Micromolar concentrations of citalopram or escitalopram inhibit glycoprotein VI-mediated and integrin αIIbß3-mediated signaling in human platelets.

Collagen and convulxin induce platelet aggregation through glycoprotein VI (GPVI)-FcRγ-Syk signaling pathway. In addition, fibrinogen induces platelet activation through integrin αIIbß3-FcγRIIa-Syk signaling pathway. We previously reported that high concentrations of selective serotonin reuptake inhibitors (SSRI) reduce platelet aggregation induced by collagen. We further investigated the effects of SSRI on GPVI- and αIIbß3-mediated signaling pathway. Citalopram and escitalopram, two relatively pure SSRI, were used in this study. Both citalopram and escitalopram concentration-dependently inhibited convulxin-induced platelet aggregation, serotonin (5-HT) release and the activation of αIIbß3. 5-HT concentration in washed platelets was unchanged after short-term treatment with citalopram. The additional 5-HT failed to fully rescue the inhibitory effect of citalopram on convulxin-induced aggregation. Convulxin-induced phosphorylation of Syk, LAT, and Akt was inhibited by citalopram and escitalopram. Citalopram inhibited the interaction between FcRγ and Syk, whereas the phosphorylation of FcRγ in response to convulxin remained unaltered. Further, citalopram inhibited the increase of the interaction between serotonin transporter and Syk induced by convulxin. In the presence of Mn2+, escitalopram inhibited the formation of lamellipodia on immobilized fibrinogen. Escitalopram did not influence the binding of fibrinogen to platelets. It inhibited the phosphorylation of Syk and PAK triggered by the adhesion on fibrinogen. Our data demonstrate that micromolar concentrations of citalopram and escitalopram inhibit GPVI- and αIIbß3-mediated platelet functions. The mechanism of the inhibitory effect of citalopram or escitalopram is not the influence on the activation of GPVI or the interaction between fibrinogen and αIIbß3, but the interaction between Syk and its upstream molecules.

Effects of epidural compression on stellate neurons and thalamocortical afferent fibers in the rat primary somatosensory cortex.

A number of neurological disorders such as epidural hematoma can cause compression of cerebral cortex. We here tested the hypothesis that sustained compression of primary somatosensory cortex may affect stellate neurons and thalamocortical afferent (TCA) fibers. A rat model with barrel cortex subjected to bead epidural compression was used. Golgi-Cox staining analyses showed the shrinkage of dendritic arbors and the stripping of dendritic spines of stellate neurons for at least 3 months post-lesion. Anterograde tracing analyses exhibited a progressive decline of TCA fiber density in barrel field for 6 months post-lesion. Due to the abrupt decrease of TCA fiber density at 3 days after compression, we further used electron microscopy to investigate the ultrastructure of TCA fibers at this time. Some TCA fiber terminal profiles with dissolved or darkened mitochondria and fewer synaptic vesicles were distorted and broken. Furthermore, the disruption of mitochondria and myelin sheath was observed in some myelinated TCA fibers. In addition, expressions of oxidative markers 3-nitrotyrosine and 4-hydroxynonenal were elevated in barrel field post-lesion. Treatment of antioxidant ascorbic acid or apocynin was able to reverse the increase of oxidative stress and the decline of TCA fiber density, rather than the shrinkage of dendrites and the stripping of dendritic spines of stellate neurons post-lesion. Together, these results indicate that sustained epidural compression of primary somatosensory cortex affects the TCA fibers and the dendrites of stellate neurons for a prolonged period. In addition, oxidative stress is responsible for the reduction of TCA fiber density in barrels rather than the shrinkage of dendrites and the stripping of dendritic spines of stellate neurons.

A Case of Multiple System Atrophy with Preexisting Alzheimer's Disease and Predating The Hot Cross Bun Sign.

PURPOSE: Synucleinopathy, tauopathy and amyloidopathy were classified as distinct clinical and pathological entities in traditional classification systems, and their interactions have been studied on neuropathology and molecular genetics recently. CASE REPORT: In this report, we present a 69-year-old male patient who had been diagnosed with probable Alzheimer's disease (AD) dementia due to progressive forgetfulness in February 2013. His Mini- Mental State Examination score was 21/30, and his Cognitive Abilities Screening Instrument score was 78/100, resulted from profound deficits in recent memory and abstract thinking domains. Initial brain magnetic resonance imaging (MRI) showed bilateral medial temporal lobe atrophy but was otherwise unremarkable. He presented with new-onset progressive gait disturbance 18 months after the diagnosis of AD, and mild ataxic gait and linear hyperintensity within the midline of the pons on axial T2-weighted MRI were documented. Neither extrapyramidal nor autonomic signs were observed. Ten months later, profound cerebellar signs, urinary incontinence, and mild axial rigidity consistent with the hot cross bun (HCB) sign were noted. Probable multiple system atrophy-cerebellar (MSA-C) type was finally diagnosed by the clinical and neuroimaging features. Of note, his diagnoses of AD and HCB sign predated the diagnosis of MSA-C by 28 and 10 months, respectively. CONCLUSION: Given that the HCB sign rarely predates overt symptoms or a diagnosis of MSA, we hypothesized that the preexisting amyloidopathy and tauopathy exerted additional neurotoxicity on the synucleinopathy. Key Words: Multiple system atrophy, hot cross bun sign, Alzheimer's disease.

(-)-Epigallocatechin gallate attenuates NADPH-d/nNOS expression in motor neurons of rats following peripheral nerve injury.

BACKGROUND: Oxidative stress and large amounts of nitric oxide (NO) have been implicated in the pathophysiology of neuronal injury and neurodegenerative disease. Recent studies have shown that (-)-epigallocatechin gallate (EGCG), one of the green tea polyphenols, has potent antioxidant effects against free radical-mediated lipid peroxidation in ischemia-induced neuronal damage. The purpose of this study was to examine whether EGCG would attenuate neuronal expression of NADPH-d/nNOS in the motor neurons of the lower brainstem following peripheral nerve crush. Thus, young adult rats were treated with EGCG (10, 25, or 50 mg/kg, i.p.) 30 min prior to crushing their hypoglossal and vagus nerves for 30 seconds (left side, at the cervical level). The treatment (pre-crush doses of EGCG) was continued from day 1 to day 6, and the animals were sacrificed on days 3, 7, 14 and 28. Nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) histochemistry and neuronal nitric oxide synthase (nNOS) immunohistochemistry were used to assess neuronal NADPH-d/nNOS expression in the hypoglossal nucleus and dorsal motor nucleus of the vagus. RESULTS: In rats treated with high dosages of EGCG (25 or 50 mg/kg), NADPH-d/nNOS reactivity and cell death of the motor neurons were significantly decreased. CONCLUSIONS: The present evidence indicated that EGCG can reduce NADPH-d/nNOS reactivity and thus may enhance motor neuron survival time following peripheral nerve injury.

Differential toxicities of intraneurally injected mercuric chloride for sympathetic and somatic motor fibers: an ultrastructural study.

BACKGROUND/PURPOSE: Mercury is a well-known neurotoxin but the susceptibility of autonomic nerves to mercury poisoning in vivo has seldom been studied. Our previous studies have shown that the hypoglossal nerve in hamsters contains somatic motor and postganglionic sympathetic fibers. The aim of this study was to investigate the ultrastructural changes in the nervous system following intraneural injection of mercuric chloride into the hypoglossal nerve in hamsters. METHODS: Six adult hamsters were used in this study. After anesthesia, the digastric muscle on the right side was removed and the trunk of the hypoglossal nerve was exposed. Two microliters of mercuric chloride aqueous solution was injected into the main trunk of the hypoglossal nerve at the bifurcation. The contralateral hypoglossal nerve was kept intact and used as the normal control. Animals were allowed to survive for 1 or 3 days and were prepared for ammonium sulfide histochemistry and electron microscopy. RESULTS: Three days after injection of mercuric chloride solution, almost all unmyelinated sympathetic fibers in the hypoglossal nerve trunk were lost, whereas myelinated somatic axons were spared. Although mercury deposition in the myelin sheaths of neuronal processes was observed in the hypoglossal nucleus, the neuronal somas were intact. By contrast, degenerated neuronal processes and mercury deposition in neuronal somas were frequently found in the superior cervical ganglia. CONCLUSION: This study demonstrated an undue susceptibility of sympathetic fibers to mercury intoxication. The mechanisms that underlie the selective reaction of sympathetic fibers to mercury warrant further investigation.

Mild hypoxic preconditioning attenuates injury-induced NADPH-d/nNOS expression in brainstem motor neurons of adult rats.

Excessive production of nitric oxide (NO) might have detrimental effects on the hypoxia-related neuropathology. This study aimed to test if mild hypoxic preconditioning (MHPC) would attenuate the pathological changes in the brainstem motoneurons having a different functional component after peripheral nerve crush injury (PNCI). Prior to PNCI treatment, young adult rats were caged in the mild hypoxic altitude chamber with 79Torr of the partial oxygen concentration ( pO(2)) (i.e., 0.5atm at 5500m in height) for 4 weeks to adapt the environmental changes. After that, all the animals having successfully crushed both the hypoglossal and vagus nerves (left-side) were allowed to survive for 3, 7, 14, 30 and 60 successive days in normoxic condition. Nicotinamine adenine dinucleotide phosphate-diaphorase (NADPH-d) histochemistry and neuronal nitric oxide synthase (nNOS) immunohistochemistry revealed that MHPC reduces NADPH-d/nNOS expression in the hypoglossal nucleus (HN) and the dorsal motor nucleus of the vagus (DMN) at different time points after PNCI. The morphological findings were further ascertained by Western blot analysis of nNOS and nitrite assay for NO production. Both the morphological and quantitative results peaked at 7 days in HN, whereas for those in DMN were progressively increased up to 60 days following PNCI. The staining intensity of NADPH-d/nNOS(+) neurons, expression of nNOS protein, NO production levels as well as the neuronal loss in HN and DMN of MHPC rats following PNCI were attenuated, especially for those having a longer survival period over 14 days. The MHPC treatment might induce minute amounts of NO to alter the state of milieu of the experimental animals to protect against the PNCI.

The spatial and segmental innervation of somatic acupoint - a study of canine Shen-Shu point (BL-23).

Although an acupuncture needle penetrates the skin, subcutaneous tissue, and underlying muscle, the most effective locus for the somatic acupoint on the needle path is not well established. We therefore investigated the sensory innervations of tissues in the needle path of the canine Shen-Shu point and evaluated their roles in initiating an acupunctural signal. Horseradish peroxidase solution was injected at all three levels within the acupoint. Only a few peroxidase-positive neurons were observed in the L1 dorsal root ganglion following intradermal injection. Following subcutaneous injection, peroxidase-labeled neurons were detected extending from spinal levels T10 to L2, with maximal labeling at T12 (46.3%). Approximately 95% of positive neurons were at spinal levels T11, T12, T13, and L1. As a result of an intramuscular injection, labeled neurons were observed at spinal levels T12 to L3, with most labeling occurring at L1 (39.9%). Approximately 95% of positive neurons were at spinal levels T13, L1, and L2. The results suggest that most afferent terminals are in the subcutaneous tissue rather than the muscular tissue, with an approximate ratio of 3.75:1. The data provide solid evidence that sensory innervation to a somatic acupoint is confined to a spinal segment and spatially organized, and we speculate that to cause a maximum effect, the centripetally transmitted signal from needling a somatic acupoint is spatio-segmental and divergently amplified.

Neurophysiologic changes after preganglionic and postganglionic nerve-root constriction: an experimental study in the rat.

STUDY DESIGN: We investigated changes in spinal somatosensory-evoked potential (SSEP) and nerve action potential (NAP), correlated behavior, and associated pathologic observation in experimental radiculopathy. OBJECTIVES: To create a rat model of sacrococcygeal radiculopathy for determining the validity of SSEP and NAP. SUMMARY OF BACKGROUND DATA: We examined the diagnostic sensitivity and value of electrophysiologic tests for evaluating lumbosacral root disease conflict. An appropriate animal model can help verify the value of these tests. METHODS: Preganglionic lesion group rats were given 2 loose ligatures around the cauda equina at the sacrum, and postganglionic lesion group rats were given 2 loose ligatures on the conjunction of the sacrococcygeal nerve roots and the caudalis nerve after they had received a laminectomy. Control group rats received a sham operation. SSEPs and NAPs were recorded preligature and postligature, and 3 times after surgery. These electrophysiologic observations were compared and correlated with tail-flick reflex and histology. RESULTS: All experimental group rats developed thermal hyperalgesia on day 14, as indicated by a significant reduction in TFL (tail-flick latency), which continued for 3 months. Amplitude decreased significantly and latency increased significantly in all SSEP recordings immediately after the operation; these changes persisted for 3 months. There were no significant differences between the experimental groups, but there were significant differences between the control and experimental groups. NAP amplitude and latency from the caudalis nerves did not change in any group in the first 2 postoperative weeks. From the second postoperative week until the 3-month follow-up, amplitude was significantly decreased and latency prolonged in the postganglionic group but unchanged in the others. CONCLUSIONS: Both SSEP and NAP are useful for evaluating electrophysiologic changes after various radiculopathies. The data also suggest that the conductivity of the peripheral nerve (NAP) was affected by the postganglionic compression of the corresponding nerve root, but not by the preganglionic lesion.

The regulatory mechanism of Tremella mesenterica on steroidogenesis in MA-10 mouse Leydig tumor cells.

Tremella mesenterica (TM), a yellow jelly mushroom, has been traditionally used as tonic food to improve body condition in Chinese society for a long time. We have previously demonstrated that TM reduced in vitro hCG-treated steroidogenesis in MA-10 mouse Leydig tumor cells without any toxicity effect. In the present study, the mechanism how TM suppressed hCG-treated steroidogenesis in MA-10 cells was investigated. MA-10 cells were treated with vehicle, human chorionic gonadotropin (hCG, 50 ng/ml), or different reagents with or without TM to clarify the effects. TM significantly suppressed progesterone production with the presences of forskolin (10 and 100 microM) or dbcAMP (0.5 and 1mM), respectively, in MA-10 cells (p<0.05), which indicated that TM suppressed steroidogenesis after PKA activation along the signal pathway. Beyond our expectation, TM induced the expression of steroidogenic acute regulatory (StAR) protein with or without hCG treatments. However, TM profoundly decreased P450 side chain cleavage (P450scc) and 3beta-hydroxysteroid dehydrogenase (3beta-HSD) enzyme activities without any influences on the expression of both enzymes. These inhibitions on steroidogenic enzyme activities might counteract the stimulation of StAR protein expression. In conclusion, results suggest that TM suppressed hCG-treated steroidogenesis in MA-10 cells by inhibiting PKA signal pathway and steroidogenic enzyme activities.

The in vivo effect of Cordyceps sinensis mycelium on plasma corticosterone level in male mouse.

Cordyceps sinensis (CS), an Ascomycetes fungus parasitic to Lepidoptera larvae, has been traditionally used as nutritious food for the enhancement in immuno-modulation in Chinese society for a long time. Previous report has demonstrated the CS water extract stimulates in vitro corticosterone production in rat primary adrenal cells. In the present studies, we determined the in vivo effects of CS and its fractions on plasma corticosterone production in mouse. Different concentrations of CS and CS fractions dissolved in water (0.02 and 0.2 mg/g body weight) were fed to immature and mature mice from 1, 3 or 7 d. The plasma levels of corticosterone were determined by radioimmunoassay (RIA), and the weight of adrenal gland and body weight were also evaluated. Results illustrated that plasma corticosterone levels were significantly induced by F2 at 0.02 mg/g body weight with 7 d feeding in immature mice, and by CS at 0.02 mg/g body weight with 3 d feeding and F3 at 0.02 mg/g body weight for 7 d feeding in mature mice, respectively (p < 0.05). There were no differences of adrenal gland weight except there was significant stimulation by CS at 0.2 mg/g body weight with 3 d feeding in mature mice (p < 0.05) and there were significant inhibitions by both dosages of F3 for 3 d feeding in immature mice and F2 for 7 d feeding in mature mice (p < 0.05), respectively. Concerning body weight, the stimulatory effects were observed with CS feeding at 0.2 mg/g body weight for 7 d and F3 feeding at 0.02 mg/g body weight for 3 and 7 d in mature mice. Whereas, the inhibitory effect were observed in F2 feeding at 0.2 mg/g body weight for 7 d in immature mice and at both dosages for 7 d in mature mice, respectively. Taken together, these studies illustrate that CS and its fractions stimulated mouse in vivo corticosterone production. However, CS and its fractions didn't have constant stimulatory or inhibitory effects on the weights of body and adrenal glands.

Ultrastructural identification of a sympathetic component in the hypoglossal nerve of hamsters using experimental degeneration and horseradish peroxidase methods.

We employed experimental degeneration, tract-tracing with wheatgerm agglutinin conjugated with horseradish peroxidase (WGA-HRP) and electron microscopy to explore the postganglionic sympathetic fibers in the hypoglossal nerve of hamsters. Quantitative results of normal untreated animals at the electron microscopic level showed the existence of unmyelinated fibers, which made up about 20% of the total fibers in the nerve, being more numerous on the left side. The nerve fibers were preferentially distributed at the periphery of the nerve. Following superior cervical ganglionectomy, most of the unmyelinated fibers underwent degenerative changes. Tract-tracing studies showed that some of the unmyelinated fibers were labeled by WGA-HRP injected into the superior cervical ganglion (SCG). It is suggested that the unmyelinated fibers represent the postganglionic sympathetic fibers originated from the SCG.

Melatonin restores the cytochrome oxidase reactivity in the nodose ganglia of acute hypoxic rats.

This study aimed to elucidate whether melatonin would exert beneficial effects on the neuronal functions of the nodose ganglion (NG) following acute hypoxic insult. The cytochrome oxidase (COX) and the nicotinamine adenine dinucleotide phosphate diaphorase (NADPH-d) histochemistry along with the nitric oxide synthase (NOS) immunofluorescence were used to examine the metabolic stage and nitric oxide production in nodose neurons respectively. Adult rats were injected intraperitoneally with melatonin at 5 or 100 mg/kg. Hypoxia was achieved by placing the rats into an altitude chamber (PO2 = 43 torr) for 4 hr. The results show that in normal untreated rats, nearly all and about 43% of the NG neurons displayed COX and NOS/NADPH-d reactivities with various staining intensities respectively. However, COX reactivity was drastically decreased while NOS/NADPH-d reactivity was significantly upregulated following hypoxia treatment. In melatonin pretreated rats, the hypoxia-induced reduction of COX reactivity was obviously prevented and the augmentation of NOS/NADPH-d reactivity was successfully suppressed. The deficit in the metabolic stage and the over-activation of NOS would contribute to the generation of oxidative stress. By effectively preventing the metabolic disruption, melatonin may have potential utility in therapeutic treatment of neuronal dysfunctions where oxidative stress is a participant.

Neuronal NADPH-d/NOS expression in the nodose ganglion of severe hypoxic rats with or without mild hypoxic preconditioning.

This study aimed to test the hypothesis that mild hypoxic preconditioning (MHPC)-induced NOS expression would attenuate the neuropathological changes in the nodose ganglion (NG) of severe hypoxic exposure (SHE) rats. Thus, the young adult rats were caged in the altitude chamber for 4 weeks prior to SHE for 4 h to gain hypoxic preconditioning. The altitude chamber was used to set the height at the level from 5500 m (0.50 atm; pO2=79 Torr) to 10,000 m (0.27 atm; pO2=43 Torr) for MHPC and SHE, respectively. The experimental animals were allowed to survive for 0, 7, 14, 30 and 60 successive days, respectively. Nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) histochemistry and neuronal nitric oxide synthase (nNOS) immunohistochemistry were used to detect NADPH-d/nNOS reactivity in the NG at various time points following hypoxic exposure. The present results showed that about 38% of the neurons in the NG displayed NADPH-d/nNOS positive [NADPH-d/nNOS(+)] in normoxic rats. In SHE rats, a peak in the percentage (71%) and staining intensity (230%) of NADPH-d/nNOS(+) nodose neurons at 0 day, which then gradually decreased at 7-60 days. About 25% of the nodose neurons died 60 days after SHE. However, in MHPC rats subjected to SHE, NADPH-d/nNOS(+) neurons peaked in the percentage (51%) and staining intensity (171%) at 0 day, which then decreased at 7-60 days. In addition, neuronal survival was markedly increased by MHPC. These results suggested that MHPC might have a neuroprotective effect that reduces the susceptibility of the nodose neurons to NOS mediated neuropathy subsequent to SHE.

Witzelsucht after right putaminal hemorrhage: a case report.

Witzelsucht is a tendency to tell inappropriate and poor jokes. It usually occurs after a focal lesion involving orbitofrontal cortical or paramedian thalamic regions, especially on the right side. Here we report a 56-year-old man developing witzelsucht and hypersexuality after a right putaminal hemorrhage. The hematoma extended to the sublenticular part of posterior internal capsule and mesencephalon. The hemorrhage might have disconnected the fibers in the ascending reticular systems, and the fibers between paramedian thalamus and orbitofrontal cortex, and thus could be responsible for the patient's rare clinical manifestations.

Differential expression of calcitonin gene-related peptide (CGRP) and choline acetyltransferase (ChAT) in the axotomized motoneurons of normoxic and hypoxic rats.

We employed a double injury model (axotomy along with hypoxia) to determine how nerve injury and hypoxic insult would affect the expression of calcitonin gene-related peptide (CGRP) and choline acetyltransferase (ChAT) in the hypoglossal nucleus (HN) and nucleus ambiguus (NA). Adult rats were subjected to unilateral vagus and hypoglossal nerve transection, following which half of the animals were kept in an altitude chamber (PO2=380 Torr). The immunoexpression of CGRP and ChAT (CGRP-IR/ChAT-IR) were examined by quantitative immunohistochemistry at 3, 7, 14, 30 and 60 days post-axotomy. The results revealed that CGRP-IR in the HN was increased at 3 days but decreased to basal levels at 7 days following nerve injury. The decline was followed by a second rise in CGRP-IR at 30 days post-axotomy, followed again by a return to basal levels at 60 days. In the NA, CGRP-IR was up-regulated at 3 days and remained increased for up to 60 days after nerve injury. Animals treated with a double injury showed a greater CGRP-IR than normoxic group in both nuclei at all post-axtomized periods. In contrast to CGRP, ChAT-IR was markedly reduced in the HN and NA at 3 days reaching its nadir at 14 days following nerve injury. Hypoxic animals showed a stronger reduction of ChAT-IR in both nuclei at all post-axtomized periods. Results of cell counting showed that neuronal loss was somewhat obvious in hypoxic HN than that of normoxic ones. The present results suggest that up-regulation of CGRP-IR may exert its trophic effects while down-regulation of ChAT-IR may correlate with the poor neurotransmission within the injured neurons. It is speculated that the enhanced expression of CGRP-IR and the pronounced reduction of ChAT-IR in hypoxic rats may result from a drastic shift of intracellular metabolic pathways, which in turn could lead to more metabolic loading to the severely damaged neurons following the double insult.

Synaptic remodeling in the nucleus ambiguus following vagal-hypoglossal nerve anastomosis in the cat.

We reported recently the occurrence of a massive and selective elimination of synaptic boutons on motoneurons in the dorsal motor nucleus of the vagus (DMV) in the cat following vagal-hypoglossal nerve anastomosis (VHA) [J. Comp. Neurol. 458 (2003) 195]. This study was aimed to explore the synaptic reorganization in the other major nucleus associated with the vagus, namely, the nucleus ambiguus (NA) following the same treatment. In view of the tremendous difference in function, the NA and DMV are considered to be two ideal nuclei for explanatory studies seeking to elucidate how VHA could induce different plasticity of brainstem neurons influenced by the newly reestablished neural pathway. The present results showed that the vagal efferent neurons in the NA had responded to VHA in a different manner compared with those in the DMV. Firstly, the numbers of axon terminals containing round (R), round with dense-cored (R+D), pleomorphic (P) or flattened (F) synaptic vesicles contacting the NA motoneurons were markedly increased at 500-day postoperation, the longest reinnervation interval. The percent increases in the synapse frequency for R, R+D, P and F boutons were 8.6%, 274.4%, 238.3% and 400.0%, respectively. Secondly, the formation of astroglial ensheathment around the motoneurons in the DMV following VHA was not evident in the NA. Another striking difference was the extensive dendritic sprouting of the NA neurons as opposed to the dendritic retraction of the DMV neurons as shown by a significant increase in distal dendrites of NA motoneurons. The different modes of neural remodeling between NA and DMV may be attributed to the unique nature of the two nuclei to structures they normally supply and their different compatibility with the newly innervated target, viz. tongue skeletal musculature.