Homologous Down-Regulation of Gonadotropin-Releasing Hormone Receptors and Desensitization of Gonadotropes: Lack of Dependence on Protein Kinase C

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Abstract
The demonstration that GnRH provokes the accumulation of diacylglycerol and the redistribution of protein kinase C to the membrane fraction in gonadotropes suggests a role for this enzyme as a mediator of GnRH action. In the present work we have investigated the possibility that protein kinase C might mediate GnRH-stimulated receptor down-regulation and densensitization. Pretreatment of pituitary cells for 6 h with GnRH (10-11-10-6 M) caused a biphasic change in GnRH receptor number [the maximum binding (Bmax) for 125I-buserelin binding was increased by 10-10 M GnRH and reduced by 10-7 and 10-6 M GnRH] and caused desensitization (pretreatment with 10-9-10-6 M GnRH reduced the proportion of cellular LH released in a subsequent challenge with GnRH). Pretreatment for 6 h with 0.2-200 nM phorbol myristate acetate (a protein kinase C-activating phorbol ester) did not cause desensitization, but at 200 nM, did reduce GnRH receptor number. As a further test of the requirement for protein kinase C for GnRH action, cells were depleted of all measurable protein kinase C (and rendered unresponsive to protein kinase C activators) by prior treatment with a high dose of phorbol myristate acetate (500 nM for 6 h followed by 12 h in plating medium). Depletion of protein kinase C did not alter the ability of GnRH to desensitize gonadotropes or down-regulate its own receptors. The demonstration that the effects of GnRH on receptor number and gonadotrope responsiveness are neither blocked by depletion of protein kinase C nor entirely mimicked by activation of protein kinase C suggests that these effects of the releasing hormone are not solely mediated by this enzyme.