Haruhide Hayashi

Compressive force induces VEGF production in periodontal tissues.

During orthodontic tooth movement, the activation of the vascular system in the compressed periodontal ligament (PDL) is an indispensable process in tissue remodeling. We hypothesized that compressive force would induce angiogenesis of PDL through the production of vascular endothelial growth factor (VEGF). We examined the localization of VEGF in rat periodontal tissues during experimental tooth movement in vivo, and the effects of continuous compressive force on VEGF production and angiogenic activity in human PDL cells in vitro. PDL cells adjacent to hyalinized tissue and alveolar bone on the compressive side showed marked VEGF immunoreactivity. VEGF mRNA expression and production in PDL cells increased, and conditioned medium stimulated tube formation. These results indicate that continuous compressive force enhances VEGF production and angiogenic activity in PDL cells, which may contribute to periodontal remodeling, including angiogenesis, during orthodontic tooth movement.

Pulpal and cutaneous inputs to somatosensory neurons in the parabrachial area of the cat

The majority of somatosensory neurons recorded from the mesencephalic parabrachial area and pontine parabrachial nucleus of the cat responded exclusively to noxious mechanical stimuli to the skin. Their receptive fields were very large. Two-thirds of the neurons tested responded to electrical stimulation of the tooth pulp. These results suggest that neurons in this area have extensive convergence of spinal and trigeminal inputs, and contribute to the affective or autonomic aspects of pain.

PHYSIOLOGICAL PROPERTIES OF PERIODONTAL MECHANOSENSITIVE NEURONES IN THE TRIGEMINAL (GASSERIAN) GANGLION OF THE RAT

Periodontal mechanosensitive (PM) primary afferent neurones were recorded from the rat trigeminal ganglion and their response to mechanical tooth stimulation studied. The majority (95%) of PM afferents were single-tooth units and most were sensitive to mechanical stimulation of the incisor. They had a sustained response to pressure applied to the tooth, and showed a directional selectivity to the stimulation. Only a small number (5%) of units were multi-tooth sensitive; their receptive fields were restricted to the molars. When the incisor was stimulated mechanically in 12 directions in a plane perpendicular to the axis of the tooth crown, the predominant response fields of the PM units were of the medium type (90-180 degree response angle) or the broad type (180-360 degrees). Each unit had single optimal stimulus direction oriented predominantly in the rostrocaudal or caudorostral direction. About 60% of the PM units responded to tooth stimulation at irregular spike intervals, whereas the remaining 40% fired at relatively regular intervals. When the tooth was stimulated at a force of 0.05 N, the mean spike interval and rate of spike-interval fluctuation [percentage of the standard deviation (SD) of the interval distribution to the mean interval (mean); SD/mean x 100] were 29.2 ms and 38.1% for the regular-interval units, and 15.1 ms and 8.8% for the irregular-interval ones. The mean spike interval of the regular units was significantly longer than that of the irregular ones.

The inhibitory effect of substance P antagonist, CP-96,345, on the late discharges of nociceptive neurons in the rat superficial spinal dorsal horn

The effects of local spinal applications of the substance P antagonist, CP-96,345, on the electrically-evoked discharges of nociceptive neurons were examined in the superficial spinal dorsal horn of adult rats anesthetized with sodium pentobarbital. Extracellular single-unit recordings were made from lumbar spinal dorsal horn neurons which were excited by noxious mechanical stimulation and had early and late discharges evoked by A- and C-fiber inputs, respectively, following transcutaneous electrical stimulation. CP-96,345 was applied directly onto the surface of the spinal cord close to the recording sites. A dose of 50 nmol of CP-96,345 drastically inhibited the late discharges of 80% of neurons, without affecting early discharges significantly. This study demonstrated the specific inhibitory effect of CP-96,345 on the late discharges of a restricted population of spinal dorsal horn neurons. These results illustrate the important role of substance P in nociceptive transmission mediated by primary afferent C-fibers in normal acute pain.

Dorsomorphin stimulates neurite outgrowth in PC12 cells via activation of a protein kinase A‐dependent MEK‐ERK1/2 signaling pathway

In this study, we investigated the effect of dorsomorphin, a selective inhibitor of bone morphogenetic protein (BMP) signaling, on rat PC12 pheochromocytoma cell differentiation. PC12 cells can be induced to differentiate into neuron‐like cells possessing elongated neurites by nerve growth factor, BMP2, and other inducers. Cells were incubated with BMP2 and/or dorsomorphin, and the extent of neurite outgrowth was evaluated. Unexpectedly, BMP2‐mediated neuritogenesis was not inhibited by co‐treatment with dorsomorphin. We also found that treatment with dorsomorphin alone, but not another BMP signaling inhibitor, LDN‐193189, induced neurite outgrowth in PC12 cells. To further understand the mechanism of action of dorsomorphin, the effects of this drug on intracellular signaling were investigated using the following signaling inhibitors: the ERK kinase (MEK) inhibitor U0126; the tropomyosin‐related kinase A inhibitor GW441756; and the protein kinase A (PKA) inhibitor H89. Dorsomorphin induced rapid and sustained ERK1/2 activation; however, dorsomorphin‐mediated ERK1/2 activation and neuritogenesis were robustly inhibited in the presence of U0126 or H89, but not GW441756. These findings suggest that dorsomorphin has the potential to induce neuritogenesis in PC12 cells, a response that requires the activation of PKA‐dependent MEK‐ERK1/2 signaling.

Morphology of central terminations of intra-axonally stained, low-threshold mechanoreceptive primary afferent fibers from oral mucosa and periodontium in the rat

Horseradish peroxidase (HRP) was injected intra-axonally into functionally identified primary afferent fibers within the rat spinal trigeminal tract in order to study the morphology of their central terminations. They were physiologically determined to be large, myelinated afferent fibers from periodontium or oral mucosa by means of electrical and mechanical stimulation of their receptive fields. Twenty-eight axons that innervated the periodontium of incisors and 21 axons that innervated the oral mucosa were stained for distances of 2-5 mm from the injection sites at the levels of the main sensory nucleus (Vms), spinal trigeminal nucleus and rostral cervical spinal cord. The collaterals of these primary afferent fibers formed terminal arbors in the medial or dorsomedial part of the Vms, and the oral and interpolar spinal trigeminal nuclei (Vo and Vi). In the caudal spinal trigeminal nucleus (Vc), the collaterals of one half of the periodontium afferent fibers terminated mainly in lamina V at the rostral and middle levels of Vc. On the other hand, the collaterals of the other half of the periodontium afferent fibers terminated mainly in lamina IV at the rostral level of Vc, and rostrally these terminal areas shifted to the most medial part of Vi. The collaterals of mucosa afferent fibers terminated in lamina V at the rostral level of Vc, and these terminal areas shifted gradually to laminae III and IV as the parent axons traveled more caudally. These shifts were staggered rostrocaudally according to the rostrocaudal locations of the receptive fields. The density of collaterals of periodontium afferent fibers in Vi was significantly larger than that of mucosa afferent fibers. The average size of the varicosities of periodontium afferent fibers was significantly larger than those of mucosa afferent fibers in Vo, Vi and Vc. The average number of varicosities belonging to single collaterals of slowly-adapting periodontium afferent fibers in Vi were significantly larger than those in Vo. In Vi, the average number of varicosities of single collaterals of slowly-adapting periodontium afferent fibers were significantly larger than those of rapidly-adapting periodontium afferent fibers.

DISTRIBUTION OF TRIGEMINAL SENSORY NUCLEUS NEURONS PROJECTING TO THE MESENCEPHALIC PARABRACHIAL AREA OF THE CAT

Horseradish peroxidase (HRP) retrograde tracing experiments in the cat demonstrated projections from the trigeminal sensory nuclei to the mesencephalic parabrachial area (PBA), which is located ventral to the inferior colliculus and dorsal to the brachium conjunctivum and includes the nucleus cuneiformis and the most lateral part of the periaqueductal gray. After HRP injection into the mesencephalic PBA, HRP-labeled neuronal cell bodies in the trigeminal sensory nuclei were mainly distributed in the interpolar and caudal spinal trigeminal nuclei (Vi and Vc) bilaterally with a clear-cut contralateral dominance. In Vi, HRP-labeled neurons were mainly distributed in the lateral and medial border regions. In Vc, labeled neurons were located mainly in laminae I and V.

Response properties of periodontal mechanosensitive neurones in the rat trigeminal sensory complex projecting to the posteromedial ventral nucleus of the thalamus

Unitary discharges from periodontal mechanosensitive (PM) neurones responding to mechanical stimulation of the tooth were recorded from the trigeminal sensory complex in the rat brainstem. Of the PM units recorded, 22% were activated by antidromic stimulation of the contralateral (20%) or ipsilateral (2%) posteromedial ventral nucleus of the thalamus. Although thalamic-projecting neurones were recorded extensively throughout the trigeminal sensory complex, they originated most often in the region from the caudal main sensory nucleus to the rostral subnucleus oralis of the trigeminal spinal tract nucleus. The response latencies of the rostral nucleus units to orthodromic stimulation of peripheral receptive fields and antidromic stimulation of the thalamus were significantly shorter than those of the caudal nucleus units. More than half were single-tooth units originating from incisor teeth. They responded continuously when pressure was applied to the tooth. The magnitude of the response varied with the direction of the stimulus. Maximal responses were obtained when the stimulus was applied labiolingually or vice versa. The threshold for mechanical stimulation of the tooth was less than 0.05 N. The rostrocaudal distribution and response properties of thalamic-projecting PM neurones were very similar to those of non-thalamic-projecting PM units that were not activated by antidromic stimulation of the thalamus.

Physiological properties of periodontal mechanosensitive neurones in the posteromedial ventral nucleus of rat thalamus.

Unitary discharges of periodontal mechanosensitive (PM) neurones responding to mechanical tooth stimulation were recorded from the posteromedial ventral nucleus (VPM) of rat thalamus. PM neurones are distributed in the ventromedial area in the rostral two-thirds of the VPM nucleus. Maxillary and mandibular tooth-sensitive neurones are arranged in dorsoventral sequence. Of the PM neurones, 36% were slowly adapting to pressure applied to the tooth and 67% were rapidly adapting. The majority of PM units were sensitive to the contralateral incisor tooth. Response magnitudes of the slowly adapting neurones varied with stimulus direction and were directionally selective to mechanical tooth stimulation. The optimal stimulus direction was labiolingual or linguolabial. Rapidly adapting neurones were directionally non-selective to tooth stimulation. The threshold for mechanical stimulation was <0.05 N. Mean response latencies evoked by electrical stimulation of the peripheral receptive fields were 4.6 ms in the slowly adapting neurones and 5.8 ms in the rapidly adapting neurones.

Functional Organization in the Orofacial Region of the Postcentral Somatosensory Cortex

Abstract The first somatosensory cortexiSIjof primates is located in the postcentral gyrus of the parietal lobe. The orofacial structures are represented most laterally in SI. The inspection of neuronal properties there should lead to understanding of the neural basis that underlie dexterous orofacial functions, such as speech, mastication, manipulation of objects, and oral stereognosis. In the orofacial representation of SI, a substantial number of neurons have unique receptive fields (RFs) resulting from the spatiotemporal integration of converging somesthetic inputs. The relative incidences of those unique neurons increase on moving caudally from area 3 towards area 2. A neural process that binds spatiotemporal information arising from functionally-related portions might enable the brain to monitor the movement of objects in contact with orofacial structures or the kinematic trajectory of orofacial structures themselves. On the other hand, the modular organization of the neocortex is a widely documented concept, in which neural connectivity, composed of nearby cortical neurons, is considered to be the functional unit of integration. From this viewpoint, studies are needed to compare physiological properties among neurons localized in a small region of the orofacial representation. As for the RF extension, a small but substantial proportion of the pairs of nearby neurons are associated with discrete but functionally-related oral portions of different structures. As for the temporal aspects, a study is now underway in our laboratory to reveal the temporal relation between the activities of nearby neurons during sustained natural stimuli.