Analysis of kinesiograph recordings and masticatory efficiency after treatment of non-reducing disk displacement of the temporomandibular joint.

The purpose of this study was to clarify the kinesiographs of chewing movement and masticatory efficiency before and after treatment in patients with non-reducing disk displacement of the temporomandibular joint (TMJ). Twenty patients who were diagnosed with unilateral non-reducing disk displacement of the TMJ were treated with pumping of the joint with injection of sodium hyaluronate. Chewing movement patterns in these patients were evaluated, using mandibular kinesiography (MKG) at their initial visit and at mean 19-month follow-up and the results were compared. Masticatory efficiency was also measured. As controls, 23 volunteers without TMJ dysfunction were employed. Far from the results of normal volunteers, chewing movement patterns of the patients on MKG did not show deviation to the chewing side in the TMJ-unaffected-side chewing in the horizontal plane. However, such patterns of the patients became similar to those of normal volunteers after treatment. Masticatory efficiency of the patients improved after treatment, though it was impaired at initial visit. The MKG and masticatory efficiency test appeared to be a useful method of comparing masticatory function before and after treatment in patients with non-reducing disk displacement of the TMJ.

Analysis of post-treatment electromyographs in patients with non-reducing disc displacement of the temporomandibular joint.

The purpose of this study was to clarify the electromyographic property of chewing movement before and after treatment in patients with non-reducing disc displacement of the temporomandibular joint (TMJ). Twenty patients who were diagnosed with unilateral non-reducing disc displacement of the TMJ were treated by pumping and injection of sodium hyaluronate into the joint. Chewing movement in these patients was evaluated by electromyography (EMG) at the initial visit and at mean 19-month follow-up and the results were compared. Chewing movement in 23 normal controls were also examined. Duration of contraction, cycle time and integrated value at the initial visit which were different from those in the controls without TMJ dysfunction tended to be at the control level mean 19 months after treatment of pumping and injection of sodium hyaluronate into the joint in patients with non-reducing disc displacement of the TMJ. Electromyography appeared to be a method of documenting the chewing movement which was impaired at initial visit improved after treatment in patients with non-reducing disc displacement of the TMJ.

Suppression of Egr-1 expression in human oral squamous carcinoma cells by okadaic acid.

We examined the expression of early growth response-1 (Egr-1) gene in human oral squamous carcinoma cell lines SCCKN and SCC-25 cells and human osteoblastic cell lines Saos-2 and MG63 cells treated with okadaic acid, a potent inhibitor of protein phosphatases type 1 and type 2A. Western blot analysis revealed that Egr-1 was strongly expressed in SCCKN cells and that okadaic acid decreased the expression of Egr-1 protein in these cells. However, Egr-1 was expressed at lower levels in SCC-25, Saos-2, and MG63 cells and transiently increased with the okadaic acid treatment. Suppression of Egr-1 protein expression in okadaic acid-treated SCCKN cells stemmed from the suppression of the Egr-1 mRNA level, as determined by the RT-RCR method. Formaldehyde-fixed and alcohol-permeabilized cultured SCCKN cells were reacted with the anti-Egr-1 antibody using immunohistochemical methods. Intense fluorescence was observed in the nuclei of the control SCCKN cells interacted with anti-Egr-1 antibody. However, only a weak reaction was observed in the nuclei in SCCKN cells treated with okadaic acid. A gel mobility shift assay showed that treatment of SCCKN cells with okadaic acid suppressed Egr-1 binding to the DIG-labeled Egr-1 consensus oligonucleotide probe. The present results indicate that the alteration of phosphorylation states in SCCKN cells regulates Egr-1 binding to its consensus sequence and its expression at the transcriptional level.

FRAP-positive and capsaicin-sensitive terminals in the substantia gelatinosa of the mouse spinal trigeminal nucleus caudalis.

FRAP (fluoride-resistant acid phosphatase)-reactivity in the substantia gelatinosa of the mouse spinal trigeminal nucleus caudalis (STNC) was examined by light and electron microscopy. Degenerated figures of terminals caused by capsaicin were compared with the FRAP-positive terminals. Scalloped (fan-like) or indented, sinuous, slender, and cap-like figures with closely packed agranular synaptic vesicles of various sizes were common to both FRAP-positive and capsaicin-sensitive terminals. These terminals had glomerular or nonglomerular endings. Sometimes FRAP-positive and capsaicin-sensitive glomerular terminals made presynapses with surrounding dendrites. Frequently, both nonglomerular terminals were in direct contact with the neuronal soma. The terminal features of FRAP-positive and capsaicin-sensitive ones in the mouse STNC are the same as those seen in the superficial dorsal horn of the spinal cord. These findings suggest that some of the FRAP-positive terminals are capsaicin-sensitive, thereby indicating their nociceptive primary afferent.

Analysis of calcitonin gene-related peptide (CGRP)-containing nerve fibres in the rat spinal cord using light and electron microscopy.

Calcitonin gene-related peptide (CGRP) content in areas surrounding the central canals of cervical, thoracic, lumbar and sacral spinal cords of rats were investigated by light microscopy, conventional transmission electron microscopy (CTEM) (100-200 kV) and high-voltage transmission electron microscopy (HVTEM) (1000 kV) using immunocytochemistry. Tissues were examined using either the peroxidase-antiperoxidase technique or an immuno-cryoultramicrotomy technique. Light microscopically, more intense CGRP localization was observed ventral to the central canals in the lumbar and sacral cord compared with the dorsal area in the same regions. HVTEM revealed that high levels of CGRP labelling were found adjacent to the basal side of ependymal cells ventral to the central canal. Analysis using CTEM operated at 200 kV demonstrated that the CGRP immunoreactivity was present within unmyelinated nerve fibres in the vicinity of the basal side of ependymal cells ventral to the central canal. Immuno-cryoultramicrotomy revealed that immuno-gold particles indicating CGRP labelling were localized on vesicle-like electron-dense bodies in unmyelinated and some fine caliber myelinated nerve fibres. Existence of CGRP as a primary afferent marker in the area surrounding the central canal suggest that there may be an intimate relationship between the modulation of nociceptive information and the area surrounding the central canal in rat spinal cord.

Relationship of substance P- and CGRP-immunoreactive central endings of the primary afferent neurons to GABAergic interneurons in the guinea pig substantia gelatinosa.

The synaptic relationships between primary afferent central endings containing substance P (SP) and calcitonin gene-related peptide (CGRP) and GABAergic interneurons in the guinea pig substantia gelatinosa of the lumbar spinal dorsal horn were studied. The pre-embedding PAP method was used for detection of GABA and the post-embedding double immunogold labeling method for SP and CGRP detection. Immunogold particles specific for SP and CGRP were mainly localized separately or together in large synaptic vesicles devoid of dense cores. SP and CGRP immunoreactivity was separate or co-localized in small roundish, slender, sinuous or large scalloped (fan-like) terminals with closely packed round agranular synaptic vesicles of various sizes and few large dense core vesicles and mitochondria, which are thought to be capsaicin-sensitive primary afferent terminals. These SP- and CGRP-immunoreactive boutons make presynaptic or symmetric contacts with GABAergic dendrites and soma. These findings suggest that the central endings of nociceptive primary afferents transmit pain stimuli to intrinsic inhibitory interneurons, thereby modulating nociceptive information via a postsynaptic circuit.

Fluoride-resistant acid phosphatase (FRAP)-positive afferent terminals make synaptic contact with interneuronal soma in the substantia gelatinosa of the mouse spinal dorsal horn.

Fluoride-resistant acid phosphatase (FRAP)-reactive terminals making contact with interneuronal soma are found in the substantia gelatinosa of the mouse spinal dorsal horn. About one half of the interneuronal somata receive FRAP-positive boutons. By electron microscopy, these FRAP-positive terminals appear small, dark, slender, roundish, cap-like, ellipsoid or sinuous and electron-dense, scalloped (fan-like) contours with clear spherical synaptic vesicles of variable size, some large dense-core vesicles and mitochondria. All these features are very similar to those of capsaicin-sensitive terminals. Thus they are considered to be nociceptive primary afferent endings. Therefore, some of the FRAP-positive terminals are suggested to have a modulatory role in the nociceptive circuit in the substantia gelatinosa.

Evidence of synaptic contacts of nociceptive primary afferent central terminals on GABAergic interneurons in the substantia gelatinosa.

Recently, we showed that capsaicin induced the degeneration of not only glomerular CI terminals but also of non-glomerular CI terminals making presynaptic contact with interneuronal soma. Studies of the nature of interneurons making contact by nonglomerular CI terminals should provide important information to facilitate our understanding of the processing of nociceptive impulses in the substantia gelatinosa. The most likely candidate molecule involved in this process in these interneurons is gamma-aminobutylic acid (GABA). Therefore, ultrastructural relationships between nonglomerular CI terminals land GABAergic interneuronal soma in the mouse substatia gelatinosa were examined by an immunocytochemical method using an antibody to GABA. Terminals with the same profiles as the CI terminals, i.e., slender, sinuous and scalloped terminals filled with clear synaptic vesicles, were found to make synaptic contacts with GABA-immunoreactive somata. Thus, nociceptive primary afferents are suggested to modulate pain transmission by themselves via GABAergic neurons in the substantia gelatinosa.

The confronting cisternae in the hippocampal neurons of rats and Japanese monkeys.

The ultrastructure of the neurons in the hippocampal formation of Wistar rats and Japanese monkeys was investigated. Confronting cisternae were observed in the cytoplasm of pyramidal neurons and granule cells. The structures were composed of closely apposed parallel cisternae, presumably continuous with the cisternae of the rough endoplasmic reticulum. The cytoplasmic matrix between the confronting cisternae contained electron dense flocculent material. In the pyramidal neurons irregular confronting cisternae were also observed. Possible functions of the confronting cisternae are discussed.

Fetal alcohol effects: decreased synaptic formations in the field CA3 of fetal hippocampus.

The effects of prenatal ethanol exposure on the synaptic formation in the field CA3 of the hippocampus of fetal rats have been investigated on gestational day 21. Significantly decreased number of synaptic junctions was observed in the fetus showing decreasing cerebral weight either with or without decreasing body weight. The administration of 0.01% zinc with ethanol or 0.02% alpha-tocopherol acetate with ethanol during pregnancy resulted in an increased cerebral weight, but did not result in an increased synaptic formation compared to ethanol alone. This result indicates that one of the most vulnerable factors in rat fetus exposed to ethanol in utero is the synaptic formation in the hippocampus. In conclusion, ethanol exposure in utero during a period of brain development roughly equivalent to the first and second human trimesters can produce consistent dysforming effect of synapses and may be associated with the functional impairement of the central nervous system in the fetal alcohol effects.

Stereoscopic observation of enzyme distribution in lowicryl K4M-embedded specimens by conventional electron microscopy.

The present investigation was undertaken to explore the value of the Lowicryl K4M embedding technique for enzyme histochemical examination of semi-thin sections. The low-temperature embedding procedure with Lowicryl K4M was found to provide favorable conditions for preservation of enzyme activity in tissue samples. We tested the histological effects of various fixatives; the best results were obtained using 4% paraformaldehyde when testing for AcPase, AlPase, TPPase, and Mg-ATPase in the dorsal root ganglion. The three-dimensional cellular fine structure could be clearly seen in stereo pair pictures under stereoscopy.

Ultracytochemical demonstration of Ca2(+)-ATPase activity in the rat saphenous artery and its innervated nerve terminal.

The localization of Ca2(+)-ATPase activity was ultracytochemically investigated in the rat saphenous artery and nerve terminals innervating the saphenous artery using a lead citrate method devised by Ando et al. (1981). Intense reaction products in the saphenous arterial endothelial cells were observed inside the caveolae and vesicles along the luminal and abluminal sides. In addition, Ca2(+)-ATPase activity was observed on the external side of the luminal, abluminal and lateral plasma membrane, and the outer membrane of mitochondria. In the smooth muscle cells, intense Ca2(+)-ATPase activity on the inside of caveolae and vesicles was observed, comparing in intensity with that on the plasma membrane of smooth muscle cells. In the nerve terminals innervating the saphenous artery, Ca2(+)-ATPase activity was demonstrated on the plasma membrane of the nerve terminal-Schwann cell interface, the axolemma of unmyelinated axons and the plasma membrane of Schwann cells. It is suggested from the above ultracytochemical results that Ca2(+)-ATPase activity plays an important role in the contraction and relaxation of the saphenous artery, and in the neurotransmitter release.

Abnormal movements in brindled mutant mouse heterozygotes: as related to the development of their offspring--biochemical and morphological studies.

As a possible preventive measure for brain dysfunction in Menkes disease, prenatal treatment by maternal administration of zinc, vitamin E and copper was examined in brindled mutant mice. During pregnancy and lactation, female heterozygous mice received 20 ppm zinc or 0.004% alpha-tocopherol acetate (vitamin E) throughout and 6 ppm copper from gestational day 13 in the drinking fluid, ad libitum. The maternal administration of zinc and vitamin E, as antioxidants, or copper resulted in decreased fetal and neonatal death of offspring, especially those of hemizygous males, as compared with the administration of water only. When offspring did not grow, maternal abnormal movements, which comprised rotatory movements of high speed with tremor and ataxia, were frequently observed. In the heterozygotes with abnormal movements, the level of lipid peroxidation in cerebrum and the concentration of copper in kidney were much higher than those in the heterozygotes with normal movement. Morphologically, in cerebellum of the heterozygotes with abnormal movements, the loss of Purkinje cells, abundance of lipofuscin granules and abnormal mitochondria or degenerative bodies of high electron density were frequently observed, as compared with heterozygotes with normal movement. These findings suggest that the development of hemizygous male mice may be influenced by both copper and oxygen radical metabolism.

[Mg++-ATPase activity in circumventricular capillaries during fetal development of rats].

We performed an ultracytochemical study of Mg++-ATPase as a marker of maturation in circumventricular capillaries at the developmental stage of rats, from 19 gestational days to 18 postnatal day, comparing with hippocampal capillaries. During the course of perinatal development, the predominant site of the Mg++-ATPase activity in circumventricular capillaries was shifted from the luminal cell membrane to the antiluminal cell membrane and the total enzyme activity was markedly increased. In all hippocampal capillaries observed, the predominant site of the Mg++-ATPase activity was the antiluminal cell membrane. It was suggested that immaturity of circumventricular capillaries may be one of the causative factors leading to intraventricular and subependymal hemorrhages in neonates.

High-voltage electron microscopy of E-PTA-stained synaptic junctions in the rat frontal cortex.

The E-PTA-stained synaptic junctions in the adult rat frontal cortex were examined with high-voltage electron microscopy (HVEM). Perforated whole synaptic junctions were clearly shown in the stereo image. The E-PTA staining procedure provides a useful marker for studies of the 3-dimensional structure of synaptic junctions by means of HVEM.

The effect of cortisol on the plastic synapses in the developing mouse brain.

We studied the effect of cortisol on the plastic synapses in the hippocampal region of the developing mouse brain. Ethanolic phosphotungustic acid-stained synaptic junctions were reduced in the cortisol-treated mouse brain dose-dependently and irreversibly. The steroid therapy in infancy requires circumspection.

Qualitative morphogenesis of synaptic glomeruli in the rat cerebellum.

The synaptic glomeruli of rat cerebellum was investigated at postnatal ages of 1 to 7, 9 and 15 days in an attempt to formulate a morphogenic analysis for synaptic junctional development, using the E-PTA stain for synapses. The synaptic junctions, which are composed of dense projection, intercleft density and postsynaptic band in synaptic glomeruli, were observed at a postnatal age of 6 days. Two types of synaptic junctions, symmetrical and asymmetrical, were recognized.

The effect of ACTH on plastic synapses in the developing mouse brain.

We studied the effect of ACTH on plastic synapses in the hippocampal gyrus of the developing mouse brain. Examination of ethanolic phosphotungustic acid stained synaptic junctions revealed no obvious differences between the ACTH-treated brains and controls qualitatively and quantitatively. Therefore, ACTH may not adversely affect the plastic synapses in the developing mouse brain.

The fine structure of the node of Ranvier in the rat cerebellar cortex.

The transitional zone between the myelinated and the terminal portions of nerve fibers was electron microscopically investigated in the cerebellar cortex of normal rats. A noteworthy finding in the present study is the existence of a "heminode" consisting of the myelinated and non-myelinated portions, as demonstrated within a single section. The heminode seems to be a characteristic structure in the granular layer of the rat cerebellar cortex.