Interaction of parathyroid hormone-related peptide-responsive dual signal transduction systems in osteoblastic osteosarcoma cells: Role in PTHrP-induced homologous desensitization

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Abstract
In Osteoblastic Umr-106 Cells, 10−7 M human (h) PTH-related peptide (PTHrP)-(1–34) significantly induced the formation of total inositol phosphates to the same degree as 10−7 M hPTH-(1–34), confirming that in addition to cAMP-dependent protein kinase (PKA), PTHrP possesses another signal transduction system, calcium/protein kinase C (Ca/PKC). Experiments were therefore performed to characterize the cross talk of these dual-signal transduction systems and its participation in the PTHrP-induced homologous desensitization of cAMP and cytosolic calcium (Cai) response in osteoblasts. Preincubation with 10−7 M hPTHrP-(1–34) caused homologous desensitization, resulting in a remarkable decrease in cAMP accumulation in response to further exposure to PTHrP. This effect was significant after 2 h pretreatment and reached a maximum at 6 h. Pretreatment with the PKC-activating phorbol ester phorbol 12-myristate-13-acetate (PMA, 10−6 M) for 30 minutes and 6 h caused a significant increase and decrease in cAMP responsiveness to PTHrP, respectively. Pretreatment with calcium ionophores (A23187 or ionomycin, 10−6 M), not for 30 minutes but for 6 h, caused a significant decrease in cAMP responsiveness to PTHrP. H-7 (an inhibitor of PKC, 50 μM) significantly blocked not only PMA- but also PTHrP-induced desensitization of the cAMP response. PTHrP caused the complete homologous desensitization of an increase in Cai within 30 minutes. Pretreatment with dibutyryl-cAMP (10−4 M) for 30 minutes caused significant inhibition of the PTHrP-induced increase in Cai, and pretreatment with Sp-cAMPS (10−4 M), a direct activator of PKA, for 30 minutes completely blocked the PTHrP-induced increase in Cai. Rp-cAMPS (10−4 M), an antagonist in the activation of PKA, slightly but significantly antagonized the PTHrP-induced homologous desensitization of the Cai response. The present study demonstrated the existence of cross-talk in PTHrP-responsive dual signal transduction systems and its participation in PTHrP-induced homologous desensitization.

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