Gonadotropin-releasing hormone receptor microaggregation. Rate monitored by fluorescence resonance energy transfer.

ABSTRACT

Gonadotropin-releasing hormone (GnRH) regulates pituitary gonadotropin release and is a therapeutic target for human and animal reproductive diseases. In the present study we have utilized the technique of fluorescence resonance energy transfer to monitor the rate of GnRH receptor-receptor interactions. This technique relies on the observation that the degree of physical intimacy of molecules can be assessed by the tendency of proximal fluorophores to exchange energy. Our data indicate that GnRH agonist, but not antagonist, occupancy of the GnRH receptor promotes physical intimacy (microaggregation) between receptors. The time course indicates that this occurs promptly (<1 min) after occupancy and persists for at least 80 min and within the physiologically relevant range of the releasing hormone. The process measured is not inhibited by 0.1 mm vinblastin, 2 microm cytochalasin D, or 3 mm EGTA, an observation that distinguishes it from macroaggregation (patching, capping, and internalization). These observations, along with reports from other laboratories, are consonant with a growing body of evidence that indicates that microaggregation is an early event following agonist occupancy of the receptor and part of the mechanism by which effector regulation occurs.