Desensitization of tumour Leydig cells by lutropin: evidence for uncoupling of the lutropin receptor from the guanine nucleotide-binding protein

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Abstract
Purified rat Leydig tumor cells were pretreated with lutropin and the effect on the subsequent response to lutropin was determined. Maximal cAMP production was achieved with the same concentration of lutropin in control and lutropin-pretreated cells; however, the maximum stimulated level in pretreated cells was only 30% of controls. The sensitivity to lutropin was decreased in lutropin-pretreated cells [ED50 60 .+-. 5.7 ng/ml and 8 .+-. 1.8 ng/ml (mean .+-. SD, 3) for controls], as was the rate of maximal cAMP production (0.58, compared with 1.89 pmol/106 cells per min for controls). However, cholera toxin-stimulated cAMP production was not decreased by lutropin pretreatment, and a potentiation was seen at all time points studied (up to 6 h). Pre-incubation with lutropin caused a decrease in specific 125I-labeled human choriogonadotropin binding; however, this decrease was abolished if the cells were washed under acidic conditions (pH 3.0 for 2 min at 4.degree. C), indicating that occupation but not loss of the lutropin receptors had taken place. The effect of pretreating the cells with lutropin on adenylate cyclase activity in purified plasma membranes was also investigated. In plasma membranes from control cells both guanosine 5''-[.beta.,.gamma.-imido]triphosphate [p(NH)ppG] plus lutropin and NaF plus lutropin caused a 50 to 60-fold linear increase in cAMP production over 40 min compared with 15-fold with p(NH)ppG and 6-fold with lutropin alone. In plasma membranes isolated from lutropin-treated cells the NaF-plus-lutropin- and the p(NH)ppG-stimulated cAMP production rates were unchanged but no effect of lutropin could be demonstrated with or without added p(NH)ppG. In contrast the plasma membranes from dibutyryl cAMP-treated cells had similar cAMP production rates to control cells with all stimulants studied. The present evidence obtained from studies both with intact cells and with isolated plasma membranes indicates that the initial lutropin-induced desensitization of the rat Leydig tumor cell is due to a lesion in the hormone-receptor coupling to the guanine nucleotide regulatory protein. This process is apparently not mediated by cAMP.