In this study, we used genetic and pharmacological means to formally identify the osteoblast as the major cell type in which the sympathetic tone, under the control of leptin signaling in the brain, acts to inhibit bone mass accrual. Results of this investigation are represented schematically in Fig. S1. We cannot exclude, based on our results, the possibility that osteocytes may also be a target of the sympathetic tone. We should emphasize here that our study is restricted to the means whereby leptin signaling in the brain, as represented by leptin ICV infusion, regulates bone mass. The reason for this focus on this aspect of leptin regulation of bone mass accrual is that a neuron-specific inactivation of the leptin receptor in the mouse recapitulates the bone phenotype of mice lacking leptin signaling, whereas its osteoblast-specific inactivation fails to do so (Shi et al., 2008
Once it was realized that there was a central control of bone mass (Ducy et al., 2000
), the next question was to decipher how the brain signals to the bone. It became rapidly apparent that the sympathetic nervous system is a critical mediator of this homeostatic function exerted by the brain. This signaling inhibits bone mass accrual by, at the same time, preventing bone formation and favoring bone resorption. The evidence supporting this view was genetic and relied on the study of a classical gene inactivation mouse model. Indeed, mice lacking the β2 adrenergic receptor in all cells had a high bone mass secondary to an increase in osteoblast proliferation and bone formation and a subsequent decrease in bone resorption (Elefteriou et al., 2005
). It was also supported by a clinical aspect such as the low bone mass of patients affected with reflex sympathetic dystrophy (Schwartzman, 2000
Several transcription factors have been implicated in mediating this function of the sympathetic tone, and it has long been assumed, although never tested, that this regulation takes place in the osteoblasts (Elefteriou et al., 2005
; Fu et al., 2005
). However, because many of these molecular players such as Adrβ2, CREB, and cMyc are by no means osteoblast-specific molecules, there was an urgent need to demonstrate that indeed it is in osteoblasts that this pathway operates. What the experiments presented in this study establish is that it is by acting in the osteoblasts that the sympathetic tone regulates bone mass accrual. It is also through its expression in osteoblasts that CREB regulates osteoblast proliferation downstream of the sympathetic tone. Likewise, it is through its expression in osteoblasts that ATF4 favors Rankl
expression and osteoclast differentiation downstream of the sympathetic tone. It is noteworthy that these two transcription factors have distinct functions. Indeed, CREB regulates osteoblast proliferation but does not affect osteoclast differentiation, whereas ATF4 affects osteoclast differentiation but is not involved in osteoblast proliferation.
In considering the overall sympathetic mode of regulating bone mass accrual, it is quite remarkable that the sympathetic tone recruits two different transcription factors in the same cell to regulate each arm of bone remodeling. Remarkably, this specificity is conserved when one looks at the main regulator of this function of the sympathetic nervous system, leptin. Indeed, long-term ICV infusion of this hormone in mutant mice lacking, in osteoblasts only, either Creb or Atf4 verified that leptin and the sympathetic tone recruit each of these two transcription factors for different purposes.
Looking more globally at these regulations of bone mass by leptin, it underscores the importance of the various roles played by CREB at several steps in this pathway. Indeed, CREB is mediating serotonin regulation of the sympathetic tone in the ventromedial hypothalamic nuclei, a regulation inhibited by leptin signaling in the brain. Thus, CREB acts upstream of the sympathetic tone (Oury et al., 2010
). The present study shows that it also acts downstream of it and in osteoblasts.
This work, by showing that the bulk of the sympathetic regulation of bone mass accrual occurs in osteoblasts, raises the question of its pharmacological relevance. The work of Bonnet et al. (2008)
showing that a low dose of β blockers acting through Adrβ2 could prevent gonadectomy-induced bone loss without affecting other function regulated by the sympathetic nervous system is in full agreement with the fact that mice lacking only one allele of Adrβ2
display high bone mass. As such, it is certainly important if this pathway could be exploited further for the purpose of treating low bone mass disease.