Fluoxetine reduces L-DOPA-derived extracellular DA in the 6-OHDA-lesioned rat striatum.

We investigated the effect of fluoxetine, a selective serotonin reuptake inhibitor (SSRI), on L-DOPA-derived extracellular dopamine (DA) levels in the striatum of rats with nigrostriatal dopaminergic denervation using in vivo microdialysis. Treatment with fluoxetine (10 mg/kg, i.p.) induced a 41% reduction in the cumulative amount of extracellular DA during 300 min following L-DOPA administration (50 mg/kg, i.p.; p < 0.01). This effect was antagonized by pretreatment with WAY-100635, a potent 5-HT1A antagonist, indicating that this effect of fluoxetine is due to its indirect 5-HT1A agonistic property. These results suggest that SSRIs may impair motor functions in patients with Parkinson's disease by reducing efflux of exogenous L-DOPA-derived DA.

Activation of 5-HT(1A) but not 5-HT(1B) receptors attenuates an increase in extracellular dopamine derived from exogenously administered L-DOPA in the striatum with nigrostriatal denervation.

In order to determine whether L-DOPA-derived extracellular dopamine (DA) in the striatum with dopaminergic denervation is affected by activation of serotonin autoreceptors (5-HT(1A) and 5-HT(1B) receptors), we applied in vivo brain microdialysis technique to 6-hydroxydopamine-lesioned rats and examined the effects of the selective 5-HT(1A) receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) and the selective 5-HT(1B) receptor agonist CGS-12066 A on L-DOPA-derived extracellular DA levels. Single L-DOPA injection (50 mg/kg i.p.) caused a rapid increase and a following decrease of extracellular DA, with a peak value at 100 min after L-DOPA injection. Pretreatment with both 0.3 mg/kg and 1 mg/kg 8-OH-DPAT (i.p.) significantly attenuated an increase in L-DOPA-derived extracellular DA and the times of peak DA levels were prolonged to 150 min and 225 min after L-DOPA injection, respectively. These 8-OH-DPAT-induced changes in L-DOPA-derived extracellular DA were antagonized by further pretreatment with WAY-100635, a selective 5-HT(1A) antagonist. In contrast, intrastriatal perfusion with the 5-HT(1B) agonist CGS-12066 A (10 nM and 100 nM) did not induce any changes in L-DOPA-derived extracellular DA. Thus, stimulation of 5-HT(1A) but not 5-HT(1B) receptors attenuated an increase in extracellular DA derived from exogenous L-DOPA. These results support the hypothesis that serotonergic neurons are primarily responsible for the storage and release of DA derived from exogenous L-DOPA in the absence of dopaminergic neurons.

Reserpine pretreatment prevents increases in extracellular striatal dopamine following L-DOPA administration in rats with nigrostriatal denervation.

The influence of L-DOPA and reserpine on extracellular dopamine (DA) levels in the striatum of intact and dopaminergic denervated rats was studied using the brain microdialysis technique. In intact rats, reserpine (5 mg/kg s.c.) reduced extracellular DA levels to 4% of basal values. L-DOPA (50 mg/kg i.p.) had no effect on extracellular DA levels in reserpine-pretreated rats. In rats with 6-hydroxydopamine-induced lesion of the nigrostriatal dopaminergic system, basal levels of extracellular DA were low but markedly increased by L-DOPA (50 mg/kg i.p.). In 6-hydroxydopamine-lesioned rats, pretreatment with reserpine (5 mg/kg s.c.) diminished L-DOPA (50 mg/kg i.p.)-induced increases in extracellular DA levels to 16% of those obtained in denervated animals not pretreated with reserpine (p<0.01). These results suggest that in the intact striatum, extracellular DA stems mainly from vesicular storage sites and that in the striatum with dopaminergic denervation, a large part of the L-DOPA-derived extracellular DA is also derived from a vesicular pool that is released by an exocytosis mechanism.

Role of serotonergic neurons in L-DOPA-derived extracellular dopamine in the striatum of 6-OHDA-lesioned rats.

The effect of L-dihydroxyphenylalanine (L-DOPA) on extracellular dopamine (DA) in the striatum was determined by microdialysis in 6-hydroxydopamine (6-OHDA)-lesioned rats treated with and without the serotonergic neurotoxin 5,7-dihydroxytryptamine (5,7-DHT). At the same time the intensity of L-DOPA-induced rotational behavior was assessed. In 6-OHDA-lesioned rats treated with 5,7-DHT, L-DOPA (50 mg/kg, i.p.) increased extracellular DA only to 20% of that measured in animals not treated with 5,7-DHT. Likewise, 6-OHDA-lesioned rats treated with 5,7-DHT exhibited a significantly lower number of L-DOPA-induced rotations. These results suggest that serotonergic terminals in the striatum can convert exogenously administered L-DOPA into DA that can be released into the extracellular space.

Loss of regulation by presynaptic dopamine D2 receptors of exogenous L-DOPA-derived dopamine release in the dopaminergic denervated striatum.

To determine whether dopamine release derived from exogenous l-DOPA is under inhibitory control of presynaptic dopamine D2 receptors in the dopaminergic denervated striatum, extracellular dopamine levels were measured in the striatum of 6-hydroxydopamine-lesioned rats using in vivo brain microdialysis. Quinpirole, a D2 agonist, dose-dependently (0.01-3 mg/kg s.c.) inhibited endogenous dopamine release both in the intact and dopaminergic denervated striatum. The dose-response curve obtained from the denervated striatum showed a shift to the right. Administration of l-DOPA (30 mg/kg i.p.) with carbidopa (25 mg/kg i.p.) increased dopamine release to 130% of basal levels in the intact striatum and 770% of basal levels in the denervated striatum. In the intact striatum, dopamine release was continuously inhibited by quinpirole pretreatment (1 mg/kg s.c.) even after l-DOPA administration. In the denervated striatum, l-DOPA-derived dopamine release was not affected by quinpirole pretreatment. These results suggest that, in the striatum with dopaminergic denervation, regulation by presynaptic D2 receptors is still operative on endogenous dopamine release but it does not work on dopamine release derived from exogenously administered l-DOPA.

[Reduction of apomorphine-induced rotation in rats with additional destruction of the pedunculopontine nucleus contralateral to the unilateral nigrostriatal lesion].

Pedunculopontine nucleus (PPN) has been proposed to be one of the responsible sites for dopamine agonist induced contraversive rotation in rats with unilateral dopaminergic denervation. However, previous experiments have not confirmed possible participation of PPN in this rotational behavior. To determine whether contralateral side of the PPN plays a vital role in this rotation, we investigated apomorphine induced rotation in rats with further destruction of unilateral PPN contralateral to the nigrostriatal lesion. First, rats with unilateral PPN lesioned with N-methyl-D aspartic acid (NMDA) showed contralateral rotation when they were injected with apomorphine 0.5 mg/kg s.c. Second, rats with right substantia nigra previously lesioned with 6-hydroxydopamine were microinjected with NMDA (PPN lesion group) or vehicle (sham lesion group) into the contralateral (left)side of the PPN. Contraversive rotation induced by apomorphine 0.05 mg/kg s.c. did not show any significant change in PPN lesion group as compared with sham lesion group. Whereas such rotation induced by apomorphine 0.5 mg/kg s.c. significantly reduced to 61% in PPN lesion group as compared with that in sham lesion group. These results could be interpreted as a partial cancellation of rotational activity to the contralateral side (left) by the potential rotational activity toward the opposite side (right) in rats with additional contralateral PPN lesion. Although PPN might not serve as an output route for the dopaminergic neurotransmission in the basal ganglia, PPN itself could rather induce some movement affecting the basal ganglia.

Strychnine-sensitive and strychnine-insensitive glycine binding sites in the spinal cord of the wobbler mouse.

Using quantitative autoradiography, the strychnine-sensitive glycine site and strychnine-insensitive glycine site of the N-methyl-D-aspartate receptor were analyzed in the cervical segment of the spinal cord of the wobbler mouse, which is a purported model of human motor neuron diseases. Significantly increased density of the strychnine-sensitive site was found in the lamina II-inner (+17%) and laminae III & IV (+17%) of wobbler mice. The strychnine-insensitive site was also increased in lamina I & II-outer (+15%), lamina II-inner (+15%), laminae III & IV (+48%), laminae V-VIII (+43%) and lamina X (+26%) of wobbler mice. However, no significant differences were observed for the both sites in the ventral horn where motor neurons are located. These findings suggest that both inhibitory and excitatory-associated glycinergic dysfunctions are involved in the wobbler mouse motor neuron disease.

[Isaacs' syndrome with abnormal F response and effects of double filtration plasmapheresis: a case report].

We reported a case of Isaacs' syndrome with abnormal F response detected electrophysiologically. A 14-year-old female was admitted to Hirosaki University Hospital with complaints of progressive myokymia and muscle cramp. PHT and CBZ were partially effective, but discontinued for drowsiness. A neurological examination revealed prominent myokymia and muscle cramp in the legs. The myokymia were worsened by exercise, bathing and diet. An electrophysiological examination showed characteristic F-response; high amplitude, long duration and increased number of phases. The epidural nerve block brought about a disappearance of the myokymia and an improvement of the abnormal features of F response. After repeated double filtration plasmapheresis, the myokymia and abnormal features of F response were remarkably reduced. Although Isaacs' syndrome is thought to have a hyperexcitability at the site of distal peripheral nerve, we suggested that the hyperexcitability might exist at the site of proximal region, and that immunological mechanisms underlie the cause of myokymia and unusual F-response in this case.

The effect of thymectomy on myasthenia gravis, thrombocytopenia, and granulocytopenia associated with thymoma: report of a case.

We report the case of a 47-year-old woman with thymoma who developed myasthenia gravis, thrombocytopenia, and granulocytopenia, simultaneously, the concurrent association of these four disorders being extremely rare. Thymectomy was performed, and, during the post-thymectomy course, there were surprising findings concerning the recovery of not only the myasthenia gravis but also of the hematologic disorders. Immediately after thymectomy, the myasthenic symptoms completely disappeared, and the granulocyte and platelet counts recovered to within the normal range within a few days. The laboratory data revealed no difference between pre- and post-thymectomy in the release of cytokines (tumor necrosis factor; TNF, interleukin; IL-2, and IL-6), anti-acetylcholine receptor antibody, or platelet-associated IgG. On the other hand, the serum level of anti-neutrophil cytoplasmic antibody (p-ANCA), against the myeloperoxidase of the granulocytes was dramatically decreased, after thymectomy, showing a significant correlation with the granulocyte count. According to our survey of the literature, this is the first report to show that the removal of a thymoma led to the dramatic resolution not only of myasthenia gravis but also of other associated diseases. It is possible that p-ANCA may be regulated by thymoma, thus causing severe granulocytopenia.

[A patient of late-onset nemaline myopathy with mononuclear cell infiltration].

We described a 50-year-old woman with late-onset nemaline myopathy with focal mononuclear cell infiltrates in her muscle biopsy. She developed difficulty in climbing stairs, and elevating her arms for 5 months after the onset of the disease. On admission, neurological examination revealed moderate weakness and atrophy in the proximal limb and neck muscles. Laboratory studies were within normal limits except mildly elevated serum CK and aldolase levels. Electromyography showed myopathic changes in the right triceps and quadriceps muscles examined. A biopsy from the left biceps brachii muscle revealed increased variation in fiber size, with numerous basophilic atrophic fibers. There were some foci of mild mononuclear cell infiltration. Most of basophilic fibers contained nemaline bodies on modified trichrome stain. On electron microscopy numerous nemaline bodies were present in fibers with marked myofibrillar degeneration. Azathioprine and prednisolone administration was not effective to improve her condition. As mononuclear cell infiltration has been occasionally described in the previously reported patients of adult-onset nemaline myopathy, inflammatory process may have some roles in formation of nemaline bodies on the way of acute myofibrillar degeneration.

Reduction of GABAA and GABAB receptor densities in the cerebellar cortex from 3-acetylpyridine-induced ataxic rats.

We measured GABAA and GABAB receptor densities in the cerebellar cortex from 3-acetylpyridine-induced ataxic rats using receptor autoradiography. GABAA and GABAB receptor densities were significantly reduced both in the molecular layer and the granule cell layer. Reduction of GABA receptor densities may be induced by loss of GABA receptors on the degenerated climbing fibers or by secondary or compensatory changes of neuronal activities in the cerebellum.

Quantitative autoradiographic distribution of glutamate receptors in the cervical segment of the spinal cord of the wobbler mouse.

The reduction of glutamate content has been observed in the spinal cord of the wobbler mouse, a purported model of amyotrophic lateral sclerosis (ALS). To elucidate glutamate receptors in the wobbler spinal cord, we measured densities of N-methyl-D-aspartate (NMDA), kainate, alpha-amino-3-hydroxy-5-methyl-4-isoxazole proprionic acid (AMPA) and metabotropic glutamate (mGlu) binding sites using in vitro autoradiography. In wobbler mice, NMDA, kainate, and AMPA binding sites were increased in the dorsal horn and kainate binding sites were also increased in the intermediate zone. However, mGlu binding was unchanged. These results disagree with those observed in ALS spinal cords, in which NMDA and kinate binding sites are decreased. The wobbler mouse may have the glutamate dysfunction, but in a different way from ALS.

Sinusoidal vessels in the periodontal ligament of hamster incisors: their distribution, structure and possible function.

Vascular architecture in the periodontal ligament of hamster incisors was investigated by use of vascular casts under a scanning electron microscope (SEM). In addition to ordinary nutrient blood vessels, anastomosing vessels of large caliber developed, surrounding the incisor. From their characteristic configuration, these vessels were regarded as "sinusoids". The plexus of sinusoidal vessels was connected with capillaries in the papillary layer of the enamel organ at the labial periodontal ligament, and with veins penetrating into the alveolar bone on the lingual side. Transmission electron microscopic (TEM) observation showed that the sinusoidal wall was composed of only a thin layer of endothelial cells, lacking a smooth muscular element, and surrounded by densely arranged collagen fibers. Although the frequent association of Ruffini-type nerve endings with sinusoidal vessels was noted, neither direct contact nor specialized structures between these was recognizable. A possible function of the periodontal sinusoids is discussed on the basis of their distribution and ultrastructural evidence.

Histochemical and immunohistochemical demonstration of macrophages and dendritic cells in the lingual periodontal ligament of rat incisors.

The distribution of macrophages in the lingual periodontal ligament of rat incisors was surveyed by histochemical and immunohistochemical methods. Numerous macrophages showing intense ACPase reactions were located primarily in the shear zone of the periodontal ligament. Immunostaining with an ED1-monoclonal antibody that recognizes various subpopulations of macrophages revealed plentiful positive cells showing flamelike profiles throughout the periodontal ligament, in addition to regular macrophages associated with sinusoidal blood vessels. A similar distribution of flamelike cells expressing Ia antigens was demonstrable with immunostaining using an OX6-monoclonal antibody. A consecutive staining of sections for ACPase histochemistry followed by immunoreactions for Ia antigens revealed the presence of two types of the flamelike cells in the periodontal ligament: one with and the other without distinct ACPase activity, corresponding to the macrophage and the dendritic cell, respectively. Either type of flamelike cells was located in the bone-related and shear zones, whereas only dendritic cells without ACPase activity were restricted to the tooth-related zone. OX6-immunonegative cells showing ACPase reactions were also found in the periphery of the sinusoidal blood vessels. Our data are the first to demonstrate the abundance of macrophages and dendritic cells expressing Ia antigens throughout the lingual periodontal ligament of rat incisors. In addition to regular macrophages, an exclusive localization of macrophages with flamelike extensions has been demonstrated in the bone-related and shear zones of the ligament. The region-specific arrangement of macrophages and dendritic cells with various histochemical and immunological features suggests that the periodontal ligament of rat incisor is a useful model for analyzing the process of differentiation of antigen-presenting cells.

Innervation of the periodontal ligament in the dog with special reference to the morphology of Ruffini endings.

The distribution and terminal formation of nerves in the periodontal ligament of dog incisors and canines were investigated by immunohistochemistry for neurofilament protein (NFP) and by electron microscopy. The NFP-immunoreactive nerve fibers were found to be densely distributed in the apical third of the periodontal ligament, while they were sparse in the coronal two thirds. Most of the nerve endings in the periodontal ligament showed a tree-like appearance and resembled those nerve endings demonstrated in the periodontal ligament of human and monkey under the category of free nerve endings. Presumable axon terminals of these were slightly thicker than preterminal portions, running along periodontal collagen fibers and tapering within them. In light microscopic images, at least, they differed from the Ruffini endings which are commonly seen in rodents, displaying a glove-like configuration with extremely expanded tips. Under the electron microscope, however, the tree-like endings of the dog appeared similar to the Ruffini endings of rodents: their terminals were filled with mitochondria, covered with a cytoplasmic process of a Schwann cell, and surrounded by collagen fibers. These ultrastructural findings, combined with the results of previous physiological studies suggest that the nerve endings demonstrated in the present study can be identified as Ruffini endings. It is even stressed that the dog-type of Ruffini ending can be regarded as a representative of the sensory receptors in the mammalian periodontal ligament. In addition to these endings, knobbed endings, corpuscular (lamellated and glomerular) endings, and free nerve endings were rarely encountered in the periodontal ligament of incisors and canines of the dog.

A possible mechanism of mechanoreception in Ruffini endings in the periodontal ligament of hamster incisors.

The topographical relationship between Ruffini endings and the surrounding collagen fibers in the periodontal ligament of hamster incisors was investigated by means of both immunohistochemistry for neurofilament protein (NFP) and electron microscopy. Periodontal Ruffini endings, a type of stretch receptor, were present exclusively in the alveolar half of the periodontal ligament. Their axon terminals were densely and regularly associated with transverse collagen fibers, possibly forming a mechanoreceptive complex. Since blood sinuses with frequent anastomoses extended throughout the alveolus-related part, the densely innervated collagen bundles were separated from each other by the vascular spaces. Electron microscopic observation of specimens stained with tannic acid revealed a linkage between the axon terminals of the Ruffini endings and the surrounding collagen filaments. The axon terminals were enveloped by multiple layers of the basal lamina, which were penetrated by collagen filaments. The irregularly arranged collagen filaments were sandwiched between electron-dense laminae of the multilayered basal lamina. The possible mechanism of mechanoreception by the periodontal Ruffini endings is discussed on the basis of the immunohistochemical and ultrastructural findings.

Immunohistochemical localization of laminin in the periodontal Ruffini endings of rat incisors: a possible function of terminal Schwann cells.

Ruffini endings in the periodontal ligament of rodents are ensheathed by a special type of terminal Schwann cell with a particularly developed rough endoplasmic reticulum and Golgi apparatus, and further enveloped by a characteristic multi-layered structure. In order to reveal the functional significance of the structures, localization of a laminin molecule in the periodontal Ruffini endings of rats was immunohistochemically investigated at the levels of light and electron microscopy. Immunostaining using an anti-laminin serum clearly demonstrated the profiles of the Ruffini endings as well as those of the blood vessels. Ultrastructurally, reaction products for laminin were deposited in the entire thickness of the multi-layered structure, supporting the idea that this structure is derived from the basal lamina. The basal lamina, immunoreacting with laminin antiserum, was penetrated by periodontal collagen fibers, possibly serving as an adhesive device between the Ruffini endings and surrounding collagen fibers. The laminin immunoreactive materials were also recognized in the vesicles and caveolae of the terminal Schwann cells which tended to gather at the interstitial surface of the cells. The terminal Schwann cells are therefore believed to be directly involved in the formation of the multilayered basal lamina through the active production of its materials.

Dopamine agonists potentiate antiakinetic effects of competitive NMDA-antagonists in monoamine-depleted mice.

The competitive NMDA-antagonists SDZ EAA-494 and CGP 37849 and the mixed D-1/D-2 dopamine agonists CI 201-678 and SDZ 205-152 reverse akinesia in monoamine-depleted mice in a dose dependent manner. Combination of threshold doses of NMDA-antagonists with dopamine agonists markedly enhances anti-akinetic effects. CI 201-678 which in addition to D-1 and D-2 receptors stimulates alpha-2 receptors produces a stronger effect than SDZ 205-152 which is devoid of alpha-2 agonist activity. The results indicate that concomitant blockade of NMDA-receptors and activation of dopamine receptors results in synergistic or at least additive motor stimulatory effects.

Sensory receptors in the periodontal ligament of hamster incisors with special reference to the distribution, ultrastructure and three-dimensional reconstruction of Ruffini endings.

The innervation of the periodontal ligament in hamster incisors was investigated by means of immunohistochemistry for nervous-specific proteins and electron microscopy. The lingual periodontal ligament was found to be exclusively innervated by Ruffini endings which appeared to be most developed in this species among rodents; the labial periodontal ligament lacked them. The Ruffini endings occupied the alveolar half of the periodontal ligament, being intertwined with transverse collagen fibers. In electron microscopy, the Ruffini endings displayed expanded axon terminals filled with large-sized mitochondria. Three-dimensional reconstructions of the Ruffini endings at the electron microscopic level revealed complicated shapes for the axon terminals and a characteristic relationship with the associated terminal Schwann cells. The axon terminals were plate- or knob-shaped, the former being predominant. Each axon terminal was covered by thick Schwann sheaths derived from more than two terminal Schwann cells whose cell bodies were located apart from the axon terminals and contained a developed Golgi apparatus and rough endoplasmic reticulum. On the other hand, each terminal Schwann cell simultaneously extended their cytoplasmic processes to several axon terminals just like astrocytes. The thick Schwann sheath, for the most part, was covered by a multiple layer of basal lamina. These findings have aided us in understanding the entire structure of the periodontal Ruffini endings.

Cholinesterase activity in terminal Schwann cells associated with Ruffini endings in the periodontal ligament of rat incisors.

A nonspecific cholinesterase activity was demonstrated in terminal Schwann cells associated with Ruffini endings in the periodontal ligament of rat incisors at the light and electron microscopic levels. The terminal Schwann cells are ultrastructurally characterized by a well-developed Golgi apparatus and rough endoplasmic reticulum. The cells in this study were positive for nonspecific cholinesterase, whereas ordinary Schwann cells associated with more proximal nerve fibers reacted negatively. The reaction products were densely deposited in the cisternae of the rough endoplasmic reticulum and along the nuclear envelop. A moderately intense labeling was found in the cytoplasmic extensions, in which the reaction products gathered in caveolae and vesicles. These findings indicate that nonspecific cholinesterase is a useful marker to distinguish terminal Schwann cells from ordinary Schwann cells and that the enzyme may be synthesized in the rough endoplasmic reticulum and conveyed toward the axon terminals. Since this enzyme has been known to be shared by the inner bulb of Pacinian corpuscles and the lamellar cells of Meissner's corpuscles, its possible involvement in mechanoreceptive functions in these specialized Schwann cells deserves further investigation.