In Vivo Ca2+ Imaging of the Insular Cortex during Experimental Tooth Movement.

Pain and discomfort are common problems for patients undergoing orthodontic treatment. We have demonstrated that cortical excitation propagation in the somatosensory and insular cortices (IC) induced by electrical stimulation of the periodontal ligament (PDL) is facilitated 1 d after experimental tooth movement (ETM). However, it is necessary to examine ETM-induced changes in neural responses at a single-cell level to understand the mechanisms of cortical plastic changes, in which excitatory glutamatergic and inhibitory GABAergic neurons are intermingled to form cortical local circuits. We performed in vivo 2-photon Ca2+ imaging by loading the Ca2+ indicator Oregon Green BAPTA with the astrocyte marker sulforhodamine. We focused on the IC region that exhibited the largest neural response to maxillary PDL (mxPDL) stimulation using a VGAT-Venus transgenic rat that expresses venus fluorescent protein in GABAergic neurons and discerned changes in the neural activities of each cortical neuronal subtype before and during ETM treatment of the maxillary incisor and first molar. Notably, 1 d after ETM treatment (1d-ETM), the number of neurons responding to mxPDL stimulation increased from 47.6% to 64.2% in excitatory neurons and from 44.5% to 66.2% in inhibitory neurons. On the other hand, only 3% to 4% of excitatory and inhibitory neurons responded to mandibular molar PDL (mbPDL) stimulation in control rats, and the 1d-ETM group showed significant increases in excitatory (14.0%) and inhibitory neurons (22.5%) responding to mbPDL stimulation. Interestingly, most mbPDL-responding neurons also responded to mxPDL stimulation. The population of excitatory and inhibitory neurons that responded only to mxPDL stimulation was comparable between the control and 1d-ETM groups. The facilitative responses in the 1d-ETM group had almost recovered 7 d after ETM treatment. These results suggest that ETM induces parallel increases in PDL-responding neurons and changes some insensitive neurons to respond to both mxPDL and mbPDL stimulation.

Early growth-responsive-1-dependent manganese superoxide dismutase gene transcription mediated by platelet-derived growth factor.

Manganese superoxide dismutase Mn-SOD plays a major role in protecting mitochondria from oxidative damage. Overexpression of Mn-SOD maintains cell survival under conditions that lead to apoptotic death. In addition to the antioxidative enzyme, platelet-derived growth factor (PDGF) is a principal survival factor that inhibits apoptosis and promotes proliferation by activating survival signaling pathways in various cells. Here we show that PDGF induced the expression of the Mn-SOD gene in NIH3T3 cells, and its induction was associated with early growth response-1 (Egr-1), a transcription factor. An electrophoretic mobility shift assay demonstrated that Egr-1 bound to the proximal promoter of the Mn-SOD gene in response to PDGF. The proximal promoter region of Mn-SOD was shown to be transcriptionally responsive to both basal and PDGF stimulation by transfection studies. Forced expression of Egr-1 in the cells activated Mn-SOD transcription in a dose-dependent manner. The pathway by which PDGF induced Egr-1 involved the mitogen-activated protein kinase kinase-1 (MEK1) and extracellular signal-regulated kinases 1 and 2 (ERK1/2), because the effect of PDGF on the induction of Egr-1 was blocked by U0126, a specific MEK1 inhibitor. These findings indicate that the induction of Mn-SOD is part of the anti-apoptotic properties mediated by PDGF.

[The role of transplanted astrocytes for the regeneration of CNS axons].

Long tract axons in the mammalian CNS do not normally regenerate for appreciable distance after they transected. But we reported transplantation of Schwann cells(SCs) or olfactory ensheathing cells induced regeneration of transected rat dorsal column (DC) axons and improved the conduction. Scar formation(gliosis), for which astrocytes(ACs) play an important role, may be one of strong and physical barriers for the regeneration of CNS axon. Oligodendrocyte and myelin associated protein or products also inhibit the regeneration of the axons, as chemical barriers. To investigate how effective the promotion or the reduction of scar or myelin formation may be for axonal regeneration, we transplanted AC into transected DCs, or radiated(X-ray) the DCs, and compared to normal DCs or regenerated DCs following by SC transplantation. DCs of adult rats were transected at Th 11 and transplanted with SCs(6 x 10(4)) of adult rats or ACs(6 x 10(4)) of neonatal rats. Five to six weeks later, the spinal cords were removed and pinned in a recording chamber, and compound action potentials (CAPs) along the DC through the transected lesion were recorded, to investigate conduction properties(conduction velocity and response after high frequency stimulations). Following transplantation of SCs or ACs, histological examination revealed regenerated axons with SC-like patterns of remyelination in transected DCs. X-ray irradiation did not enhance the regeneration of DC axons. SC transplantation improved the conduction properties of transected DCs and increased the number of regenerated axons, compared to transected DCs without cell transplantation. AC transplantation resulted in improvement of the conduction properties, but the number of regenerated axons was similar to that of transected DCs without the transplantation. X-ray irradiation (40 Gy) three days before DC transection and AC transplantation prevented the electrophysiological continuity of axons through the transected lesion. This evidence revealed that AC transplantation secondarily enhanced the regeneration of axons, probably endogeneous SCs of dorsal roots migrated into the transected lesion and enhanced the axonal regeneration.

Transplantation of an acutely isolated bone marrow fraction repairs demyelinated adult rat spinal cord axons.

The potential of bone marrow cells to differentiate into myelin-forming cells and to repair the demyelinated rat spinal cord in vivo was studied using cell transplantation techniques. The dorsal funiculus of the spinal cord was demyelinated by x-irradiation treatment, followed by microinjection of ethidium bromide. Suspensions of a bone marrow cell fraction acutely isolated from femoral bones in LacZ transgenic mice were prepared by centrifugation on a density gradient (Ficoll-Paque) to remove erythrocytes, platelets, and debris. The isolated cell fraction contained hematopoietic and nonhematopoietic stem and precursor cells and lymphocytes. The cells were transplanted into the demyelinated dorsal column lesions of immunosuppressed rats. An intense blue beta-galactosidase reaction was observed in the transplantation zone. The genetically labeled bone marrow cells remyelinated the spinal cord with predominately a peripheral pattern of myelination reminiscent of Schwann cell myelination. Transplantation of CD34(+) hematopoietic stem cells survived in the lesion, but did not form myelin. These results indicate that bone marrow cells can differentiate in vivo into myelin-forming cells and repair demyelinated CNS.

Transplantation of clonal neural precursor cells derived from adult human brain establishes functional peripheral myelin in the rat spinal cord.

We examined the myelin repair potential of transplanted neural precursor cells derived from the adult human brain from tissue removed during surgery. Sections of removed brain indicated that nestin-positive cells were found predominantly in the subventricular zone around the anterior horns of the lateral ventricle and in the dentate nucleus. Neurospheres were established and the nestin-positive cells were clonally expanded in EGF and bFGF. Upon mitogen withdrawal in vitro, the cells differentiated into neuron- and glia-like cells as distinguished by antigenic profiles; the majority of cells in culture showed neuronal and astrocytic properties with a small number of cells showing properties of oligodendrocytes and Schwann cells. When transplanted into the demyelinated adult rat spinal cord immediately upon mitogen withdrawal, the cells elicited extensive remyelination with a peripheral myelin pattern similar to Schwann cell myelination characterized by large cytoplasmic and nuclear regions, a basement membrane, and P0 immunoreactivity. The remyelinated axons conducted impulses at near normal conduction velocities. This suggests that a common neural progenitor cell for CNS and PNS previously described for embryonic neuroepithelial cells may be present in the adult human brain and that transplantation of these cells into the demyelinated spinal cord results in functional remyelination.

Peroxynitrite induces GADD34, 45, and 153 VIA p38 MAPK in human neuroblastoma SH-SY5Y cells.

Peroxynitrite, one of the most reactive radicals, is produced from superoxide anion and nitric oxide. A peroxynitrite generator, 3-morpholinosydonimine (SIN-1), was found to induce the expression of three different growth arrest and DNA damage-inducible (GADD) mRNA, GADD34, GADD45, and GADD153, at the early phase during cell death in human neuroblastoma SH-SY5Y cells. In addition, peroxynitrite activated p38 MAPK just before induction of three GADD mRNA. A specific inhibitor of p38 MAPK, SB202190, markedly suppressed peroxynitrite-induced expression of three GADD mRNA in SH-SY5Y cells. The expression of three GADD genes and also p38 MAPK phosphorylation were suppressed by treatment with radical scavengers, superoxide dismutase plus catalase and glutathione. Glutathione depletion by L-buthionine-S, R-sulfoximine (BSO), increased the vulnerability of the cells to peroxynitrite. These findings indicate that peroxynitrite-mediated oxidative stress activated p38 MAPK to induce three GADD genes.

Peroxynitrite induces neuronal cell death in aging and age-associated disorders: A review.

Peroxynitrite produced from nitric oxide and superoxide has been proposed to cause neuronal dysfunction and cell death in aging and age-related degenerative diseases. 3-Nitrotyosine, an oxidation product of tyrosine by peroxynitrite, was reported to increase in degenerating brains. In this paper, involvement of peroxynitrite in neuronal cell death was studied by analyses of human brains and in vitro experiments on cell death induced by a peroxynitrite-generating agent, SIN-1. 3-Nitrotyrosine-containing proteins were detected in lipofuscin, a typical aging-related pigment in human brains. The cytotoxicity of peroxynitrite was examined in human dopaminergic SH-SY5Y cells by use of SIN-1. SIN-1 induced apoptotic cell death in the cells, and increased the level of 3-nitrotyrosine-containing proteins. The intracellular transduction of death signal was studied in apoptosis induced by peroxynitrite. Apoptosis was induced by sequential death cascade, collapse of mitochondrial membrane potential, activation of caspases and fragmentation of nuclear DNA. In addition, phosphorylation of p38 mitogen activated phosphokinase (MAPK) was found to be associated with apoptosis by SIN-1, as shown by inhibition of apoptotic process by SB202190, a p38 inhibitor. Involvement of peroxynitrite in the cell death is discussed in relation to neuronal degeneration in aging and age-associated diseases.

[Remyelination by Schwann cell transplantation for CNS demyelinated axons: functional comparison with developmental myelination].

Transplantation of Schwann cells(SCs) induced remyelination of demyelinated rat dorsal column(DC) axons and improved conduction. To investigate the difference between developmental oligodendrocytic myelination and SC myelination in conductive functions of axons, we compared normally developmental DCs(10 days old, 22-23 days old, and adult rat), demyelinated DCs, and remyelinated DCs by SC transplantation. DCs of adult rats were demyelinated at T 11 by X-ray irradiation and ethidium bromide, and transplanted with SCs(3 x 10(4)) of adult rats. Three weeks later, the spinal cord was removed and pinned in a recording chamber and compound action potentials (CAPs) were recorded, to investigate conduction properties(conduction velocity, amplitude of CAP and response after high frequency stimulation). Normal DCs were recorded in same manner. Following transplantation of SCs, histological examination revealed SC-like patterns of remyelination of demyelinated axons. The conduction velocities of 10 days old DCs(1.7 +/- 3.4 m/s, n = 4) increased early to 8.1 +/- 3.3 m/s(22-23 days old, n = 7) and 12.2 +/- 1.5 m/s (adult, n = 5). After SC transplantation the velocity significantly improved 7.7 +/- 1.5 m/s(n = 5) compared to demyelinated axons(1.2 +/- 0.4 m/s, n = 7), but less than adult. A 600 Hz 0.5 sec stimulus train led to an amplitude decrement of 7.1 +/- 7.5%(n = 7) in demyelinated axons. Following transplantation, amplitude decreased in 66.2 +/- 11.9%(SC, n = 5), same as 22-23 days(71.7 +/- 11.7%, n = 7) old or adult(76.6 +/- 15.6%, n = 6). The recovery properties after high frequency stimulation developed earlier than conduction velocity. Following SC transplantation, the recovery properties improved to that of normal adult, but not conduction velocity. Lower conduction velocity may be due to shorter internode distance of remyelination after transplantation, same as 22-23 days old pups, than normal adult rats. Though anatomical difference and/or time after transplantation influenced the conduction, these result suggested SC remyelination might result in insufficient for conduction velocity.

[Characteristic improvement of the function following Schwann cell transplantation for demyelinated spinal cord].

Transplantation of Schwann cells (SCs) induced remyelination of demyelinated rat dorsal column (DC) axons and improved conduction. To investigate the difference between oligodendrocyte (OL) and SC myelination in conductive functions of axons, we compared normal DCs, demyelinated DCs, demyelinated DCs remyelinated by SC transplantation, and normal dorsal roots. All of the axons was originated from dorsal root ganglion neurons. Dorsal roots of adult rats were demyelinated at T11 by X-ray irradiation and ethidium bromide, and transplanted with SCs (3 x 10(4)) of adult rats. Three weeks later, the spinal cord was removed and pinned in a recording chamber and compound action potentials (CAPs) were recorded, to investigate conduction properties (conduction velocity and response after high frequency stimulation). Normal DCs or dorsal roots were recorded in same manner. Following transplantation of SCs, histological examination revealed SC-like patterns of remyelination in demyelinated DCs. SC transplantation improved significantly conduction properties compared to demyelinated axons, but less than normal DC. Moreover, remyelinated axons by SC transplantation showed as low amplitude of CAP as dorsal roots, but lower conduction velocity than dorsal roots. Though anatomical difference and/or time after transplantation influenced the conduction, these result suggested that SC myelination resulted in lower amplitude of CAP than OL, and SC remyelination might be insufficient for conduction velocity.

[Comparison of myelin-forming cells as candidates for therapeutic transplantation in demyelinated CNS axons].

Demyelination of axons resulted in distinct reduction of conduction velocity or block of conduction. Remyelination by transplantation of myelin-forming cells may provide a therapeutic approach for demyelinated diseases. However, which cell type will be the most appropriate candidate for such a cell therapy is not established. To investigate how effective grafted neonatal brain cell (BC) (including oligodendrocyte and astrocyte) isolated from neonatal fronto-temporal lobes, adult olfactory ensheathing cell (OEC) or adult Schwann cell (SC) may be for demyelinated CNS axons in vivo, dorsal columns(DCs) of adult rat spinal cord were demyelinated at Th 11 by X-ray irradiation (day 0) and the injection of ethidium bromide (day 3), and transplanted 5 x 10(4) of BCs, 3 x 10(4) of OECs, or 3 x 10(4) of SCs into the lesion (day 6). Day 28-31, spinal cord were removed and transferred an in vitro recording chamber to record field potentials using glass micropipettes, to investigate conduction properties at 36 degrees. Normal DCs were recorded in same manner. Histological examination revealed that OECs and SCs resulted in substantial SC-like patterns of remyelination to equal degree, BC transplantation resulted in less myelination. The conduction velocities were significantly improved to 4.2 +/- 2.4 m/s(BC, n = 5), 8.5 +/- 3.3 m/s(OEC, n = 6) and 7.7 +/- 1.5 m/s(SC, n = 5), compared to demyelinated axons(1.2 +/- 0.4 m/s, n = 7). A 600 Hz 0.5 sec stimulus train led to an amplitude decrement of 7.1 +/- 7.5% (n = 7) in demyelinated axons. Following transplantation, the amplitude decreased in 31.3 +/- 18.7% (BC, n = 5), 49.9 +/- 19.9% (OEC, n = 6) and 66.2 +/- 11.9% (SC, n = 5). Transplanted OECs and SCs enhanced the remyelination of demyelinated CNS axons, and improved conduction properties were similar, and more effective than that induced from isolated CNS tissue which included oligodendrocyte.

Transplantation of human olfactory ensheathing cells elicits remyelination of demyelinated rat spinal cord.

Human olfactory ensheathing cells (OECs) were prepared from adult human olfactory nerves, which were removed during surgery for frontal base tumors, and were transplanted into the demyelinated spinal cord of immunosuppressed adult rats. Extensive remyelination was observed in the lesion site: In situ hybridization using a human DNA probe (COT-1) indicated a similar number of COT-1-positive cells and OEC nuclei within the repaired lesion. The myelination was of a peripheral type with large nuclei and cytoplasmic regions surrounding the axons, characteristic of Schwann cell and OEC remyelination. These results provide evidence that adult human OECs are able to produce Schwann cell-like myelin sheaths around demyelinated axons in the adult mammalian CNS in vivo.

Excitability changes of dorsal root axons following nerve injury: implications for injury-induced changes in axonal Na(+) channels.

Electrophysiological recordings were obtained from rat dorsal roots in a sucrose gap chamber to study changes in Na(+) currents following nerve injury. Application of 4-aminopyridine unmasks a prominent and well-characterized depolarization (delayed depolarization) following the action potential. In our previous studies, this potential, which is only present in cutaneous afferent axons, has been shown to correlate with activation of a slow Na(+) current. The delayed depolarization in the dorsal root was reduced 1 week after sciatic nerve ligation, suggesting a reduction in the kinetically slow Na(+) currents on dorsal root axons [control: 44. 2+/-7.3% (n=5); injury: 7.3+/-4.7% (n=5), P<0.001]. The refractory period of the action potential was reduced following nerve injury, in agreement with biophysical studies indicating faster "repriming" of fast Na(+) currents on cutaneous afferent cell bodies. Dorsal root ligation near the spinal cord also results in a reduction in the delayed depolarization. These results indicate that changes in Na(+) channel organization occur on dorsal root axons following either central or peripheral target disconnection, suggesting trophic support can be derived from either the CNS or the PNS.

[Diagnosis of brain tumor with proton MR spectroscopy--the quantification of gliomas compared with normal brain].

We conducted quantification of proton MR spectroscopy (MRS) using water signal as an internal standard, in order to quantify the concentrations of metabolites in normal adult brains and in gliomas in vivo. Single-voxel spectra were acquired using a point-resolved spectroscopic (PRESS) pulse sequence as part of the Probe. P spectroscopy package on a GE Signa Horizon Hispeed LX1.5T scanner (TR/TE/Ave = 3000 msec/30 msec/64). The volume of interest (VOI) varied from 15.0 x 15.0 x 15.0 mm3 to 20.0 x 20.0 x 20.0 mm3 for the brain. The present study included 26 healthy volunteers and 12 patients with gliomas, whose diagnoses were verified by histologic examination. The calculated concentrations of N-acetyl-aspartate (NAA), creatine (Cre) and choline (Cho) in normal hemispheric white matter were 23.66 +/- 1.94 mM (mean +/- SD), 12.97 +/- 1.44 mM, and 4.38 +/- 0.60 mM, respectively. We found they were not necessarily uniform in different parts of the brain, for example, in the pons and basal ganglia. The concentrations of NAA and Cre decreased in all gliomas (p < 0.001). Cho concentration also decreased in the glioma (p < 0.005). The NAA/Cre, NAA/Cho, and Cre/Cho ratios can distinguish normal brain from gliomas, and NAA/Cho ratio can distinguish low-grade astrocytoma from the high-grade group. The results indicate that this noninvasive method offers reasonable estimation of metabolite concentrations in the brain in vivo and therefore is useful in diagnoses of gliomas.

432 Somali women's birth experiences in Canada after earlier female genital mutilation.

BACKGROUND: Women with previous female genital mutilation (sometimes referred to as circumcision) are migrating, with increasing frequency, to countries where this practice is uncommon. Many health care professionals in these countries lack experience in assisting women with female genital mutilation during pregnancy and birth, and they are usually untrained in this aspect of care. Somali women who customarily practice the most extensive form of female mutilation, who were resident in Ontario and had recently given birth to a baby in Canada, were surveyed to explore their perceptions of perinatal care and their earlier genital mutilation experiences. METHOD: Interviews of 432 Somali women with previous female genital mutilation, who had given birth to a baby in Canada in the past five years, were conducted at their homes by a Somali woman interviewer. RESULTS: Findings suggested that women's needs are not always adequately met during their pregnancy and birth care. Women reported unhappiness with both clinical practice and quality of care. CONCLUSIONS: Changes in clinical obstetric practice are necessary to incorporate women's perceptions and needs, to use fewer interventions, and to demonstrate greater sensitivity for cross-cultural practices and more respectful treatment than is currently available in the present system of care.

[A case of tentorial meningioma presented with pure word deafness].

It has been known that an isolation of Wernicke's area from auditory input results in pure word deafness. In this report, a 73-year-old female case with tentorial meningioma suffering from pure word deafness is reported. The patient initially presented with hydrocephalus, and was treated with a ventriculo-peritoneal(V-P) shunt. A year after the V-P shunt, she suffered from a symptom of deafness. On admission, her repetition and auditory comprehension were severely impaired, while reading and visual comprehension were almost normal. Auditory brain stem response(ABR) revealed normal latency between wave I and V, while wave VI and VII was disappeared. Middle latency response(MLR) showed no wave peak. On MRI, tentorial meningioma compressed bilateral medial geniculate bodies, but not auditory radiation or temporal lobe. 99mTc-HMPAO single photon emission computed tomography(SPECT) showed hypoperfusion in the left temporal lobe, considered as a diaschisis resulting from the isolation of left temporal lobe from auditory input via bilateral medial geniculate bodies.

Mitogen-activated protein kinase pathway mediates peroxynitrite-induced apoptosis in human dopaminergic neuroblastoma SH-SY5Y cells.

Peroxynitrite, a product of nitric oxide and superoxide, is one of the most potent oxidants and it has been suggested to be involved in many neurodegenerative disorders. The mechanism of the cytotoxicity by peroxynitrite was examined using 3-morpholinosydonimine (SIN-1) as a peroxynitrite donor and SH-SY5Y cells as a model of dopamine neurons. SIN-1 was found to induce apoptotic cell death with typical nucleosomal DNA fragmentation with activation of caspase 3-like proteases. The signal transduction of apoptosis was studied in concern to mitogen-activated protein kinases (MAPKs). After SIN-1 treatment, phosphorylation of p38 was detected, followed by that of Erk. SB202190, an inhibitor of p38, suppressed Erk phosphorylation to the basal level and partially reduced the activation of caspase 3-like proteases and also the cell death. These results suggest that peroxynitrite may activate p38 MAPK pathway to induce apoptosis in dopamine cells via activation of caspase 3-like proteases.

Family history of subarachnoid hemorrhage and the incidence of asymptomatic, unruptured cerebral aneurysms.

OBJECT: Previously the authors reported a significant correlation between a family history of subarachnoid hemorrhage (SAH) and the discovery of an unruptured aneurysm in a group of healthy volunteers. This study corroborates and extends previous findings regarding the relationship between genetic and acquired factors in the formation of cerebral aneurysms. METHODS: The incidence of asymptomatic, unruptured cerebral aneurysms was studied among patients with a family history of SAH within the second degree of consanguinity. Forty-one unruptured cerebral aneurysms were found in 34 (13.9%) of 244 patients. This incidence was significantly higher than that found in a control group of healthy volunteers (6%). Furthermore, patients who had a family history of SAH combined with multiple systemic risk factors were found to have the highest incidence of unruptured aneurysms (32%; odds ratio 3.49, 95% confidence interval 1.37-8.9). CONCLUSIONS: These findings suggest that patients with a family history of SAH with or without the presence of more than one systemic risk factor are at significantly higher risk of harboring cerebral aneurysms. This high-risk group should be periodically screened and treated with appropriate surgical or other forms of therapy when necessary.

[Usefulness of galea suturing method for scalp closure].

A method for scalp closure to prevent alopecia along a suture line is described. Only the galea is sutured. By tacking a sufficient width of the galea on both sides with an absorbable suture material, the sutured wound forms a ridge. The outer layer is then closed with skin staples to keep the blood circulation undisturbed. This procedure contrasts with the traditional method in which the galea is sutured with the overlying subcutaneous tissue and consequently the hair follicles are strangled. By adopting the method of suturing the galea, development of alopecia along a suture line has been effectively prevented and scarring has become less conspicuous.