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Desensitization of vasopressin sensitive adenylate cyclase by vasopressin and phorbol esters
1Department of Molecular Pharmacology L521, Smith Kline & French Laboratories, King of Prussia, PA 19406-0939.
Abstract:
Desensitization of vasopressin V2 receptor-mediated adenylate cyclase was studied in canine kidney cell line, MDCK cells. Overnight treatment of MDCK cells with arginine vasopressin (AVP) resulted in a loss of vasopressin receptors and an inhibition of cAMP accumulation in response to AVP. Both the loss of receptor and reduction in cAMP accumulation were time- and AVP concentration-dependent. Desensitization was selective for AVP because cAMP formation in response to isoproterenol, prostaglandin E1 (PGE1) and forskolin was not affected by AVP pre-treatment. Pre-treatment of MDCK cells with phorbol dibutyrate (PDBu) also caused a dose-dependent inhibition of AVP mediated cAMP accumulation, but not of isoproterenol-, PGE1- and forskolin-induced cAMP accumulation. PDBu pre-treatment did not cause loss of vasopressin receptors. Instead, the affinity for vasopressin was changed by PDBu treatment. Pre-treatment of the cells with pertussis toxin (PT) had no effect on the desensitization and downregulation of vasopressin (V2) receptors, suggesting that the desensitization may not be mediated by pertussis toxin sensitive G-protein. Our data suggest that pre-treatment of MDCK cells with AVP or PDBu caused desensitization of AVP-mediated cAMP accumulation and that downregulation of V2 receptors required agonist occupancy of the receptors, whereas the affinity of the receptors was changed by phorbol ester treatment.
Insights
Arginine vasopressin (AVP) desensitizes kidney cells by reducing vasopressin receptors and cAMP levels. Phorbol ester also inhibits AVP signaling, affecting receptor affinity but not receptor number.
Area of Science:
- Cell Biology
- Pharmacology
- Endocrinology
Background:
- Vasopressin V2 receptors regulate water balance via cAMP signaling.
- Understanding receptor desensitization is crucial for managing conditions involving vasopressin dysregulation.
Purpose of the Study:
- To investigate the mechanisms of vasopressin V2 receptor desensitization in MDCK cells.
- To differentiate between receptor downregulation and alterations in signaling pathways.
Main Methods:
- Treatment of canine kidney (MDCK) cells with arginine vasopressin (AVP) or phorbol dibutyrate (PDBu).
- Measurement of vasopressin receptor levels and cAMP accumulation.
- Assessment of signaling pathways using isoproterenol, prostaglandin E1, and forskolin.
Main Results:
- AVP treatment led to time- and concentration-dependent loss of vasopressin receptors and reduced cAMP.
- Desensitization was selective for AVP, sparing other cAMP-inducing agents.
- Phorbol dibutyrate inhibited AVP-mediated cAMP accumulation by altering receptor affinity, not by reducing receptor number.
- Pertussis toxin did not affect AVP-induced desensitization or receptor downregulation.
Conclusions:
- AVP-induced desensitization involves both V2 receptor downregulation (requiring agonist occupancy) and potential post-receptor events.
- Phorbol ester treatment desensitizes the V2 receptor pathway through a mechanism distinct from downregulation, possibly by modifying receptor affinity.
- The signaling pathway for AVP desensitization may not involve pertussis toxin-sensitive G-proteins.