Multiple G-protein involvement in parathyroid hormone regulation of acid production by osteoclasts

✍ Scribed by Lisa G. May; Carol V. Gay

• Publisher: John Wiley and Sons
• Year: 1997
• Tongue: English
• Weight: 135 KB
• Volume: 64
• Category: Article
• ISSN: 0730-2312
• DOI: 10.1002/(sici)1097-4644(199701)64:1<161::aid-jcb18>3.0.co;2-o

✦ Synopsis

The involvement of multiple G-proteins in parathyroid hormone regulation of acid production was demonstrated in a highly enriched osteoclast population. Osteoclasts were isolated from the endosteum of 2.5 to 3-week-old chicken tibia using sequential enzymatic digestion. Single cell analysis of acid production was accomplished using microscope photometry and vital staining with acridine orange, a hydrogen ion concentration sensitive fluorescent dye. Lithium chloride, an uncoupler of G-proteins from their respective receptors, blocked parathyroid hormone stimulated production of acid. Cholera toxin, which permanently activates G s -proteins, mimicked PTH stimulation. Pertussis toxin, which prevents receptor interaction with G i -and G o -proteins, blocked both 10 28 M and 10 211 M PTH stimulated acid production, suggesting that the pertussis toxin-sensitive G-protein is utilized at both PTH concentrations. Immunoblots of osteoclast plasma membrane proteins, using a panel of antibodies generated against specific G-protein a subunits, revealed a 48 kDa G s a, a 41 kDa G o a, a 34 kDa G i a-3, and a unique 68 kDa Ga subunit, with the 41 kDa and 34 kDa bands being the most intense. Immunoblots of osteoblast plasma membrane proteins had a substantially different profile with the most intense bands being a G s a (48 kDa) and a G o a (36 and 38 kDa). The studies suggest the utilization of at least two different G-proteins in the parathyroid hormone regulation of acid formation by osteoclasts, a G s and a pertussis toxin-sensitive G-protein (G o and/or G i a-3).