Striatal [3H]GTP binding in developing rats: involvement of sulfhydryl residues, Ca2+ and Mg2+

The influence of sulfhydryl reagents and cations on specific [3H]GTP binding to striatal membranes was investigated in developing rats. Two components of non-cooperative [3H]GTP binding sites were observed in 15, 30, 70 and 360 day old rats but only a single component in 1 and 7 day old ones. The KD for low affinity binding increased with age. Bmax values for both high and low affinity binding increased with age and reached a peak at 30 days, followed by a decrease at 70 and 360 days. At 7 and 70 days, NaCl 1-100 mM did not affect [3H]GTP binding but CaCl2 and MgCl2 significantly inhibited the binding over a concentration range of 1-100 mM. TLC analysis of [3H]GTP and the metabolites in the binding medium and membranes showed that [3H]GTP in both membranes and in the medium was decreased by addition of 1 mM CaCl2 and 1 mM MgCl2 into the binding medium. On days 7 and 70, p-chloromercuriphenyl sulfonate strongly inhibited [3H]GTP binding, and dithiothreitol significantly increased binding but dopamine, apomorphine, spiperone and alpha-flupenthixol did not increase binding up to 0.1 mM. It is suggested that sulfhydryl residues, Ca2+ and Mg2+ are involved in the regulation of guanine nucleotide binding and that the regulatory mechanism becomes functional at 7 days. Ca2+ and Mg2+ seem to act by stimulating degradation of [3H]GTP. In addition, the density of GTP binding sites reaches a peak at around 30 days and the affinity decreases with age.