HIGH-AFFINITY FSH RECEPTOR-BINDING IS A SLOWLY DEVELOPING NON-REVERSIBLE PROCESS THAT APPEARS NOT TO BE IMPORTANT FOR RAPID RECEPTOR ACTIVATION

The kinetics of FSH-induced stimulation of adenylyl cyclase and of FSH receptor binding have been studied, in order to test the hypothesis t hat FSH-induced receptor activation and high affinity FSH receptor bin ding are mediated by different hormone/receptor complexes. Within 2 mi n after addition of FSH to rat Sertoli cells, cAMP production was stim ulated at a constant production rate for only 5 min. After 10 min aden ylyl cyclase activity returned to basal levels. The steady state level of cAMP production was proportional to the FSH concentration. Since s timulation of adenylyl cyclase activity is correlated with receptor oc cupancy, different steady state levels of FSH/receptor complex must ha ve been formed within 2 min after hormone addition. The rapid establis hment of steady state receptor activation is in sharp contrast with th e slow development of high affinity receptor binding, which requires s everal hours before an apparent steady state was reached. Steady state receptor activation occurs when FSH receptor binding is between 0 and 5% of the maximal binding. Moreover, in the presence of steady state receptor activation, FSH receptor binding increases linearly in time. Since receptor binding of FSH was only partially (approximate to 40%) reversible, the slow development of the high affinity binding complex suggests a transformation process of the initial FSH/receptar complex that is involved in transmembrane signalling, into a more stable bindi ng complex. Consistent with the NaCl dependent affinity of the recepto r for FSH (van Loenen et al., 1994) 10-fold higher FSH concentrations were required to induce a similar rate of cAMP production when Krebs R inger buffer was used instead of Krebs Ringer buffer without NaCl but with 200 mM sucrose. The kinetics of the cAMP response were not affect ed by NaCl. On the basis of our experimental findings and those report ed by other investigators, we propose a model with at least two distin ct FSH/receptor complexes: H + R double left right arrow HR' (relative ly low affinity and coupled with signal transduction and internalizati on) double left right arrow HR'' (higher affinity and non-reversible). With this model the effects of NaCl on initial hormone/receptor compl ex formation and on the development of high affinity binding might be explained. The properties of the high affinity complex appear to be mi sleading for a proper understanding of the signal transduction propert ies of the receptor. The individual kinetic constants of the relativel y low affinity complex, rather than the value of the apparent equilibr ium constant of the high affinity complex, might be important.