Lonnie R. Johnson

Sodium channel Nav1.6 accumulates at the site of infraorbital nerve injury

BackgroundSodium channel (NaCh) expressions change following nerve and inflammatory lesions and this change may contribute to the activation of pain pathways. In a previous study we found a dramatic increase in the size and density of NaCh accumulations, and a remodeling of NaChs at intact and altered myelinated sites at a location just proximal to a combined partial axotomy and chromic suture lesion of the rat infraorbital nerve (ION) with the use of an antibody that identifies all NaCh isoforms. Here we evaluate the contribution of the major nodal NaCh isoform, Nav1.6, to this remodeling of NaChs following the same lesion. Sections of the ION from normal and ION lesioned subjects were double-stained with antibodies against Nav1.6 and caspr (contactin-associated protein; a paranodal protein to identify nodes of Ranvier) and then z-series of optically sectioned images were captured with a confocal microscope. ImageJ (NIH) software was used to quantify the average size and density of Nav1.6 accumulations, while additional single fiber analyses measured the axial length of the nodal gap, and the immunofluorescence intensity of Nav1.6 in nodes and of caspr in the paranodal region.ResultsThe findings showed a significant increase in the average size and density of Nav1.6 accumulations in lesioned IONs when compared to normal IONs. The results of the single fiber analyses in caspr-identified typical nodes showed an increased axial length of the nodal gap, an increased immunofluorescence intensity of nodal Nav1.6 and a decreased immunofluorescence intensity of paranodal caspr in lesioned IONs when compared to normal IONs. In the lesioned IONs, Nav1.6 accumulations were also seen in association with altered caspr-relationships, such as heminodes.ConclusionThe results of the present study identify Nav1.6 as one isoform involved in the augmentation and remodeling of NaChs at nodal sites following a combined partial axotomy and chromic suture ION lesion. The augmentation of Nav1.6 may result from an alteration in axon-Schwann cell signaling mechanisms as suggested by changes in caspr expression. The changes identified in this study suggest that the participation of Nav1.6 should be considered when examining changes in the excitability of myelinated axons in neuropathic pain models.

Increased sodium channel immunofluorescence at myelinated and demyelinated sites following an inflammatory and partial axotomy lesion of the rat infraorbital nerve

Abstract The localization of sodium channels (NaChs) change following nerve lesions and this change may contribute to the development of increased pain states. Here we examine the change in distribution of NaChs within the rat infraorbital nerve (ION) two weeks after a combined inflammatory/partial axotomy lesion that results in behavior showing increased sensitivity to mechanical stimuli. Sections from experimental and normal control IONs were double‐stained for indirect immunofluorescence using an antibody that identifies all NaCh isoforms and caspr‐antibody to identify nodes of Ranvier, and a confocal microscope z‐series of optically sectioned images were then obtained. ImageJ (NIH) software was used to quantify the area of pixels showing maximum NaCh intensity within both caspr and non‐caspr associated accumulations. Analysis showed that the lesioned IONs had many more split nodes, heminodes and caspr‐negative “naked” accumulations, a significantly increased area of NaCh staining within typical nodes and “naked” accumulations, as well as an increased density and size of significant accumulations when compared to normal IONs. This study demonstrates a dramatic redistribution and increased immunofluorescence of NaChs especially at myelinated and demyelinated sites in fibers located just proximal to the lesion. The remodeling of NaChs seen in this study may represent an important event associated with the development of increased nerve excitability after lesions.

Sodium channel expression and localization at demyelinated sites in painful human dental pulp.

UNLABELLED The expression of sodium channels (NaCh(s)) change after inflammatory and nerve lesions, and this change has been implicated in the generation of pain states. Here we examine NaCh expression within nerve fibers from normal and painful extracted human teeth with special emphasis on their localization within large accumulations, like those seen at nodes of Ranvier. Pulpal tissue sections from normal wisdom teeth and from teeth with large carious lesions associated with severe and spontaneous pain were double-stained with pan-specific NaCh antibody and caspr (paranodal protein used to visualize nodes of Ranvier) antibody, while additional sections were triple-stained with NaCh, caspr and myelin basic protein (MBP) antibodies. Z-series of images were obtained with the confocal microscope and evaluated with NIH ImageJ software to quantify the density and size of NaCh accumulations, and to characterize NaCh localization at caspr-identified typical and atypical nodal sites. Although the results showed variability in the overall density and size of NaCh accumulations in painful samples, a common finding included the remodeling of NaChs at atypical nodal sites. This remodeling of NaChs included prominent NaCh expression within nerve regions that showed a selective loss of MBP staining in a pattern consistent with a demyelinating process. PERSPECTIVE This study identifies the remodeling of NaChs at demyelinated sites within the painful human dental pulp and suggests that the contribution of NaChs to spontaneous pulpal pain generation may be dependant not only on total NaCh density but may also be related to NaCh expression at atypical nodal sites.

Ultrastructure of degenerative changes following ricin application to feline dental pulps

SummaryThe ultrastructure of degenerative changes within the ipsilateral trigeminal ganglion, and partes caudalis and interpolaris of the spinal trigeminal nucleus in the cat is described following the application of the potent toxin ricin to the tooth pulps of unilateral maxillary and mandibular posterior teeth, including the cuspids. Survival times ranged from 6 to 10 days. Typical changes identified within the ipsilateral trigeminal ganglion included myelin fragmentation and ‘compartmentalization’ of the axoplasm of medium-sized myelinated axons, while small myelinated and unmyelinated axons underwent a more variable response ranging from electron-lucent to electron-dense changes. The affected cell body was characterized by the presence of swollen, electron-lucent mitochondria, a reduction of cytoplasmic ribosomes and a filamentous hyperplasia. Other changes often included an eccentric nucleus and satellite cell proliferation. Degenerative changes often occurred in isolated elements surrounded by normal profiles, suggesting specificity of ricin within the trigeminal ganglion. Changes within brainstem axons showed both an electron-dense and a lucent, fragmenting type of axonal alteration. Terminal changes ranged from electron-dense to lucent and also included filamentous hyperplasia and ‘hyperglycogenesis’. The altered axonal knobs contained round synaptic vesicles that were presynaptic to dendritic profiles and postsynaptic to terminals containing flattened synaptic vesicles. The above brainstem alterations were identified specifically in the following areas: ventrolateral, medial and dorsomedial pars interpolaris; the ventrolateral and mid-dorsal to dorsomedial areas of the marginalis and outer substantia gelatinosa layers of pars caudalis; and in ventral pockets corresponding to lamina V of the medullary dorsal horn. Dense alterations within terminals containing flattened synaptic vesicles that are typically presynaptic to primary afferents in these areas were rare findings, but along with vacuolization of dendritic profiles suggest a trans-synaptic effect possibly due to the exocytosis of ricin. The results are discussed in relation to different reports of dental projections and with regards to patterns of transganglionic degeneration.

Localization of the Nav1.8 sodium channel isoform at nodes of Ranvier in normal human radicular tooth pulp

The activation of voltage-gated sodium channels is necessary for action potential propagation and multiple sodium channel isoforms have been identified that show a differential distribution throughout the nervous system. An evaluation of sodium channel localization in the radicular pulp from normal human extracted third molars established the presence of the Na(v)1.8 isoform at nodes of Ranvier in a subpopulation of the myelinated axons as demonstrated with immunofluorescence confocal microscopy. A caspr antibody was used to identify the paranodal region of nodes of Ranvier and quantitative analysis revealed that 16.5% of the nodes contained significant Na(v)1.8 immunoreactivity. Since the Na(v)1.6 isoform has been described as the predominant sodium channel at essentially all nodes, the finding of Na(v)1.8 in a subpopulation of nodes suggests that multiple isoforms may coexist at some nodes of Ranvier and also suggests that this isoform may be an important nodal sodium channel type in the peripheral sensory nervous system of humans.

Light microscopic localization of calcitonin gene-related peptide in the normal feline trigeminal system and following retrogasserian rhizotomy

Calcitonin gene‐related peptide (CGRP) is a neuropeptide that has been implicated in the transmission and modulation of primary afferent nociceptive stimuli. In this study, we describe the light microscopic distribution of CGRP immunoreactivity (IR) within the feline trigeminal ganglion and trigeminal nucleus of normal adult subjects and in subjects 10 and 30 days following complete retrogasserian rhizotomy. Within the trigeminal ganglion of normal subjects, cell bodies and fibers showed CGRP‐IR, whereas immunoreactive fibers were rare in the central root region. Within the normal spinal trigeminal and main sensory nuclei, CGRP‐IR was seen to form a reproducible pattern that varied between the different nuclei. Following rhizotomy, most, but not all, of the CGRP‐IR was lost from the spinal trigeminal and main sensory nuclei, except in regions where the upper cervical roots and cranial nerves VII, IX, and X project into the trigeminal nucleus. The pattern seen at 10 days contained more CGRP‐IR than that seen at 30 days and suggests that degenerating fibers still show CGRP‐IR. In contrast to the decrease seen in the nuclei after rhizotomy, examination of the central root that was still attached to the trigeminal ganglion showed an increase in CGRP‐IR within fibers, some of which ended in growth conelike enlargements. Rhizotomy induced a dramatic increase in CGRP‐IR within trigeminal motoneurons and their fibers, which was strongest 10 days after rhizotomy and weaker at 30 days, which was still stronger than normal. These results indicate that the majority of CGRP‐IR found in the trigeminal nucleus originates from trigeminal primary afferents and that an upregulation of CGRP‐IR occurs in trigeminal motoneurons and in regenerating fibers in the part of the central root that was still attached to the ganglion. In addition, the persistence of CGRP‐IR fibers in the trigeminal nucleus provides one possible explanation for the preservation of pain in humans following trigeminal rhizotomy.

Oral Health Knowledge and Behaviors among Adolescents with Type 1 Diabetes

Early onset and more advanced periodontal disease has been reported for children with diabetes. We surveyed oral health knowledge, attitudes, and behaviors among adolescents with diabetes in order to inform potential intervention strategies. Study subjects were youth (ages 12–19 years) with type 1 diabetes (N = 90) participating in a cohort study investigating determinants of periodontal disease at a regional pediatric diabetes specialty clinic. Over 90% of the youth had been instructed on how to brush and floss and had preventive dental care in the past year. However, 44% knew that periodontal disease is associated with diabetes and 32% knew that it can start in childhood with bleeding gums. Despite being at high risk for developing periodontal disease, the mean toothbrushing frequency was once per day and 42% did not floss. Significant opportunity exists for improving periodontal disease knowledge and adoption of preventive oral hygiene behaviors in adolescents with diabetes.

Light- and Electron-Microscopic Localization of Primary Dental Afferents to Medullary Dorsal Horn (Pars Caudalis)

Light-microscopic (LM) and ultrastructural (electron-microscopic, or EM) identification of primary dental afferents to medullary dorsal horn (MDH) was demonstrated in the cat following injections of horseradish peroxidase (HRP) into pulpal chambers of unilateral maxillary and mandibular posterior teeth, including the cuspids. Use of a new osmication protocol improved and simplified the EM localization of reaction product within the brain stem terminals. LM examination showed that the projection pattern varied between the different levels of MDH. At caudal levels, the labeling was primarily confined to a narrow band consisting of a dense projection to the dorsomedial portion of laminae I and superficial II and a less intense projection to lamina V. The pattern to rostral levels became increasingly more dense and extensive within these same laminae. LM examination of the tooth apex region showed that a limited spread to the periodontal ligament occurred in some cases. EM investigation of the ipsilateral MDH demonstrated reaction product in terminals with synaptic vesicles that are presynaptic to small and medium-sized dendrites. Labeled axonal endings in close association with cell bodies were also observed. No labeled structures were identified in the contralateral MDH. Some of the reaction product found with EM was below the LM limit of resolution, and thus ultrastructural investigation is necessary for a complete analysis of any pathway when using HRP.

Retrogasserian rhizotomy causes expression of nerve growth factor receptor-immunoreactive protein in motoneurons within the adult feline trigeminal motor nucleus

The effect of lesions on nerve growth factor receptor (NGFr) immunoreactivity (IR) in motoneurons within the mature feline trigeminal motor nucleus was investigated. Ten days following complete unilateral retrogasserian trigeminal rhizotomy including transection of the trigeminal motor root, motoneurons within the ipsilateral trigeminal motor nucleus showed NGFr-IR. In contrast, motoneurons within the contralateral trigeminal motor nucleus and within both trigeminal motor nuclei in unoperated control subjects did not show NGFr-IR. It is suggested that the appearance of NGFr-IR in motoneurons within the ipsilateral trigeminal motor nucleus after rhizotomy represents an attempts towards recovery and may be associated with the regrowth of its cut axons.

Loss of Alveolar Bone Due to Periodontal Disease Exhibits a Threshold on the Association With Coronary Heart Disease

BACKGROUND A number of epidemiologic studies were published that looked at the association between coronary heart disease (CHD) and periodontal disease. However, debate exists about whether this association is a true relationship or simply an example of an uncontrolled confounder. This retrospective cohort study examines the relationship between periodontal disease and CHD. METHODS Digital panoramic radiographs were used to assess alveolar bone loss (ABL) using a Schei ruler. Participants consisted of Veterans Administration (VA) patients who were eligible for dental benefits and had a digital panoramic radiograph taken at the VA Medical Center, Denver, Colorado. Information on CHD and other important clinical variables were obtained from electronic medical records. RESULTS The examination of the relationship between ABL and CHD revealed a significant non-linear relationship with a threshold at ≈ 20% bone loss with a doubling of the probability ratios of CHD compared to those at 7.5% bone loss. CONCLUSIONS To our knowledge, this is the first study to demonstrate a non-linear relationship between ABL and CHD. A significant positive association between ABL and CHD was found at even low levels of bone loss between 10% and 20%.