Studies were undertaken to determine whether PTH-related
protein (
PTHrP) (107-139) mobilizes [Ca(2+)](i) in osteoblastic
osteosarcoma UMR 106 cells.
PTHrP (107-139), in a manner similar to
PTHrP (107-111), induced a rapid [Ca(2+)](i) response in these cells that was dose dependent (EC(50) of approximately 0.1 pM) and more efficient than that of
PTHrP (1-36) (EC(50) of approximately 1 nM). This effect of
PTHrP (107-139) was abrogated by micromolar doses of
verapamil or
nifedipine. However, it was unaffected by 10 microM
U73122 (a
phospholipase C inhibitor), 100 microg/ml
heparin (an
inositol 1,4,5-trisphosphate receptor inhibitor), or 400 ng/ml
pertussis toxin (a G(i) inhibitor), which inhibited the [Ca(2+)](i) response to
PTHrP (1-36), or by either 25 nM
bisindolylmaleimide I (BIM), a
protein kinase (PK) C inhibitor, or 1 microM phorbol-12-myristate-13-acetate preincubation (22 h).
PTHrP (107-139) and
PTHrP (1-36), at 100 nM, desensitized the [Ca(2+)](i) response to a second challenge with the same
peptide, but not with the other
peptide in these cells.
PTHrP (7-34), a type 1
PTH/PTHrP receptor (PTH1R) antagonist, decreased the effect of
PTHrP (1-36) on [Ca(2+)](i). In contrast,
PTHrP (107-111), but neither
PTHrP (109-138) nor
PTHrP (7-34), abolished this effect of
PTHrP (107-139). Both
PTHrP (107-139) and
PTHrP (1-36), added together at submaximal doses, induced a higher [Ca(2+)](i) response. Moreover,
PTHrP (107-139) increased the efficacy of
PTHrP (1-36) on [Ca(2+)](i), but decreased its induced increase in PKA activity in these cells.
Verapamil or
nifedipine (at 50 microM) or 25 nM BIM, but not 25 microM
adenosine 3',5'-cyclic monophosphorothioate, Rp-isomer, a
PKA inhibitor, abolished the
PTHrP (107-139)-induced increase in
interleukin 6 messenger RNA (assessed by RT, followed by PCR) in UMR 106 cells. This
peptide also increased c-fos
messenger RNA in these cells; an effect inhibited by BIM, but unaffected by either
verapamil or
EGTA. These findings support the existence of high-affinity receptors for
PTHrP (107-139), associated with an induced Ca(2+) influx, different from the PTH1R in UMR 106 cells. The present results suggest that
PTHrP could affect bone turnover by interacting with the PTH1R and other yet unknown receptors in bone cells through complex mechanisms.