Since periodontal ligament (PDL) cells exhibit several osteoblastic traits, we hypothesized that human PDL cells will respond to hormonal stimulation in an osteoblast-like manner. Confluent and pre-confluent PDL cells from six patients were challenged with PTH(1-34). Cell number, ALP, osteocalcin, osteoprotegerin, and RANKL expression were determined. Intermittent PTH(1-34) treatment of confluent PDL cells caused a significant increase in proliferation, whereas differentiation and osteoprotegerin production decreased significantly. In pre-confluent PDL cells, this treatment regimen induced a biphasic decrease in proliferation, but a biphasic increase in differentiation and osteoprotegerin production. Continuous PTH(1-34) exposure enhanced proliferation but inhibited osteocalcin production in confluent cells and stimulated osteoprotegerin production in pre-confluent PDL cells. RANKL was hardly detectable and unaffected by PTH(1-34) treatment. These results indicate that human PDL cells respond to PTH(1-34) in an osteoblast-like manner, and that the PTH(1-34) effect depends on the maturation state of the cells and on the mode of administration.