The distribution of calbindinD-28k and parvalbumin immunoreactive neurons in the somatosensory area of the pigeon pallium.

GABAergic interneurons regulate the degree of glutamatergic excitation and output of projection neurons. In this study, we investigated the distribution of calbindinD-28k (CB) and parvalbumin (PV) in the somatosensory area of the pigeon pallium using immunohistochemical method. Our results show that anatomical structures of the somatosensory area of the pigeon pallium consisted of several subdivisions including the hyperpallium, intercalated hyperpallium, mesopallium, nidopallium and basorostralis. Neuronal density was significantly higher in the intercalated hyperpallium and basorostralis than that in the other subdivisions. The density of the CB immunoreactive neurons was generally similar in all the subdivisions; however, the density of PV immunoreactive neurons was particularly prominent in the basorostralis compared with that in the other subdivisions. In addition, the mean proportion of PV immunoreactive neurons to total neurons was higher than that in the CB immunoreactive neurons in all the subdivisions. In brief, our present study shows that PV immunoreactive neurons in the somatosensory area of the pigeon pallium were significantly abundant compared with CB immunoreactive neurons. This finding needs more studies regarding CB- and PV-related functions in the somatosensory area of the avian pallium.

The location of projection neurons to the biceps brachii muscle in the telencephalon of the pigeon.

Few studies regarding the anatomical distribution of motor neurons innervating muscles of the arm have been demonstrated in avian brains. The purpose of this study was to finely determine the localization of cerebral neurons innervating the biceps brachii muscle in the pigeon. The cholera toxin B subunit (CTB) was employed as a retrograde tracer to determine the location of neurons controlling the biceps brachii muscle in the telencephalon following intramuscular injection in male pigeons (n = 7), which were killed 14 days after intramuscular injection with CTB. We found that CTB-labelled neurons were located contralaterally in the hyperpallium apicale of the rostral telencephalon and that most of the CTB-labelled neurons were pyramidal in shape. This study shows that CTB is easily taken up by nerve terminals which innervate the biceps brachii muscle of the pigeon and that cerebral motor neurons controlling the biceps brachii muscle are located in the hyperpallium apicale.

An extract of Polygonum multiflorum protects against free radical damage induced by ultraviolet B irradiation of the skin.

Over the last decades, the incidence of ultraviolet B (UVB)-related skin problems has been increasing. Damages induced by UVB radiation are related to mutations that occur as a result of direct DNA damage and/or the production of reactive oxygen species. We investigated the anti-oxidant effects of a Polygonum multiflorum thumb extract against skin damage induced by UVB irradiation. Female SKH-1 hairless mice were divided into three groups: control (N = 7), distilled water- (N = 10), and P. multiflorum extract-treated (PM, N = 10) groups. The PM (10 g) was extracted with 100 mL distilled water, cryo-dried and 9.8 g was obtained. The animals received a topical application of 500 microL distilled water or PM extract (1, 2, 4, 8, and 16%, w/v, dissolved in distilled water) for 30 min after UVB irradiation (wavelength 280-320 nm, 300 mJ/cm(2); 3 min) of the dorsal kin for 14 days, and skin immunohistochemistry and Cu,Zn-superoxide dismutase (SOD1) activity were determined. SOD1 immunoreactivity, its protein levels and activities in the skin were significantly reduced by 70% in the distilled water-treated group after UVB irradiation compared to control. However, in the PM extract-treated groups, SOD1 immunoreactivity and its protein and activity levels increased in a dose-dependent manner (1-16%, w/v, PM extract) compared to the distilled water-treated group. SOD1 protein levels and activities in the groups treated with 8 and 16%, w/v, PM extract recovered to 80-90% of the control group levels after UVB. These results suggest that PM extract strongly inhibits the destruction of SOD1 by UV radiation and probably contains anti-skin photoaging agents.

Changes of pyridoxal kinase expression and activity in the gerbil hippocampus following transient forebrain ischemia.

In the previous study, we observed chronological alterations of glutamic acid decarboxylase (GAD), which is the enzyme converting glutamate into GABA. GAD isoforms (GAD65 and GAD67) differ substantially in their interactions with cofactor pyridoxal 5'-phosphate, which is catalyzed by pyridoxal kinase (PLK). In the present study, we examined the chronological changes of PLK expression and activity in the hippocampus after 5 min transient forebrain ischemia in gerbils. PLK immunoreactivity in the sham-operated group was detected weakly in the hippocampus. Ischemia-related change of PLK immunoreactivity in the hippocampus was significant in the hippocampal cornu ammonis (CA1)region, not in the hippocampal CA2/3 region and dentate gyrus. PLK immunoreactivity was observed in non-pyramidal GABAergic neurons at 30 min to 3 h after ischemic insult. At 12 h after ischemic insult, PLK immunoreactivity was shown in many CA1 pyramidal cells as well as some non-pyramidal cells. At this time point, PLK immunoreactivity and protein content was highest after ischemia. Thereafter, PLK immunoreactivity and protein content is decreased time-dependently by 4 days after ischemic insult. Four days after ischemia, some astrocytes expressed PLK in the CA1 region. The specific PLK activity was not altered following ischemic insult up to 2 days after ischemic insult. Thereafter, the specific PLK activity decreased time-dependently. However, total activity of PLK was significantly increased 12-24 h after ischemic insult, and thereafter total activity of PLK decreased. Therefore, we suggest that the over-expression of PLK in the CA1 pyramidal cells at 12 h after ischemia may induce increase of GAD in the CA1 pyramidal cells, which plays an important role in delayed neuronal death via the increase of GABA or enhancement of GABA shunt pathway.

Ischemia-related changes of adrenocorticotropic hormone immunoreactivity and its protective effect in the gerbil hippocampus after transient forebrain ischemia.

In the present study, the temporal and spatial alterations of adrenocorticotropic hormone (ACTH) immunoreactivity in the gerbil hippocampus after 5 min transient forebrain ischemia were investigated as followed up 7 days after ischemic insult, and the effects of ACTH after ischemic insult were also investigated 4 days after ischemic insult. The ectopic expression of ACTH (1-24 fragments) immunoreactive neurons in the cornus ammonis 1 (CA1) region of hippocampus and hilar region of the dentate gyrus 1 day after the ischemic insult was observed. Judging from the double immunofluorescence study, these neurons contain GABA. Four days after ischemic insult, the ACTH immunoreactivity was localized in CA1 pyramidal cells and glia near the stratum pyramidale, which normally do not express ACTH. In addition, in the saline-treated groups, the percentage of the detected Cresyl Violet positive neurons was 11.2% compared with the sham-operated group 4 and 7 days after ischemic insult. In these groups, the OX-42 immunoreactive microglia were detected in the strata pyramidale, oriens and radiatum. However, in the Org2766 (analog of ACTH)-treated group, 57.8% neurons compared with the sham-operated group were stained with Cresyl Violet 4 and 7 days after ischemic insult. In these groups, the OX-42 immunoreactive microglia were significantly reduced in the stratum pyramidale. These results suggest that transient forebrain ischemia may provoke selective ectopic and enhanced expression of ACTH in the hippocampus, and further suggest that ACTH plays an important role in reducing the ischemic damage.

The alteration of corticotropin-releasing factor (CRF) receptor immunoreactivity in the gerbil hippocampus and neocortex following ischemic insults.

Recently, we suggested that the ectopic expression of corticotropin-releasing factor (CRF) is associated with processes linked to neuronal injury and/or degeneration in response to an ischemic insult. However, little experimental data currently links the CRF receptor directly to neuronal death induced by ischemia. Therefore, in the present study, we investigated the temporal and spatial changes in CRF receptor immunoreactivity in the hippocampus and the neocortex after transient ischemia. CRF receptor immunoreactivity in the hippocampus was reduced up to 24h after ischemia insult, as compared to the sham. Interestingly, CRF receptor immunoreactivity disappeared in the CA1 region of the hippocampus at 4 days in the post-ischemic group. The other regions of hippocampus maintained their immunoreactivities at this time point. On the other hand, in the neocortex, 3h after transient ischemia, the CRF receptor immunoreactivity was elevated in regions vulnerable to ischemia. At 12h post-ischemia, its immunoreactivity had decreased versus the sham operated animals. These results suggest that the selectively ectopic expression of CRF following ischemia, which we reported previously, may regulate inflammatory responses. In addition, these findings also suggest that the mechanisms of neuronal death as mediated by CRF receptor differ in the hippocampus and the neocortex.

Multiclinic evaluation of the antihypertensive effect of a methyldopa, hydrochlorothiazine, and amiloride combination.

A multiclinic, double-blind, randomized study in 361 patients with essential hypertension compared Aldoretic, (Merck & Co., Inc., New Jersey), a combination of methyldopa, hydrochlorothiazide, and amiloride with the combination of hydrochlorothiazide and amiloride and with methyldopa alone. At the end of week 12, the proportion of patients who were classified as normotensive (diastolic less than or equal to 90 mm Hg) was 61% in the group treated with Aldoretic, which was statistically significantly greater than the 48% in the hydrochlorothiazide/amiloride group and the 35% in the methyldopa group.

Effects of arachidonic acid and cyclo (his-pro) on zinc transport across small intestine and muscle tissues.

Previously we have shown that arachidonic acid (AA) plus zinc or cyclo (his-pro) (CHP) plus zinc improve clinical signs of diabetes in streptozotocin-induced diabetic rats. Since streptozotocin destroys pancreatic beta-cells, we hypothesize that the effect of either AA or CHP, plus zinc on glucose metabolism is via mobilization of intracellular zinc which in turn stimulates glucose uptake by peripheral tissues. We now report the relationship between zinc and AA and between zinc and CHP in controlling zinc influx and efflux across hindlimb muscle cells isolated from three-month old rats. Although CHP increased muscle zinc influx in a dose-dependent manner, AA was not effective. However, AA was more effective in stimulating zinc efflux than CHP. We have previously demonstrated that AA stimulates intestinal zinc uptake and absorption, and now present evidence that CHP also influences intestinal zinc transport. These results suggest that both AA and CHP affect glucose uptake in muscle cells via stimulating intestinal zinc absorption and muscle cell zinc flux.

Analysis of physicochemical factors related to the automatic pellicle removal in Korean chestnut (Castanea crenata).

Analyses were carried out for condensed tannin content, soluble sugar content, color, firmness, and peeling ratio of the pellicle of 14 varieties of Korean chestnut. The correlation between these physicochemical components and the peeling ratio was measured. The total soluble sugar content ranged from 4.20 to 11.83%. Condensed tannin contents in the inner and outer chestnut shells were 7.83-71.42% and 0.31-2.04%, respectively. In the outer shell of chestnut, CIE L* value was 26.98-33.34, CIE a value was 7.56-14.90, and CIE b value was 7.36-15.67. Firmness of chestnut was 4.41-9.20 kg. Peeling ratio of the pellicle by the peeling machine was significantly different with variety. The average peeling ratio was 63.79%, and that of the Okkwang variety was the highest at 85.29%. A negative correlation was found between tannin content of inner shell and peeling ratio (r = -0.57), but the correlation coefficients between other components and the peeling ratio were not significant.

Purification and characterization of a bacteriocin produced by Lactococcus lactis subsp. lactis H-559 isolated from kimchi.

Lactic acid bacteria were isolated from kimchi and screened for bacteriocin production. Strain H-559, identified as Lactococcus lactis subsp. lactis, exhibited the strongest antibacterial activity among them and was active against pathogenic bacteria such as Listeria monocytogenes and Staphylococcus aureus as well as many lactic acid bacteria. The antimicrobial substance produced by L. lactis subsp. lactis H-559 was inactivated by alpha-chymotrypsin, and protease type IX and XIV and was confirmed to be a bacteriocin. The bacteriocin activity was stable from pH 2.0-11.0 and up to 10 min heating at 100 degrees C. The bacteriocin was sequentially purified by ammonium sulfate precipitation, ion-exchange chromatography, and reversed-phase high-performance liquid chromatography (HPLC). Its molecular weight was determined to be 3343.7 Da by MALDI-mass spectrometry. Isoleucine was detected as the first N-terminal amino acid residue but the remaining amino acid sequence could not be determined by the Edman degradation method. It was different from other bacteriocins in terms of pH stability, molecular weight, amino acid composition, and the partial amino acid sequences of peptides obtained by acid hydrolysis.

Phlegmonous enteritis in a patient with congestive heart failure and colon cancer.

Phlegmonous enteritis is a rare infective inflammatory disease of the intestine, predominantly involving the submucosal layer. It is difficult to diagnose and often fatal. Its association with alcoholism and various liver diseases, although rarely reported, is well documented. We report a case of phlegmonous enteritis in a male patient with congestive heart failure and colon cancer, and describe the ultrasonographic and CT findings.

Evaluation of FOXP3 expression in canine mammary gland tumours.

The expression of the transcription factor forkhead box P3 (FOXP3) was examined in 62 canine mammary gland tumours via immunohistochemical analysis and association with the known expression of the oestrogen receptor-alpha (ERα), progesterone receptor (PR), c-erbB-2 receptor (HER2/neu), and survival. Positive staining for FOXP3 was present in 22.6% of the tumours and was associated with the histological type. Negative staining for FOXP3 was associated with positive ERα and PR expression (P < 0.001). The expression of FOXP3 in canine mammary gland tumours was significantly associated with the disease-free survival time (P = 0.029). The FOXP3 expression was not an independent prognostic factor in the multivariate analysis, though. The negative expression of FOXP3 in the canine mammary gland tumours was found to be related to histopathologic benignity and a longer survival time in canine mammary gland tumours.

Pituitary adenylate cyclase-activating polypeptide-immunoreactive cells in the ageing gerbil hippocampus.

In the present study, we investigated age-related changes in pituitary adenylate cyclase-activating polypeptide (PACAP) immunoreactivity and its protein levels in the gerbil hippocampus at various ages using immunohistochemistry and western blot analysis. In the post-natal month 1 (PM 1) group, PACAP-immunoreactive cells were found in all hippocampal subregions. The number of PACAP-immunoreactive cells was decreased in the PM 3 group and was still more decreased in the PM 6 and 12 groups. Thereafter, in the PM 18 and 24 groups, PACAP-immunoreactive cells were significantly increased again. However, in the mossy fibre zone, PACAP immunostaining was very strong in the adult group, especially in the PM 6 group. In addition, PACAP protein level was highest at PM 6, showing a slight decrease at PM 24. These results indicate that PACAP-immunoreactive cells are lowest in the adult stage and highest in the aged stage. However, PACAP immunoreactivity in the mossy fibre zone and PACAP protein level in the hippocampus are highest in the adult stage.

Microtubule associated protein 2 and choline acetyltransferase immunoreactivity in the lumbar spinal cord of young adult and aged dogs.

German Shepherds are a good model for research about aging and neurological disorders such as lumbosacral spinal canal stenosis. We compared neurons, glia and cholinergic neurons in the ventral horn of the lumbar spinal cord (L(3)) between adult (1-2 years old) and aged (10-12 years old) groups. Any pathological findings were not found by hematoxylin and eosin staining and neurological examination, and the number of NeuN (a marker for neurons)-positive neurons were similar in both groups. Microtubule-associated protein 2 (MAP2) immunoreactive dendrites in the aged dog were decreased without any change in ß-tubulin protein level. Glial fibrillary acidic protein (a marker for astrocytes) and ionized calcium-binding adapter molecule 1 (a marker for microglia) immunoreactivity were not significantly changed in both groups. The number of ChAT immunoreactive neurons was decreased; however, its protein level was not significantly changed in the aged group. These results suggest that numbers of ventral horn neurons are not changed, but cholinergic neurons may change in aged dogs compared to adult dogs.

Immunoreactivity and protein levels of olfactory marker protein and tyrosine hydroxylase are not changed in the dog main olfactory bulb during normal ageing.

The immunoreactivity and protein expression of olfactory marker protein (OMP) and tyrosine hydroxylase (TH) in the main olfactory bulb (MOB) of the dog during normal ageing was investigated. OMP immunolabelling was observed only in nerve bundles of the olfactory nerve (ONL) and glomerular layers (GL) and there was no OMP expression within cell bodies of any layer. TH immunolabelling was detected in all layers of the MOB except for the ONL. Most of the neurons expressing TH were distributed in the juxtaglomerular region and had a morphological appearance consistent with periglomerular, external tufted or superficial short axon cells. Dendrites of TH-immunoreactive neurons were closely apposed to OMP-immunoreactive nerve bundles within the glomeruli. There was no significant age-related loss of OMP and TH immunoreactivity and protein concentrations of these molecules were consistent in dogs of different ages. These results suggest that olfactory signal transduction to the GL via axons of olfactory receptor neurons remains unchanged during ageing in the dog.

Sensory and instrumental analysis for slipperiness and compliance of food during swallowing.

In spite of its importance, there have been few attempts to evaluate the sensory attributes of the food bolus during swallowing. In the current study, the slipperiness, the degree of slide for the food bolus through the mucosal surface of the oro-pharynx, and the compliance, how easily the shape of a food bolus can be transformed for automatic and comfortable swallowing, were derived among several sensory attributes related to the swallowing. Therefore, the study aims were twofold: (1) to develop the methods of sensory and instrumental analyses for determining the slipperiness and compliance of the food bolus during swallowing and (2) to examine the appropriateness of the newly designed devices by correlating the data between the sensory and instrumental analyses. Six commercial food products were evaluated by 10 trained panelists for each attribute. The devices for assessing each attribute were developed in consideration of the oro-pharyngeal movements. The sensory and instrumental analyses showed high correlation and regression coefficients as well as intensity differences of the 6 samples for each attribute. In conclusion, the slipperiness and the compliance were suitable for acquiring a better understanding of the sensory attributes of the food bolus during swallowing, and the newly developed devices showed a high potential for determining those attributes.

Immunohistochemical localization of glutamate in the gerbil main olfactory bulb using an antiserum directed against glutamate.

Information on the localization and the roles of glutamate in the nervous system is becoming valuable because the axon terminals of the olfactory sensory neurons and the synapses of the mitral and tufted output cells appear to be glutamatergic. In this study, we have analysed the distribution of glutamate immunoreactivity in the main olfactory bulb (MOB) of the Mongolian gerbil using an antiserum directed against glutamate. Glutamate immunoreactivity in the MOB was present in the olfactory nerve layer (Onl), glomerular layer (GL), external plexiform layer (EPL) and mitral cell layer (ML), but not in the granule cell layer (GCL). Glutamate immunoreactivity detected in the Onl was thought to be terminal ramifications of glomeruli. Some neurons in the periglomerular region showed glutamate immunoreactivity. In the EPL, glutamate immunoreactivity was found in some neuronal somata (tufted cells) and processes. In addition, mitral cells in the ML were labelled by the glutamate antibody. The pattern of glutamate immunoreactivity in the mitral cells was similar to that in the tufted cells. In brief, glutamate in the gerbil MOB is the neurotransmitter used by primary afferents and output neurons.

Comparative study on calretinin immunoreactivity in gerbil and rat retina.

Expression of calretinin in retina has been ascribed to multiple biological and functional aspects in the visual system. In this study, we examined the distribution patterns of calretinin immunoreactivity in gerbil and rat retina. In the gerbil, calretinin immunoreactivity was present in bipolar and amacrine cells of the inner nuclear layer and in neurones of the ganglion cell layer. In the rat, amacrine and ganglion cells showed calretinin immunoreactivity, but bipolar cells did not contain calretinin immunoreactivity. In both species, calretinin immunoreactivity was absent in cones, cone bipolars, and horizontal cells. In conclusion, gerbil as well as rat has a rod-dominant retina. The differences in calretinin expression between rat and gerbil require further investigations under various functional and developmental conditions.

Histochemical and electron microscopic study on motor neurone degeneration following transient spinal cord ischaemia at normothermic conditions in rabbits.

This study was carried out to investigate the motor neurone degeneration in the ventral horn following transient spinal cord ischaemia at normothermic conditions in rabbits. Transient spinal cord ischaemia was induced by occlusion of the abdominal aorta underneath the left renal artery for 15 min at normothermia (38.7 degrees C). Sections at the level of L7 were examined using histochemical and electron microscopic methods. Cresyl violet-positive motor neurones began to reduce in number at 3 h after ischaemia reperfusion, and were not detectable at 48 h after ischaemia reperfusion. Acid fuchsin-positive motor neurones were detected at 1 h after ischaemia reperfusion, significantly increased up to 6 h after the ischaemia reperfusion, and eventually disappeared by 48 h after ischaemia reperfusion. In electron microscopic findings, the disintegration of cytoplasmic membranes, and the disruption of mitochondria and endoplasmic reticulum were observed in motor neurones at 30 min after ischaemia reperfusion. Motor neurones showed necrotic findings with pyknotic degeneration at 1 h after ischaemia reperfusion. The necrotic degeneration became severer time dependently after ischaemia reperfusion. At 48 h after ischaemia reperfusion, cellular components were not detectable in motor neurones. In conclusion, we suggest that the degeneration pattern of motor neurones of the ischaemic spinal cord was necrotic after ischaemia reperfusion under normothermic conditions.

The analysis of titration studies in phase III clinical trials.

Clinical trials commonly employ the titration design for certain drugs such as antihypertensives. In a Phase III trial the design has purposes distinct from those of a Phase I or II trial, as well as from those of a trial with a parallel design. In this paper we compare the titration design with the usual parallel design in their respective purposes for Phase III trials, explore the relevant questions addressed, and examine typical data from such trials. We also discuss work which focuses primarily on the Phase I or II titration trials. We formulate the problem in the framework of one-way contingency table augmented with incomplete data and obtain the maximum likelihood estimates of the parameters and their estimated variances/covariances via the EM algorithm. An example of a Phase III study of an antihypertensive agent illustrates the proposed procedure.