Elsevier

Brain Research

Volume 845, Issue 2, 23 October 1999, Pages 165-175
Brain Research

Research report
Masseteric inflammation-induced Fos protein expression in the trigeminal interpolaris/caudalis transition zone: contribution of somatosensory–vagal–adrenal integration

https://doi.org/10.1016/S0006-8993(99)01913-7Get rights and content

Abstract

The effects of vagotomy and adrenalectomy on the expression of Fos protein in brainstem neurons following the inflammation of masseter muscle were examined in order to differentiate the Fos activation related to nociceptive processing in contrast to that due to somatoautonomic processing. The inflammation was induced by a unilateral injection of complete Freund's adjuvant (CFA) into the masseter muscle under methohexital anesthesia after a small skin-cut (S-cut). After the CFA injection, Fos positive neurons were identified in bilateral spinal trigeminal nucleus (VSP), nucleus tractus solitarius (NTS), ventrolateral medulla (VLM) and inferior medial olivary nucleus (IOM). At the level of the trigeminal subnucleus interpolaris/caudalis (Vi/Vc) transition zone, there was a selective induction of Fos-like immunoreactivity (LI) in the VSP and NTS, when compared to control rats (anesthesia with or without S-cut). A major portion of the Fos-LI in the VSP at the level of the caudal Vc was apparently activated by S-cut. Bilateral adrenalectomy or a unilateral vagotomy resulted in a selective reduction of inflammation-induced Fos-LI in the VSP at the Vi/Vc transition zone (P<0.05) and NTS (P<0.05), but had less effect on Fos-LI in the caudal Vc. These results suggest that the inflammation of the masseter muscle, an injury of orofacial deep tissue, results in a widespread change in neuronal activity in the VSP and NTS that depends in part on the integrity of the adrenal cortex and vagus. Thus, in addition to somatotopically organized nociceptive responses, orofacial deep tissue injury also is coupled to somatovisceral and somatoautonomic processing that contribute to central neural activation.

Introduction

The expression of Fos, the protein product of an immediate early gene (IEG), c-fos, has been widely used as a measure of neural activation. The change in the expression of IEGs induced by various stimuli may contribute to the establishment of long-term functional changes in the central nervous system (CNS) following nociceptive stimulation 7, 17, 31, 32. The induction of Fos-like immunoreactivity (LI) in the medullary and rostral cervical spinal dorsal horns after noxious orofacial stimulation has been systematically studied 1, 8, 14, 15, 26, 34, 45, 46, 48, 51. Following noxious orofacial stimulation, Fos-LI is induced in widespread brainstem structures including the spinal trigeminal nucleus (VSP), the nucleus tractus solitarius (NTS), the paratrigeminal nucleus, the lateral reticular nucleus, and the inferior olivary nucleus.

Although a major portion of the Fos-LI observed in these previous studies is coupled to afferent input originating at the injury site, some of it appears to be induced by non-trigeminal somatic and visceral input 14, 34, 45, 51. It has been proposed that some Fos-LI is indirectly related to primary afferent input from the stimulated site due to polysynaptic input, or that it reflects neuronal activity associated with somatoautonomic processing, descending control and environmental stress 34, 45, 51. For example, Fos-LI is often induced on the side contralateral to orofacial stimulation and numerous Fos-labeled neurons are found in the bilateral NTS 14, 29, 51. The Fos-labeling in the ventral portion of the subnucleus interpolaris/caudalis (Vi/Vc) transition zone may be involved in autonomic responses after corneal stimulation [4]. Additionally, the expression of Fos in the VSP was also observed following electrical stimulation of the vagus nerve 11, 50. It is also well known that the adrenal steroids modulate the expression of IEGs and neuronal activity in CNS 3, 23, 25, 26, 28, 36. The purpose of the present study was to directly test the hypothesis that Fos-LI after orofacial inflammation is partly related to neural activity associated with autonomic processing and stress, indirect reactions to the inflammation and hyperalgesia.

Inflammation of the masseter muscle activates small myelinated (Group III or Aδ) and unmyelinated (Group IV or C) fibers, and second-order neurons in the VSP [33]. The injection of the small-fiber irritant, mustard oil, to the deep masseter muscle produces a short-lasting facilitatory effect on nociceptive neurons in VSP [16]. We have used complete Freund's adjuvant (CFA) to induce models of persistent orofacial inflammation 19, 51. In the present study, CFA was injected into the masseter muscle to produce inflammation in orofacial deep tissue. We first examined the regions of the VSP and other brainstem nuclei that are activated by the masseteric inflammation, using expression of Fos protein as a marker of neuronal excitation. We then investigated the contribution of the adrenals and vagal afferent input to the widespread expression of Fos protein in the brainstem following inflammation of the masseter muscle. The results indicate that unilateral masseteric inflammation induces Fos-LI in bilateral VSP and other brainstem nuclei. A portion of the Fos expression in the trigeminal Vi/Vc transition zone was related to anesthesia, and both adrenalectomy and vagotomy reduced the number of Fos positive neurons in VSP and NTS after masseteric inflammation. Some of these findings have been reported in abstract form [18].

Section snippets

Animal preparation

Male Sprague–Dawley rats (Harlan, Indianapolis, 200–395 g) were kept under 12 h light–dark cycles with free access to food and water. CFA (Mycobacterium tuberculosis, Sigma) suspended in an oil:saline (1:1) emulsion was used as the inflammatory agent. Rats were divided into the following eight groups (n=3 per group).

(1) The masseter-CFA (M-CFA) group received a unilateral injection of CFA (0.05 ml, 25 μg Mycobacterium tuberculosis) into the masseter muscle after the muscle was exposed by a 5-mm

Fos-LI induced by masseteric inflammation vs. S-cut alone

Two hours after the injection of CFA into the masseter muscle, Fos-LI neurons appeared throughout the VSP from the Vi to the C1–3 cervical spinal dorsal horn. The Fos-LI exhibited a bimodal distribution with one peak centered at the periobex level and the other peak at the caudal Vc. The Fos-LI at the periobex level was bilateral but significantly greater ipsilaterally (P<0.05). At the caudal Vc, the Fos-LI was predominantly ipsilateral to the CFA injection (Fig. 1Fig. 2AFig. 3, M-CFA). A

Discussion

The present results show that the masseteric inflammation-induced Fos-LI in the brainstem and upper cervical spinal cord can be attributed to several factors. Two new findings have emerged from these studies. First, the caudal Vc and Vi/Vc transition zone process cutaneous and deep muscle input differentially. By comparing the masseteric inflammation induced Fos-LI to that induced by anesthesia and S-cut, it was found that the bilateral but ipsilaterally dominant periobex peak of Fos-LI in

Acknowledgements

We thank Ms. S. Zou and Ms. E.B. Wade for excellent technical assistance. Supported by NIH grant DE11964.

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