Desensitization of human renal D1 dopamine receptors by G protein-coupled receptor kinase 4.
BACKGROUND
The D1 dopamine receptor, expressed in several nephron segments, participates in the regulation of water and electrolyte transport. Because the renal D1 receptor is desensitized in genetic hypertension, we sought to determine the mechanism(s) of the desensitization of D1 receptors endogenously expressed in renal proximal tubules.


METHODS
The mechanisms involved in the homologous desensitization of the D1 receptor in human renal proximal tubule cells were studied by measuring the production of cAMP in response to stimulation or inhibition of G protein-coupled receptor kinase (GRK) activity and expression. Protein expression was assessed by immunoblotting.


RESULTS
In human renal proximal tubule cells, the D1 agonist, fenoldopam, increased cAMP accumulation (73 +/- 2%). Fenoldopam pre-treatment decreased the responsiveness to subsequent fenoldopam stimulation (t(1/2) approximately equal to 20 min) with complete desensitization at 30 minutes. Recovery occurred gradually (t(1/2) approximately equal to 20 min) with full recovery at 60 minutes. Forskolin pretreatment minimally affected the fenoldopam effect, indicating a minor involvement of protein kinase A in the homologous desensitization process. Because GRKs are involved in the homologous desensitization process, we determined the consequences of inhibition of GRK expression and activity. Heparin, an inhibitor of GRK activity, decreased the expression of GRK2 and GRK4 and attenuated the desensitization of the D1 receptor (85 +/- 1%). Antisense oligonucleotides (GRK4> GRK2) blunted the D1 receptor desensitization. However, the first 20 minutes of homologous desensitization were not affected by either heparin or GRK antisense oligonucleotides.


CONCLUSIONS
These studies document the critical role of GRK4, relative to GRK2, in the homologous desensitization of D1 receptors in renal proximal tubule cells. However, the early phase of homologous desensitization is regulated by a non-GRK-mediated pathway.
Watanabe
Hidetsuna
H
Department of Pathology, University of Virginia Health Sciences Center, Charlottesville, VA 22908, USA.
Xu
Jing
J
Bengra
Chikh
C
Jose
Pedro A
PA
Felder
Robin A
RA
D000925:Anticoagulants / Q000494:pharmacology; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism; D018491:Dopamine Agonists / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D018818:Fenoldopam / Q000494:pharmacology; D054771:G-Protein-Coupled Receptor Kinase 4; D006493:Heparin / Q000494:pharmacology; D006801:Humans; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000378:metabolism*; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D017447:Receptors, Dopamine D1 / Q000378:metabolism*; D051552:beta-Adrenergic Receptor Kinases
D000925:Anticoagulants; D018491:Dopamine Agonists; D016376:Oligonucleotides, Antisense; D017447:Receptors, Dopamine D1; D005576:Colforsin; D006493:Heparin; D000242:Cyclic AMP; D017346:Protein Serine-Threonine Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D051552:beta-Adrenergic Receptor Kinases; D054771:G-Protein-Coupled Receptor Kinase 4; C517855:GRK4 protein, human; D018818:Fenoldopam