An Experimental Research Of Nerve Innervation In The Rabbit Normal Cervical Intervertebral Discs

Objective: The pathogenesis of cervical vertigo is complicated, and the sympathetic nerves factors play an important role in the pathogenesis of cervical vertigo, but the concrete mechanism of the sympathetic nerve leading to cervical vertigo is poorly understood. Recently, the therapeutic effect of percutaneous laser disc decompression for cervical vertigo is noticeable. Those facts denoted that there are some relationships between cervical intervertebral disc and the vertebral artery, and the medium of the relationship is very likely to be the sympathetic nerve. So the innervations of the cervical intervertebral disc become to be the question that attracts the most attention. So far, there is a great deal of studies on innervations of intervertebral disc, but most of the studies were concentrated in lumbar intervertebral disc. Only few studies are related to cervical intervertebral disc. Many investigations have proved that the outer third of the annulus fibrous of the lumbar intervertebral disc is innervated, especially in herniated disc tissue and in degenerated disc tissue. Those nerve fibers were demonstrated to be sensory nerve terminals and autonomic nerve terminals. Those studys also demonstrated that the dorsal portion of the lumbar intervertebral disc was innervated from the upper dorsal root ganglions. the nerve fibers enter the paravertebral sympathetic trunks through each ramus communicans at the L2 and/or more cranial levels, and reach the dorsal portion of the lumbar intervertebral disc via the corresponding ramus communicans. The studies of the lumbar intervertebral disc provided some useful experiences for the studies on innervations of the cervical intervertebral disc. The purpose of this study is to investigate whether the anulus fibrosus of rabbit normal Cervical Intervertebral Discs is innervated, the type of the never fibers and the origin of the nerve fibers, we investigated rabbit normal cervical intervertebral disc using horseradish peroxidase (HRP) tracing method and immunohistochemical method. At the same time, we selected the lumbar intervertebral disc for immunohistochemical experiment as comparison.Methods: 15 rabbits were in this study, they were divided into two groups randomly: the Intervertebral Disc (IVD) group and the Dorsal Root Ganglion (DRG) group. The number of the rabbit of the two groups was 5 and 10 respectively.1,In the Intervertebral Disc (IVD) group, the rabbits were sacrificed directly. The C5/6 cervical intervertebral disc was selected for H-E staining. Tissues were fixed in 4% neutral buffered paraformaldehyde (0.1M; pH 7.4) and embedded in paraffin wax. Each disc was serially sectioned at 5-μ, they were stained with haematoxylin and eosin to observe the histological anatomy of the normal cervical intervertebral disc. The C6/7 cervical intervertebral disc was selected for immunohistochemical experiment. Tissues were fixed in 4% neutral buffered paraformaldehyde (0.1M; pH 7.4) and embedded in paraffin wax. 5μm step serial sections were taken in each sample, the sections were stained with neurofilament protein(NF) as a general nerve marker, substance P(SP, a neurotransmitter with nociceptive activity) as a sensory marker, and C-flanking peptide of neuropeptide Y(CPON) as a sympathetic marker, and to investigate the innervation of cervical intervertebral disc. While, in the control group, the L4/5 and L5/6 lumbar intervertebral disc were selected for H-E staining and immunohistochemical experiment respectively.2,The DRG group was further divided into nontreated group (n=5) and sympathectomized group (n=5). The DRG group was used to performed horseradish peroxidase tracing experiment. In the nontreated group, the paravertebral sympathetic trunks was reserved; but in the sympathectomized group the paravertebral sympathetic trunks was abscised below the inferior cervical ganglion. The two groups were used for horseradish peroxidase experiment. The horseradish peroxidase was injected into the dorsal portion of the cervical intervertebral disc of C4/5,C5/6,C6/7 using microinjector in this group. 48 hours after the application of horseradish peroxidase to the dorsal portion of the intervertebral disc, the rabbits were sacrificed, and bilateral cervical (C4~8) and upper thoracic (T1~5) DRGs were obtained to observed the number of labeled neurons in the reserved DRGs by optical microscopy.Results: In the immunohistochemical study, we found that the lumbar intervertebral disc has an affluent innervation, This is concentrated in the perianular connective tissue. In the sections of lumbar intervertebral disc, eighty percent (4/5) showed immunoreactivity for NF in the dorsal portion of the lumbar intervertebral disc. The immunoreactivities for COPN and SP in the dorsal portion of the lumbar intervertebral discs were 60% (3/5) and 40% (2/5) respectively. Meanwhile, nerve fibers immunoreactived for NF, COPN and SP were found in the anterior portion of the lumbar intervertebral discs.Eighty percent (4/5) sections showed immunoreactivity for NF in the posterior connective tissue of the cervical intervertebral disc; the immunoreactivity for CPON in the posterior connective tissue of the cervical intervertebral disc was 20% (1/5). No sections showed immunoreactivity for SP in the posterior connective tissue of the cervical intervertebral disc.In the horseradish peroxidase experiment, no HRP labeled neuron was found both the nontreated group and sympathectomized group.Conclusion: The cervical intervertebral disc has an affluent innervation, This is concentrated in the perianular connective tissue. Those nerve fibers include the sympathetic nerve fibers immunoreactived for CPON. Those never fibers in the perianular connective tissue of the cervical intervertebral disc may participate in the pathogenesis of the cervical vertigo. The mechanisms of the cervical vertigo were still unclear.