Membrane signalling and progesterone in female and male osteoblasts. I. Involvement of intracellular Ca2+ inositol trispbosphate, and diacylglycerol,but not cAMP

Bone is a target tissue of progestins, but the mechanisms by which they act are still unclear. We examined the early (5-60 s) effects of progesterone and progesterone covalently bound to BSA (P-CMO BSA), which does not enter the cell, on the cytosolic free Ca2+ concentration (ICa2+]i) and the format ion of inositol 1,4,5 trisphosphate (InsP3) and diacylglycerol (DAC) in con fluent Female and male rat osteoblasts. P-CMO BSA like Free progesterone in creased [Ca2+]i via Ca2+ influx through L-type Ca2+ channels and Ca2+ mobil ization from the endoplasmic reticulum. Both progestins increased InsP(3) a nd DAC formation within 10 s, and the increase was blocked by phospholipase C inhibitors (neomycin and U-73122). Progesterone and P-CMO BSA mobilized calcium from the endoplasmic reticulum via the activation of a phospholipas e C linked to a pertussis toxin-insensitive G-protein in both osteoblast ty pes, and this process was controlled by protein kinase C. Neither progestin had any effect on cAMP formation in male and female osteoblasts. The membr ane effects were not blocked by a progesterone nuclear antagonist. They wer e independent of the concentration of nuclear receptors and not linked to g ender. Thus, progesterone appears to act in female and male rat osteoblasts via unconventional cell-surface receptors which belong to the class of mem brane receptors coupled to phospholipase C via a pertussis toxin-insensitiv e C-protein. The bifurcating pathways leading to the formation of InsP(3) a nd DAG may provide a certain flexibility in controlling cell responses, bot h by their nature and by their rates of formation and degradation. (C) 2000 Wiley-Liss, Inc.