Histamine Induces ATP Release from Human Subcutaneous Fibroblasts,via Pannexin-1 Hemichannels,Leading to Ca2+ Mobilization and Cell Proliferation

Changes in the regulation of connective tissue ATP-mediated mechano-transduction and remodeling may be an important link to the pathogenesis of chronic pain. It has been demonstrated that mast cell-derived histamine plays an important role in painful fibrotic diseases. Here we analyzed the involvement of ATP in the response of human subcutaneous fibroblasts to histamine. Acute histamine application caused a rise in intracellular Ca²⁺ (Ca²⁺]_i) and ATP release from human subcutaneous fibroblasts via H₁ receptor activation. Histamine-induced Ca²⁺]_i rise was partially attenuated by apyrase, an enzyme that inactivates extracellular ATP, and by blocking P2 purinoceptors with pyridoxal phosphate-6-azo(benzene-2,4-disulfonic acid) tetrasodium salt and reactive blue 2. Ca²⁺]_i accumulation caused by histamine was also reduced upon blocking pannexin-1 hemichannels with ¹⁰Panx, probenecid, or carbenoxolone but not when connexin hemichannels were inhibited with mefloquine or 2-octanol. Brefeldin A, an inhibitor of vesicular exocytosis, also did not block histamine-induced Ca²⁺]_i mobilization. Prolonged exposure of human subcutaneous fibroblast cultures to histamine favored cell growth and type I collagen synthesis via the activation of H₁ receptor. This effect was mimicked by ATP and its metabolite, ADP, whereas the selective P2Y₁ receptor antagonist, MRS2179, partially attenuated histamine-induced cell growth and type I collagen production. Expression of pannexin-1 and ADP-sensitive P2Y₁ receptor on human subcutaneous fibroblasts was confirmed by immunofluorescence confocal microscopy and Western blot analysis. In conclusion, histamine induces ATP release from human subcutaneous fibroblasts, via pannexin-1 hemichannels, leading to Ca²⁺]_i mobilization and cell growth through the cooperation of H₁ and P2 (probably P2Y₁) receptors.