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Selective Regulation of H₁ Histamine Receptor Signaling by G Protein-Coupled Receptor Kinase 2 in Uterine Smooth Muscle Cells

Reproductive Sciences Section (J.M.W., A.H.T., J.C.K.), Department of Cancer Studies and Molecular Medicine, University of Leicester, Leicester Royal Infirmary, Leicester LE2 7LX, United Kingdom; and Department of Cell Physiology and Pharmacology (J.M.W., H.S., R.A.J.C.), University of Leicester, Leicester LE1 9HN, United Kingdom

Address all correspondence and requests for reprints to: Jonathon M. Willets, Reproductive Sciences Section, Department of Cancer Studies and Molecular Medicine, University of Leicester, Clinical Sciences Building, Leicester Royal Infirmary, Leicester LE2 7LX, United Kingdom. E-mail: jmw23{at}le.ac.uk.

Histamine stimulates uterine contraction; however, little is known regarding the mechanism or regulation of uterine histamine receptor signaling. Here we investigated the regulation of G_q/11-coupled histamine receptor signaling in human myometrial smooth muscle cells using the inositol 1,4,5-trisphosphate biosensor pleckstrin homology domain of phospholipase-1 tagged to enhanced green fluorescent protein and the Ca²⁺-sensitive dye Fluo-4. Histamine addition caused concentration-dependent increases in inositol 1,4,5-trisphosphate and [Ca²⁺]_i in the ULTR human uterine smooth muscle cell line and primary human myometrial cells. These effects were completely inhibited by the H₁ histamine receptor antagonist, diphenhydramine, and were unaffected by the H₂ histamine receptor antagonist, cimetidine. ULTR and primary myometrial cells were transfected with either dominant-negative G protein-coupled receptor kinases (GRKs) or small interfering RNAs targeting specific GRKs to assess the roles of this protein kinase family in H₁ histamine receptor desensitization. Dominant-negative GRK2, but not GRK5 or GRK6, prevented H₁ histamine receptor desensitization. Similarly, transfection with short interfering RNAs (that each caused >70% depletion of the targeted GRK) for GRK2, but not GRK3 or GRK6, also prevented H₁ histamine receptor desensitization. Our data suggest that histamine stimulates phospholipase C-signaling in myometrial smooth muscle cells through H₁ histamine receptors and that GRK2 recruitment is a key mechanism in the regulation of H₁ histamine receptor signaling in human uterine smooth muscle. These data provide insights into the in situ regulation of this receptor subtype and may inform pathophysiological functioning in preterm labor and other conditions involving uterine smooth muscle dysregulation.

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