• Restorative Dentistry and Endodontics
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• v.24 no.1
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• pp.100-115
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• 1999

The purpose of this study was to investigate the distribution of calcitonin gene-related peptide containing nerve fibers in rat pulp after dentinl injury by means of immunohistochemistry and confocal laser scanning microscope. The Spague-Dawley rats weighing about 250-300gm were used. The animals were devided into normal control and experimental groups. Experimental animals were sacrified 1, 2, 4, 7, 10, 21days after dentinal injury (dentin cutting, and then acid etching with 35% phosphoric acid) on the maxillary molar teeth. The maxillary teeth and alveolar bone were removed and immersed in the 4% paraformaldehyde in 0.1M phosphate buffer (pH 7.4), then were decalcified with 15% formic acid for 10 days. Serial frozen $50{\mu}m$ thick sections were cut on a cryostat. The rabbit CGRP antibody was used as a primary antibody with a dilution of 1:2000 in 0.01M PB. The sections were incubated for 48 hours at $4^{\circ}C$, and placed into biotinylated antirabbit Ig G as a secondary anti body with dilution of 1:200 in 0.01M PB and incubated in ABC(avidin-biotin complex). The peroxidase reaction was visualized by incubating the sections in 0.05% 3,3 diaminobenzidine tetrahydrochloride containing 0.02% $H_2O_2$. For the confocal laser scanning microscopic examination, Primary antibody reaction was same as immunoperoxidase stainning, but fluorescein isothiocyanate(FITC)-conjugate antirabbit IgG as a secondary antibody was used. The confocal laser scanning microscope was used for the examination. A series of images of optical sections was collected with a 20x objective at $3{\mu}m$ intervals throughout the depth of specimen. FITC fluerescence was registrated through a 488nm and 568nm excitation filter, and images were saved on optical disk. The stereoscopic images and three dimentionnal images were reconstructed by computer software, and then were analyzed. The results were as follows : 1. In normal control group, CGRP containing nerve fibers were coursed through the root with very little branching, and then formed a dense network of terminals in coronal pulp. 2. A slight increase in CGRP containing nerve fibers at 1 and 2day postinjury was noted subjacent to the injury site. In the 4day group, there were an extensive increase in the number of reactive fibers, followed by a partial return toward normal levels at 7~10 day postinjury, and return by 21days. 3. The sprouting of the CGRP containing nerve fibers was evident within 2day after dentinal injury, and by 4days there was a maximal increased, but was decreased at 7days and returned to normal 10~21 day postinjury. 4. In confocal laser scanning microscopic exammination, the distinct distribution pattern and sprouting reaction of CGRP containing nerve fibers were observed in stereoscopic images and three dimentional images. These results suggest that CGRP containing nerve fiber can be important role in the response to dental injury and pain regulation.

• Korean Journal of Veterinary Research
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• v.39 no.3
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• pp.435-447
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• 1999

The localization of substance P(SP), calcitonin gene-related peptide(CGRP) and three calcium binding proteins, calbindin D-28k(CB), calretinin(CR) and parvalbumin(PA) was immunohistochemically examined in the myenteric and submucous plexuses of Korean native goat duodenum. In the neurons of myenteric and submucous plexuses of duodenum, immunoreactivities of SP, CGRP and CB were confirmed in both nerve cell bodies and fibers. In contrast, CR immunoreactivity was found only in nerve fibers of myenteric plexuses, while PA immunoreactivity was found only in nerve cell bodies of submucous plexuses. In the inner circular muscle layer, dense SP-like immunoreactive fibers were prominent but only a few CGRP-like immunoreactivities were observed. Most of SP- and CGRP-like immunoreactive neurons of both plexuses colocalized with CB. This result showed that SP and CGRP may have a important role for the movement of small intestine. The colocalizations of CB with SP or CGRP in myenteric and submucous plexuses suggest that CB may serve neuromodulatory role for SP- and CGRP-immunoreacted neurons on the movement of intestinal wall.

• Clinical and Experimental Pediatrics
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• v.49 no.5
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• pp.533-538
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• 2006

Purpose : The headache, one of the symptoms of meningitis, is related to abrupt elevation of intracranial pressure(ICP) or stimulation of intracranial nociceptive structure. However, in cases of mild elevation of ICP or normal findings of cerebrospinal fluid(CSF) analysis, patients sometimes complain of headaches. Therefore, other pathways may contribute to the occurrence of headaches in aseptic meningitis or meningismus. We intend to investigate the role of substance P(SP) and calcitonin gene-related peptide(CGRP) in aseptic meningitis or meningismus. Methods : We measured leukocyte count, the concentration of protein and glucose in CSF and ICP of patients with meningeal irritation sign. We also measured SP and CGRP levels by using immunoassay. We analyzed the relationship between the presence of headache and the value of SP and CGRP. Results : The concentrations of CGRP($18.8{\pm}10.5ng/mL$) in CSF and ICP($14.8{\pm}4.5cmH_2O$) in aseptic meningitis group were significantly higher than in those($14.1{\pm}7.4ng/mL$ and $12.0{\pm}5.1cmH_2O$, respectively) of the meningismus group(P<0.05). There was no significant difference in the SP levels between the two groups. In the aseptic meningitis group, the concentrations of SP and CGRP were significantly higher in the normal ICP group than in the elevated ICP group(P<0.05). Conclusion : Headaches in children with aseptic meningitis or meningismus is considered to be related to the elevation of the CSF levels of SP and CGRP.

• The Korean Journal of Physiology and Pharmacology
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• v.2 no.5
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• pp.573-580
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• 1998

The study aims to identify the mechanism (s) underlying the altered vasodilatory responses of the pial artery of spontaneously hypertensive rats (SHR) under a hypothesis that calcitonin gene-related peptide (CGRP) exerts a modulator role in the autoregulation of cerebral blood flow (CBF). The animals were divided into four groups: 1) Sprague-Dawley rats (SDR), 2) Wistar rats (WR), 3) SHR with high blood pressure $(BP{\ge}150\;mmHg),$ and 4) SHR with normotensive BP $({\le}150\;mmHg).$ The lower limit of CBF autoregulation in SHR shifted to a higher BP $(82.8{\pm}9.3\'mmHg,\;P<0.05)$ than that in SDR $(58.9{\pm}5.7\;mmHg)$. In SHR, whether the BP levels were high or normotensive, the vasodilator responses to a stepwise hypotension were significantly attenuated unlike with SDR and WR. When artificial cerebrospinal fluid (CSF) containing capsaicin $(3{\times}10^{-7}\;M)$ was suffused over the cortical surface, a transient increase in pial arterial diameter was observed in the SHR with high or normotensive BP. In contrast, SDR and WR showed a large increase in diameter, and the increase was sustained for over 10 minutes. In line with these results, the basal releases of CGRP-like immunoreactivity (CGRP-LI) in the isolated pial arteries from SHR with high and normotensive BP were $12.5{\pm}1.4\;and\;9.8{\pm}2.8\;fmole/mm^2/60\;min\;(P<0.05)$, while those from SDR and WR were $25.5{\pm}3.1\;and\;24.6{\pm}3.1\;fmole/mm^2/60\;min,$ respectively. The isolated basilar arteries showed similar results to those of the pial arteries in SHR. Thus, it is summarized that, in the SHR, the reduced autoregulatory vasodilator responses to stepwise hypotension and capsaicin may be, in part, ascribed to the decreased release of CGRP from the perivascular sensory nerve fibers of the pial arteries, and that altered vasomotor activity in SHR may not be related with the hypertensive tone.

• Biomedical Science Letters
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• v.11 no.3
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• pp.375-382
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• 2005

This study aimed to investigate whether nociceptin is implicated in the, trigeminovascular responses to electrical stimulation of trigeminal ganglion in rats. An open cranial window was prepared on the right parietal bone of male Sprague-Dawley rats. Trigeminovascular system was stimulated by electrical stimulation of trigeminal ganglion (ETS; 5ms, 5Hz, 3V). Neonatal capsaicin treatment was performed with subcutaneous administration of capsaicin (50mg/kg) within the first 24 hours after birth. Changes in regional cerebral blood flow were continuously measured through the cranial window by laser-Doppler flowmetry, and the expression of nociceptin-like immunoreactivity was determined by immunohistochemistry. ETS caused increases in regional blood flow of pial arteriole in a voltage-dependent manner. ETS markedly and voltage-dependently increased the expression of nociceptin-like immunoreactivity in dura mater ipsilateral rather than contralateral to ETS. The nociceptin-like immunoreactivity was markedly reduced by pretreatments with calcitonin gene-related peptide(8-37) ($CGRP_{8-37},\;a\;CGRP_1$ receptor antagonist), L-733060 (a $NK_1$ receptor antagonist), and $[Nphe^1]$ nociceptin(1-13)$NH_2$ (a selective and competitive nociceptin receptor antagonist) as well as by neonatal capsaicin treatment. These results suggest that the electrical stimulation of trigeminal ganglion causes prominent expression of nociceptin within dura mater, in which not only neuropeptides inducing substance P and CGRP but also nociceptin are implicated in the trigeminovascular responses to electrical trigeminal ganglion stimulation.

• Journal of Dental Rehabilitation and Applied Science
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• v.28 no.2
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• pp.171-178
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• 2012

Botulinum toxin type A (BoNT-A) has been applied successfully to treat chronic migraine, dystonia, spasticity and temporomandubular disorders(TMDs) as well as frontal wrinkle and glabella wrinkle. Recently it has been reported that BoNT-A, reversibly blocks presynaptic acetylcholine release, also inhibits the release of substance P, CGRP(calcitonin gene related peptide) and glutamate related to peripheral sensitization and neurogenic inflammation in sensory nerve, In this study we reviewed animal nerve injury model such as rat and rabbit and identify the analgesic effect and mechanism of nerve injury pain after dental treatment.

• Journal of the korean academy of Pediatric Dentistry
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• v.23 no.4
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• pp.775-787
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• 1996

The purpose of this study was to investigate the postnatal development of pulpal innervation with the tooth development and eruption process in developing rat mandibular molars (postnatal 1, 5, 10, 15, 20, 25, 35day-old and adult rats). Immunohistochemical method was carried out for the detection of nerve fibers, using the antibody against calcitonin gene-related peptide(CGRP). The results were as follows: The feature of CGRP-IR nerve fibers were shown in a bead-like appearance. The time of nerve entering into the dental papilla of tooth follicle began at the occured advanced dentinogenesis. The development of Raschkow plexus began at the formative stage of the roots and was accelerated at the stage of the crown emerged into the oral cavity. The number of nerve fibers entering the odontoblastic layer increased with the tooth eruption and mastication. The development of innervation was shown to be related with the stage of the development of individual teeth rather than the chronological age of the rat.

• Biomedical Science Letters
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• v.11 no.3
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• pp.365-373
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• 2005

The aim of this study was to investigate the alterations in meningeal blood flow by stimulation of trigeminovascular system. An open cranial window was prepared on the right parietal bone of male Sprague-Dawley rats. Trigeminovascular system was stimulated by electrical stimulation of trigeminal ganglion (ETS), somatosensory (whisker) stimulation, or topical applications of capsaicin and neuropeptides including substance P and calcitonin gene-related peptide (CGRP). Neonatal capsaicin pretreatment was performed with subcutaneous administration of capsaicin (50 mg/kg) within the first 24 hours after birth. Changes in regional blood flow of dural artery (rDBF) and pial artery (rPBF) were continuously measured through the cranial window by laser-Doppler flowmetry. Both ETS and capsaicin caused a chain of alterations in rPBF and rDBF responses, i.e., an immediate transient decrease followed by rapid and marked increase in rPBF, which were significantly attenuated not only by pretreatments with L-733,060, a $NK_1$ receptor blocker, $CGRP_{8-37}$, a $CGRP_1$ receptor blocker, and 7-nitroindazole monosodium salt (7-NINA), a neuronal nitric oxide synthase inhibitor but also by neonatal capsaicin treatment. Exogenous neuropeptides including substance P and CGRP increased the meningeal blood flow, which was significantly attenuated not only by pretreatment with L-733,060 and $CGRP_{8-37}$, respectively, but also by pretreatment with 7-NINA. The rPBF response to whisker stimulation was significantly attenuated not only by trigeminovascular system injuries including nasociliary nerve denervation and neonatal capsaicin treatment but also by pretreatments with L-733,060, $CGRP_{8-37}$ and 7-NINA. These results suggest that the stimulation of trigeminovascular system causes prominent alterations in meningeal blood flow, and that neuropeptides as well as nitric oxide in the trigeminovascular system are importantly implicated in the regulation of meningeal blood flow.

• Journal of Korean Neurosurgical Society
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• v.59 no.2
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• pp.98-105
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• 2016

Objective : Lumbar disc herniation can induce sciatica by mechanical compression and/or chemical irritation. The aim of this study was to compare the effects of GCSB-5 (Shinbaro$^{(R)}$) and NSAIDs on pain-related behavior and on the expressions of microglia, astrocytes, CGRP, TRPV1, IL-6, and CX3CL1 in a rat model of lumbar disc herniation. Methods : 112 male Sprague-Dawley rats underwent implantation of nucleus pulposus to a dorsal root ganglion (DRG). Rats were divided into five groups as follows; a saline group (the vehicle control group) (n=27), a 10 mg/kg aceclofenac group (the aceclofenac group) (n=22), and 100, 300 or 600 mg/kg GCSB-5 groups (the GCSB-5 100, 300, or 600 groups) (n=21 for each group). Rats were tested for mechanical allodynia at 3 days after surgery and at 1 day, 3 days, 7 days, 14 days, 21 days, 28 days, 35 days, 42 days, 49 days, and 56 days after treatment commencement. Immunohistochemical staining of microglia (Iba1), astrocytes (GFAP), CGRP, and TRPV1, and PCR for IL-6 and CX3CL1 were performed on spinal dorsal horns and DRGs at 56 days after medication commencement. Results : After 56 days of GCSB-5 300 administration, mechanical withdrawal thresholds were significantly increased (p<0.05), and immunohistochemical expressions of Iba1, GFAP, CGRP, and TRPV1 were reduced than other groups, but this difference was not statistically significant. Conclusion : These results indicate GCSB-5 reduces mechanical allodynia and downregulates neuroglial activity and the expressions of CGRP and TRPV1 in the spinal segments of a rat model of lumbar disc herniation.

• Restorative Dentistry and Endodontics
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• v.20 no.2
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• pp.527-537
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• 1995

The effect of calcitonin gene-related peptide (CGRP), substance P (SP) and electrical stimulation of the tooth on the intradental nerve activtiy (INA) was investigated in anesthetized cats. The INA was recorded from single pulp nerve units dissected from the inferior alveolar nerve under stereomicroscope. The INA elicited by 3 minute application of 4M NaCl in deep dentinal cavity was compared before and after stimulation at 10 minute intervals. The magnitude of INA was calculated as the total number of nerve impulses produced in given period, and the changes of INA are expressed as % of control INA. The results obtained were as follows. 1. 16 single pulp nerve units were classified as 14 $A{\delta}$-fibers (3.4~19.4m/sec) and 2-fibers (1.5~1.7m/sec) according to the conduction velocity. 2. 4M NaCl evoked an irregular bursts of spikes which continued until washing out. Isotonic saline did not affect INA to subsequent applications of the hypertonic NaCl solution (P>0.05). 3. Local application of CGRP ($200{\mu}g$/ml) in deep dentinal cavity reduced the INA induced by 4M NaCl in $A{\delta}$-fiber units (P<0.01) and some units of those responded to CGRP during application. 4. Local application of SP ($100{\mu}g$/ml) in deep dentinal cavity reduced the INA induced by 4M NaCl in AS-fiber units (p<0.05), but increased the INA in C-fiber unit coincided with large reduction of the INA of $A{\delta}$-fiber units. 5. Monopolar electrical stimulation applied to the crown at intensities high enough to excite C-fibers (12V, 5ms, 10Hz, 10~30min) decreased the INA in $A{\delta}$-fiber units (P<0.01) and systemic pretreatment with phenoxybenzamine (3mg/kg, i.v.) enhanced this inhibitory effect (P<0.01). On the contrary, electrical stimulation increased the INA in C-fiber unit.