Evidence for a dense and intimate innervation of the bone tissue, including glutamate-containing fibers

The recent demonstration in bone cells of receptors for glutamate (Glu), a major neuromediator, suggests that Glu may also act as a signaling molecule in bone and regulate bone cell metabolism. Although bone is known to be innervated, the distribution and characteristics of nerve fibers in this tissue have not been well documented. We have studied the anatomical distribution of nerve fibers and the presence of glutamate-immunoreactive ones in sections of long bones from neonatal, 15-, and 25-day-old rats, using immunocytochemistry with antibodies directed against several neuronal markers and Glu. We showed by electron microscopy that bone is rich in nerve-like processes running along vessels adjacent to bone trabeculae, in the vicinity of hematopoı̈etic cells and bone cells. Immunocytochemical studies at the tissue and cellular level confirmed the presence of a dense network of thin nerve processes immunolabeled for neurofilament 200, tyrosine hydroxylase, and microtubule associated protein-2, three markers of nerve fibers. Some of these nerve processes showed local dilatations in contact with medullary cells and bone cells that were immunolabeled for synaptophysin, a nerve terminal marker. Glu was largely expressed in these thin nerve processes in proximity to bone cells. These findings show evidence for a dense and intimate network of nerve processes in bone, some of which were containing Glu, suggesting glutamatergic innervation in bone.