SYNTHESIS AND EVALUATION OF [I-125] (S)-IODOZACOPRIDE, A HIGH-AFFINITY RADIOLIGAND FOR 5HT(3) RECEPTORS

We have developed a high specific activity radioiodinated ligand for t he biochemical evaluation and autoradiographic localization of 5HT3 re ceptors in the brain. [I-125]-(S)-iodozacopride was synthesized by rad ioiodination of deschloro-(S)-zacopride using chloramine-T, and the pr oduct was purified by HPLC. The equilibrium kinetics and pharmacology of the binding of this radioligand were studied in homogenates of rat cerebral cortex, while the distribution of binding was examined by qua ntitative autoradiography. [I-125]-(S)-iodozacopride bound to a single , saturable, specific binding site (K(d) = 192+/-9 pM, B(max) = 1.2+/- 0.2 fmol/mg protein). The binding had the pharmacological properties o f a 5HT3 receptor, being potently inhibited by a variety of 5HT3 agoni sts and antagonists including (S)-zacopride (K(i) = 0.032 nM), Quipazi ne (K(i) = 0.45 nM), LY278584 (K(i) = 0.5 nM), (1-m-chlorophenyl)-bigu anide (K(i) = 0.6 nM) and ICS 205-930 (K(i) = 1.0 nM). Autoradiographi c studies were undertaken by incubating sections with 400 pM [I-125]-( S)-iodozacopride and exposing them to film for 3-7 days to obtain suit able autoradiograms. Specific binding of [I-125]-(S)-iodozacopride was found at various amounts in a variety of brain regions. The highest l evels of binding were found in the brainstem, principally the nucleus of the solitary tract with somewhat lower levels in the area postrema, substantia gelatinosa of the trigeminal nucleus and dorsal motor nucl eus of the vagus. In the rat forebrain, moderate levels of specific bi nding were found in the glomerular layer of the olfactory bulb, anteri or olfactory nucleus and various subnuclei of the amygdala. Lower leve ls of binding were seen in the superficial laminae of the parietal cer ebral cortex and diffusely distributed throughout the hippocampal form ation. In conclusion, [I-125]-(S)-iodozacopride binds to a receptor si te with the pharmacological properties and distribution that is consis tent with the 5HT3 receptor. [I-125]-(S)-iodozacopride represents a si gnificant improvement in autoradiographic studies of the 5HT3 receptor by reducing the required exposure time for producing autoradiograms f rom the 3-6 months required for [H-3]-labeled ligands to 3-7 days.