Vascular Abnormalities in Mice Deficient for the G Protein-Coupled Receptor GPR4 That Functions as a pH Sensor

Abstract

GPR4 is a G protein-coupled receptor expressed in the vasculature, lung, kidney, and other tissues. In vitro ectopic overexpression studies implicated GPR4 in sensing extracellular pH changes leading to cyclic AMP (cAMP) production. To investigate its biological roles in vivo, we generated GPR4-deficient mice by homologous recombination. Whereas GPR4-null adult mice appeared phenotypically normal, neonates showed a higher frequency of perinatal mortality. The average litter size from GPR4^−/− intercrosses was ∼30% smaller than that from GPR4^+/+ intercrosses on N3 and N5 C57BL/6 genetic backgrounds. A fraction of knockout embryos and neonates had spontaneous hemorrhages, dilated and tortuous subcutaneous blood vessels, and defective vascular smooth muscle cell coverage. Mesangial cells in kidney glomeruli were also significantly reduced in GPR4-null neonates. Some neonates exhibited respiratory distress with airway lining cell metaplasia. To examine whether GPR4 is functionally involved in vascular pH sensing, an ex vivo aortic ring assay was used under defined pH conditions. Compared to wild-type aortas, microvessel outgrowth from GPR4-null aortas was less inhibited by acidic extracellular pH. Treatment with an analog of cAMP, a downstream effector of GPR4, abolished microvessel outgrowth bypassing the GPR4-knockout phenotype. These results suggest that GPR4 deficiency leads to partially penetrant vascular abnormalities during development and that this receptor functions in blood vessel pH sensing.

SUPPLEMENTAL MATERIAL

We thank Shirley Quan, LaKeisha Perkins, Donghui Chen, James Johnson, and Mireille Riedinger for expert technical assistance; Alfonso Luque, Sanaz Memarzadeh, Shuling Guo, and Gregory Ferl for helpful discussion; Nathan Lee and Jen Hofmann for help with the confocal microscope; Antoni Ribas and Roger Lo for critically reading the manuscript; the Division of Laboratory Animal Medicine at UCLA for mouse histological evaluation and blood cell counting; and Barbara Anderson for help with manuscript preparation.

L.V.Y. is an Associate and O.N.W. is an Investigator of the Howard Hughes Medical Institute. L.V.Y. is partially supported by NCI/NIH award no. T32 CA009056.