Homologous desensitization with gonadotropin-releasing hormone (GnRH) also diminishes gonadotrope responsiveness to maitotoxin: a role for the GnRH receptor-regulated calcium ion channel in mediation of cellular desensitization.

Maitotoxin (MTX) stimulates gonadotropin release from pituitary cell cultures. The time course and efficacy of LH release in response to GnRH and to MTX are similar; both secretagogues require extracellular Ca2+ and are inhibited by the selective Ca2+ ion channel antagonist methoxyverapamil (D600). LH release in response to either GnRH or MTX is not measurably inhibited by two other chemical classes of Ca2+ ion channel inhibitors represented by nifedipine and by diltiazem. The two secretagogues are nonadditive in their action on LH release when presented at high doses and prior studies indicate that MTX has no endogenous ionophoretic activity. These observations indicate that MTX likely stimulates LH release due to activation of the GnRH receptor associated Ca2+-ion channel in the gonadotrope. We have therefore assessed the functional state of this channel during the development of homologous desensitization of the gonadotrope to GnRH by measuring the ability of MTX to stimulate LH release. Cells were desensitized with GnRH in the presence of 3 mM EGTA. Under these conditions, the cells become refractory to GnRH in the absence of gonadotropin release since the latter process, but not the former, requires extracellular Ca2+. Accordingly, this approach allows assessment of the degree of desensitization in the absence of the influence of gonadotropin depletion. Such desensitized cells are less responsive to GnRH. Desensitized pituitary cells also respond with diminished efficacy and potency to MTX three or more hours after GnRH treatment but not at an earlier time (1 h) when GnRH receptors are diminished. These data are consistent with a model in which homologous desensitization is viewed as developing in two phases.(ABSTRACT TRUNCATED AT 250 WORDS)

Stimulation of luteinizing hormone (LH) release and phospholipid breakdown by guanosine triphosphate in permeabilized pituitary gonadotropes: antagonist action suggests association of a G protein and gonadotropin-releasing hormone receptor.

The stimulation of gonadotropin release from pituitary cell cultures by GnRH has been linked to inositol phospholipid breakdown to diacylglycerols and subsequent activation of protein kinase C as well as Ca2+ mobilization. In order to examine the means of receptor coupling to a phospholipase C-type reaction, we evaluated the role of guanine nucleotides in inositol phospholipid breakdown. In these studies ATP (50 microM) was used for cell permeabilization to allow guanine nucleotides access to the intracellular compartment. Under these conditions GTP and the GTP analog, guanylylimidodiphosphate (GMP-PNP), stimulated a time- and dose-dependent increase in LH release and inositol phosphate accumulation. These actions of GTP and GMP-PNP were not observed unless ATP was included in the treatment media. Other closely related nucleotides and nucleosides alone, or in the presence of ATP, did not elevate LH release above basal levels. We also evaluated the actions of pertussis toxin and cholera toxin on mediating the effect of GTP, GMP-PNP, and GnRH on LH release and inositol phosphate accumulation. After treatment with these agents, no changes were observed in the ability of GnRH, GTP, or GMP-PNP to stimulate either LH release or inositol phosphate accumulation. The additional observation that GnRH-, GTP-, or GMP-PNP-stimulated LH release and inositol phosphate accumulation were blocked by a potent GnRH antagonist suggests that a G protein is functionally associated with the GnRH receptor recognition site.