Involvement of sulfhydryl groups in cerebral cortical [3H]clonidine binding in developing rats

The involvement of essential sulfhydryl groups in alpha 2-adrenoceptor function was investigated in the cerebral cortex of 7 and 70 day old rats. N-Ethylmaleimide (NEM) inhibited specific [3H]clonidine binding to cerebral cortical membranes in a concentration-dependent and biphasic manner in both infant and adult rats. The inhibitory effect of NEM was attenuated by simultaneous addition of dithiothreitol 30 microM and 1 mM, though dithiothreitol up to 1 mM did not affect the binding. p-Chloromercuriphenylsulfonic acid also caused a significant reduction in [3H]clonidine binding at both stages. Scatchard analysis of [3H]clonidine binding showed that NEM 10 and 100 microM caused a significant decrease in Bmax of high affinity binding sites without changing KD. Neither Bmax nor KD values were changed by NEM 10 and 100 microM in low affinity sites. The treatment with NEM 10 and 100 microM significantly reduced the increase of binding induced by Mn2+ 10 and 100 microM which was observed on day 70 but not on day 7. It is suggested that two distinct categories of essential sulfhydryl groups are involved in cerebral cortical alpha 2-adrenoceptors and are functionally mature by day 7. Furthermore, sulfhydryl groups are involved in the Mn2+-induced increase of alpha 2-receptors in adult rats.