revealed high levels of 5-HT1A-stimulated 35S]GTPgammaS binding in the hippocampus, lateral septum, prelimbic cortex, entorhinal cortex, and dorsal raphe nucleus. 5-HT1A-stimulated 35S]GTPgammaS binding in sections was blocked by the addition of the 5-HT1 antagonist methiothepin. These results show that the use of agonist-stimulated 35S]GTPgammaS autoradiography for the 5-HT1A receptor system should provide new information regarding signal transduction in specific brain regions.