ABSTRACT
We describe a case series of suspected metallic embolism after coil embolization for intracranial aneurysms. Between January 2012 and December 2014, 110 intracranial aneurysms had been treated by coil embolization in our institution. In 6 cases, the postprocedural MR imaging revealed abnormal spotty lesions not detected on the preprocedural MR imaging. The lesions were also undetectable on the postprocedural CT scan. They were demonstrated as low-intensity spots on T1WI, T2WI, DWI, and T2*-weighted imaging. On DWI, they were accompanied by bright "halo," and on T2*-weighted imaging, they showed a "blooming" effect. In 3 of the 6 cases, follow-up MR imaging was available and all the lesions remained and demonstrated no signal changes. Although histologic examination had not been performed, these neuroradiologic findings strongly supported the lesions being from metallic fragments. No specific responsible device was detected after reviewing all the devices used for the neuroendovascular treatment in the 6 cases.
Subject(s)
Embolization, Therapeutic/adverse effects , Intracranial Aneurysm/therapy , Intracranial Embolism/etiology , Adult , Aged , Aged, 80 and over , Diffusion Magnetic Resonance Imaging , Female , Humans , Intracranial Embolism/diagnostic imaging , Magnetic Resonance Imaging , Male , Metals , Middle Aged , Retrospective Studies , Tomography, X-Ray Computed , Treatment OutcomeABSTRACT
Metabotropic glutamate 1 (mGlu) receptor has been proposed as a target for the treatment of metastatic melanoma. Studies have demonstrated that inhibiting the release of glutamate (the natural ligand of mGlu1 receptors), results in a decrease of melanoma tumor growth in mGlu1 receptor-expressing melanomas. Here we demonstrate that mGlu1 receptors, which have been previously characterized as oncogenes, also behave like dependence receptors by creating a dependence on glutamate for sustained cell viability. In the mGlu1 receptor-expressing melanoma cell lines SK-MEL-2 (SK2) and SK-MEL-5 (SK5), we show that glutamate is both necessary and sufficient to maintain cell viability, regardless of underlying genetic mutations. Addition of glutamate increased DNA synthesis, whereas removal of glutamate not only suppressed DNA synthesis but also promoted cell death in SK2 and SK5 melanoma cells. Using genetic and pharmacological inhibitors, we established that this effect of glutamate is mediated by the activation of mGlu1 receptors. The stimulatory potential of mGlu1 receptors was further confirmed in vivo in a melanoma cell xenograft model. In this model, subcutaneous injection of SK5 cells with short hairpin RNA-targeted downregulation of mGlu1 receptors resulted in a decrease in the rate of tumor growth relative to control. We also demonstrate for the first time that a selective mGlu1 receptor antagonist JNJ16259685 ((3,4-Dihydro-2H-pyrano[2,3-b]quinolin-7-yl)-(cis-4-methoxycyclohexyl)-methanone) slows SK2 and SK5 melanoma tumor growth in vivo. Taken together, these data suggest that pharmacological inhibition of mGlu1 receptors may be a novel approach for the treatment of metastatic melanoma.
Subject(s)
Cell Survival/physiology , Glutamic Acid/metabolism , Melanoma/metabolism , Receptors, Metabotropic Glutamate/metabolism , Animals , Cell Death/drug effects , Cell Death/physiology , Cell Line, Tumor , Cell Survival/drug effects , Humans , Melanoma/drug therapy , Mice , Mice, Inbred C57BL , Mice, Nude , Quinolines/pharmacologyABSTRACT
OBJECTIVE: To examine whether conservative treatment with oral contraceptives is effective in the shrinkage of a peritoneal inclusion cyst (PIC). This is a case report of two patients with a PIC that developed after gynecological surgery. CASES: Both cases were suspected of a PIC based on the medical history, laboratory data, and image findings. It was difficult in differentiate a PIC from an ovarian tumor. Surgery was chosen at first. However, PICs in both cases recurred after surgery and were treated with oral contraceptives as a conservative treatment. PICs shrank after the treatment of oral contraceptives in both cases. CONCLUSION: Due to the high rate of recurrence following surgery, conservative treatment is recommended to treat PICs. Hormone therapy using oral contraceptives seems to have some therapeutic benefit for the PICs.
Subject(s)
Contraceptives, Oral, Combined/administration & dosage , Contraceptives, Oral, Hormonal/administration & dosage , Cysts/drug therapy , Ethinyl Estradiol/administration & dosage , Levonorgestrel/administration & dosage , Peritoneal Diseases/drug therapy , Adult , Cysts/diagnosis , Cysts/etiology , Cysts/surgery , Drug Combinations , Female , Gynecologic Surgical Procedures/adverse effects , Humans , Middle Aged , Peritoneal Diseases/diagnosis , Peritoneal Diseases/etiology , Peritoneal Diseases/surgery , Recurrence , Sclerotherapy , Tissue Adhesions/surgeryABSTRACT
Because of radioactive fallout resulting from the Fukushima Daiichi Nuclear Power Plant (NPP) accident, water discharge from many outdoor swimming pools in Fukushima was suspended out of concern that radiocesium in the pool water would flow into farmlands. The Japan Atomic Energy Agency has reviewed the existing flocculation method for decontaminating pool water and established a practical decontamination method by demonstrating the process at eight pools in Fukushima. In this method, zeolite powder and a flocculant are used for capturing radiocesium present in pool water. The supernatant is discharged if the radiocesium concentration is less than the targeted level. The radioactive residue is collected and stored in a temporary storage space. Radioactivity concentration in water is measured with a NaI(Tl) or Ge detector installed near the pool. The demonstration results showed that the pool water in which the radiocesium concentration was more than a few hundred Bq L was readily purified by the method, and the radiocesium concentration was reduced to less than 100 Bq L. The ambient dose rates around the temporary storage space were slightly elevated; however, the total increase was up to 30% of the background dose rates when the residue was shielded with sandbags.
Subject(s)
Decontamination/methods , Fukushima Nuclear Accident , Schools , Swimming Pools/standards , Aluminum Hydroxide/chemistry , Cesium Radioisotopes/chemistry , Cesium Radioisotopes/isolation & purification , Decontamination/economics , Flocculation , Radiation Protection , Time Factors , Water/chemistry , Zeolites/chemistryABSTRACT
BACKGROUND AND PURPOSE: Measurement of perfusion parameters is typically done using an intravenous injection of contrast medium. This purpose of this study was to evaluate the feasibility of measuring regional and global CBV using C-arm CT with IA injections of contrast medium. MATERIALS AND METHODS: Twelve canines were studied. CBV measurement was performed using standard PCT, and then using C-arm CT with IV and IA contrast. Values obtained using C-arm CT were compared with those using PCT. RESULTS: C-arm CT CBV maps using IA injections required less contrast than ones with IV injections. PCT and C-arm CT using IV and AA injections provided comparable maps. In controls, C-arm CT with a CCA or VA injection provided comparable maps to PCT. In animals with a stroke, a CCA or VA injection did not provide maps comparable to ones made with PCT. IV and AA C-arm CT showed excellent quantitative agreement with PCT, while CCA and VA C-arm CT studies did not. CONCLUSIONS: Measurement of global CBV using C-arm CT in conjunction with either an IV or an AA injection was feasible in controls and dogs with a stroke. Measurement of regional CBV with C-arm CT using either CCA or VA injection, in normal canines, provided CBV maps qualitatively comparable with those obtained with PCT; the absolute CBV values from these maps were in poor agreement with PCT measurements. Valid measurement of CBV using C-arm CT requires all tissue in a target region to be fully and equally opacified during any acquisition. Using CCA or VA injections, it was impossible to document if and when this had been achieved. CBV measurements using these routes of injection were therefore not reliable.
Subject(s)
Blood Volume Determination/methods , Blood Volume/physiology , Brain Ischemia/physiopathology , Cerebral Arteries/physiology , Cerebrovascular Circulation/physiology , Perfusion Imaging/methods , Tomography, X-Ray Computed/methods , Algorithms , Animals , Brain Ischemia/diagnostic imaging , Contrast Media/administration & dosage , Dogs , Injections, Intra-Arterial , Radiographic Image Interpretation, Computer-Assisted/methods , Reproducibility of Results , Sensitivity and SpecificityABSTRACT
BACKGROUND AND PURPOSE: There is no satisfactory parameter that can predict the need for assistant devices for endovascular aneurysm coiling. Our aim was to evaluate the utility of MOA as a predictor of the need for stent-assisted coiling in ICA sidewall aneurysms. MATERIALS AND METHODS: From a retrospective review of an internal data base, 55 consecutive ICA sidewall aneurysms were identified. Thirty-two of the aneurysms were treated by using endovascular techniques. Because 23 of the 55 aneurysms were either untreated or clipped, 3 experienced interventionalists reviewed the 3D images of these aneurysms and then made a decision as to whether stent-assisted coiling would have been required. Thirty-one of the 55 aneurysms would have required stent-assisted coiling, while 24 would not. Neck width, DNR, AR, and MOA were obtained from each aneurysm by using prototype software. These parameters were then correlated with the requirement of stent-assisted coiling. RESULTS: MOA and neck width of aneurysms requiring stent-assisted coiling were significantly larger than those not requiring stent-assisted coiling (P < .001 and <0.001, respectively). Although the DNR and AR of aneurysms requiring stent-assisted coiling were smaller than those not requiring it, the difference was not significant (P = .22 and 0.12, respectively). ROC analysis revealed that MOA was the parameter that best correlated with the need for stent-assisted coiling. Inclusion of MOA with the rest of the parameters significantly increased the predictive performance regarding the need for stent-assisted coiling (P = .005). CONCLUSIONS: In this small study, MOA was a useful parameter to predict the need for stent-assisted coiling in ICA sidewall aneurysms. Further prospective study of this parameter for aneurysms at multiple locations is required to determine its ultimate value.
Subject(s)
Carotid Artery, Internal, Dissection/diagnostic imaging , Carotid Artery, Internal, Dissection/therapy , Embolization, Therapeutic/methods , Intracranial Aneurysm/diagnostic imaging , Intracranial Aneurysm/therapy , Stents , Adult , Aged , Carotid Artery, Internal/diagnostic imaging , Cerebral Angiography , Cohort Studies , Databases, Factual , Humans , Middle Aged , Predictive Value of Tests , Retrospective StudiesABSTRACT
BACKGROUND AND PURPOSE: Imaging is a key element in the study of many rodent models of human diseases. The application of DSA has been limited in these studies in part because of a lack of a method that allows serial intra-arterial examinations to be performed during an extended period of time. It was our intent to develop and test a method for performing sequential arterial catheterizations and DSA in rats. MATERIALS AND METHODS: Using a transfemoral approach, we subjected 12 adult male Harvey rats to 3 sequential DSA examinations during a 6- to 8-week period. At each examination, 2 selective arterial catheterizations and a DSA were performed. Animals were monitored for ill effects, and images from the 3 examinations were compared for quality and the presence of any arterial injury. RESULTS: Ten of the 12 rats survived all 3 examinations. There were no adverse effects noted and no evidence of arterial injury from the examinations. CONCLUSIONS: With the technique described, it is possible to perform serial arterial catheterizations and DSA in rats. This technique will be useful as an adjunct in the use of rodents for the study of human diseases.
Subject(s)
Angiography, Digital Subtraction/methods , Catheterization/methods , Cerebral Angiography/methods , Angiography, Digital Subtraction/mortality , Animals , Catheterization/mortality , Cerebral Angiography/mortality , Cerebrovascular Circulation , Disease Models, Animal , Femoral Artery , Male , Morbidity , Rats , Rats, Inbred Strains , RodentiaABSTRACT
BACKGROUND AND PURPOSE: CBV is a key parameter in distinguishing penumbra from ischemic core. The purpose of this study was to compare CBV measurements acquired with standard PCT with ones obtained with C-arm CT in a canine stroke model. MATERIALS AND METHODS: Under an institutionally approved protocol, unilateral MCA strokes were created in 10 canines. Four hours later, DWI was used to confirm the presence of an infarct. CBV maps acquired with PCT were compared with ones acquired by using C-arm CT. Three experienced observers, blinded to the technique used for acquisition, evaluated the CBV maps. RESULTS: An ischemic stroke was achieved in 9 of the 10 animals. Areas of reduced CBV were detected in 70%-75% of the PCT studies and in 83%-87% of the C-arm CT examinations, with false-positives in 1.7% and 3.3%, respectively. False-negatives were found in 25% of the PCT and 12.2% of the C-arm CT studies. In all studies, there was a significant difference between the absolute CBV values in normal and abnormal tissue (P < .005) and no significant difference between PCT and C-arm CT CBV values in either the normal or the abnormal parenchyma (P > .05). CONCLUSIONS: CBV measurements made with C-arm CT compare well with ones made with PCT. While further work is required both to fully validate the technique and to define its ultimate clinical value, it appears that it offers a feasible method for assessing CBV in the angiography suite.
Subject(s)
Blood Volume Determination/methods , Brain Ischemia/diagnostic imaging , Cerebrovascular Circulation , Stroke/diagnostic imaging , Tomography, X-Ray Computed/methods , Acute Disease , Animals , Cerebral Angiography/methods , Cerebral Angiography/standards , Disease Models, Animal , Dogs , False Negative Reactions , False Positive Reactions , Tomography, X-Ray Computed/standardsABSTRACT
F(1)-ATPase is a rotary motor enzyme in which a single ATP molecule drives a 120 degrees rotation of the central gamma subunit relative to the surrounding alpha(3)beta(3) ring. Here, we show that the rotation of F(1)-ATPase spontaneously lapses into long (approximately 30 s) pauses during steady-state catalysis. The effects of ADP-Mg and mutation on the pauses, as well as kinetic comparison with bulk-phase catalysis, strongly indicate that the paused enzyme corresponds to the inactive state of F(1)-ATPase previously known as the ADP-Mg inhibited form in which F(1)-ATPase fails to release ADP-Mg from catalytic sites. The pausing position of the gamma subunit deviates from the ATP-waiting position and is most likely the recently found intermediate 90 degrees position.
Subject(s)
Adenosine Triphosphate/chemistry , Proton-Translocating ATPases/chemistry , Adenosine Diphosphate/chemistry , Catalysis , Kinetics , Magnesium/chemistry , Proton-Translocating ATPases/antagonists & inhibitorsABSTRACT
Neuronal nicotinic acetylcholine receptors (nAChRs) play a significant role in sympathetic transmission in the superior cervical ganglia (SCG), with most of the signal carried by a nAChR containing an alpha3 subunit. Work has shown that transection of the postganglionic nerves (axotomy) of the SCG results in a decrease in mRNA transcripts for alpha3, alpha5, alpha7 and beta4 and in protein expression of alpha7 and beta4. To evaluate effects of axotomy on alpha3 protein in the SCG, quantitative immunoblotting was used to demonstrate a dramatic decrease (> 80%) in the levels of this subunit 4 days after axotomy. Similarly, immunocytochemistry showed a marked decline in the number and the intensity of stained neurons for the alpha3 subunit as well as tyrosine hydroxylase. Ganglia explanted into culture for 4 days also showed a substantial decrease in alpha3 subunit protein. This decrease was partially prevented by the addition of nerve growth factor (NGF) to the culture medium at the time of explantation. Additionally, this decrease was reversed by the addition of NGF to the culture medium following 4 days in culture in the absence of NGF. These findings suggest that the loss of alpha3 subunit contributes to the reported decrease in ganglionic synaptic transmission that follows axotomy, and that NGF plays an important role in regulating the expression of alpha3-containing nAChRs in the SCG.
Subject(s)
Axotomy , Nerve Growth Factor/physiology , Neurons/metabolism , Receptors, Nicotinic/metabolism , Superior Cervical Ganglion/metabolism , Animals , Blotting, Western , Culture Techniques , Male , Nerve Growth Factor/pharmacology , Rats , Rats, Sprague-Dawley , Receptors, Nicotinic/drug effects , Reference Values , Superior Cervical Ganglion/drug effects , Sympathetic Nervous System/cytology , Sympathetic Nervous System/metabolismABSTRACT
The ascidian egg is a well-known mosaic egg. In order to investigate the molecular nature of the maternal genetic information stored in the egg, we have prepared cDNAs from the mRNAs in the fertilized eggs of the ascidian, Halocynthia roretzi. The cDNAs of the ascidian embryo were sequenced, and the localization of individual mRNA was examined in staged embryos by whole-mount in situ hybridization. The data obtained were stored in the database MAGEST (http://www.genome.ad.jp/magest) and further analyzed. A total of 4240 cDNA clones were found to represent 2221 gene transcripts, including at least 934 different protein-coding sequences. The mRNA population of the egg consisted of a low prevalence, high complexity sequence set. The majority of the clones were of the rare sequence class, and of these, 42% of the clones showed significant matches with known peptides, mainly consisting of proteins with housekeeping functions such as metabolism and cell division. In addition, we found cDNAs encoding components involved in different signal transduction pathways and cDNAs encoding nucleotide-binding proteins. Large-scale analyses of the distribution of the RNA corresponding to each cDNA in the eight-cell, 110-cell and early tailbud embryos were simultaneously carried out. These analyses revealed that a small fraction of the maternal RNAs were localized in the eight-cell embryo, and that 7.9% of the clones were exclusively maternal, while 40.6% of the maternal clones showed expression in the later stages. This study provides global insights about the genes expressed during early development.
Subject(s)
Gene Expression , Urochordata/genetics , Animals , DNA, Complementary , Expressed Sequence Tags , Humans , Ovum , RNA , Urochordata/embryologyABSTRACT
The enzyme F1-ATPase has been shown to be a rotary motor in which the central gamma-subunit rotates inside the cylinder made of alpha3beta3 subunits. At low ATP concentrations, the motor rotates in discrete 120 degrees steps, consistent with sequential ATP hydrolysis on the three beta-subunits. The mechanism of stepping is unknown. Here we show by high-speed imaging that the 120 degrees step consists of roughly 90 degrees and 30 degrees substeps, each taking only a fraction of a millisecond. ATP binding drives the 90 degrees substep, and the 30 degrees substep is probably driven by release of a hydrolysis product. The two substeps are separated by two reactions of about 1 ms, which together occupy most of the ATP hydrolysis cycle. This scheme probably applies to rotation at full speed ( approximately 130 revolutions per second at saturating ATP) down to occasional stepping at nanomolar ATP concentrations, and supports the binding-change model for ATP synthesis by reverse rotation of F1-ATPase.
Subject(s)
Adenosine Triphosphate/metabolism , Molecular Motor Proteins/metabolism , Proton-Translocating ATPases/metabolism , Catalytic Domain , Hydrolysis , Kinetics , Microscopy , Microspheres , Molecular Motor Proteins/chemistry , Protein Subunits , Proton-Translocating ATPases/chemistry , RotationABSTRACT
The binding change model for the F(1)-ATPase predicts that its rotation is intimately correlated with the changes in the affinities of the three catalytic sites for nucleotides. If so, subtle differences in the nucleotide structure may have pronounced effects on rotation. Here we show by single-molecule imaging that purine nucleotides ATP, GTP, and ITP support rotation but pyrimidine nucleotides UTP and CTP do not, suggesting that the extra ring in purine is indispensable for proper operation of this molecular motor. Although the three purine nucleotides were bound to the enzyme at different rates, all showed similar rotational characteristics: counterclockwise rotation, 120 degrees steps each driven by hydrolysis of one nucleotide molecule, occasional back steps, rotary torque of approximately 40 piconewtons (pN).nm, and mechanical work done in a step of approximately 80 pN.nm. These latter characteristics are likely to be determined by the rotational mechanism built in the protein structure, which purine nucleotides can energize. With ATP and GTP, rotation was observed even when the free energy of hydrolysis was -80 pN.nm/molecule, indicating approximately 100% efficiency. Reconstituted F(o)F(1)-ATPase actively translocated protons by hydrolyzing ATP, GTP, and ITP, but CTP and UTP were not even hydrolyzed. Isolated F(1) very slowly hydrolyzed UTP (but not CTP), suggesting possible uncoupling from rotation.
Subject(s)
Protein Conformation , Proton-Translocating ATPases/metabolism , Purine Nucleotides/metabolism , Pyrimidine Nucleotides/metabolism , Adenosine Triphosphate/metabolism , Catalytic Domain , Guanosine Triphosphate/metabolism , Hydrolysis , Inosine Triphosphate/metabolism , Kinetics , Models, Chemical , Uridine Triphosphate/metabolismABSTRACT
Previous studies have suggested that the localization of the NMDA receptor NR1 subunit may be determined by the splice variant form of NR1 present. Functional studies have also supported selective targeting of NR2A and NR2B to synaptic and extrasynaptic populations, respectively. We set out to determine whether rat cortical and cerebellar NR1 splice variants and NR2 subunits are differentially localized to the postsynaptic density. Using western blot techniques, we measured the percentage of NR1 containing each cassette and the enrichment of the different cassettes and other proteins in the preparations. The results indicate that: (1) no single cassette of NR1 is differentially enriched in the postsynaptic densities and (2) the NR2A and NR2B subunits are similarly enriched at the synapse. The enrichment profiles of postsynaptic density-associated proteins demonstrated similar enrichment levels for postsynaptic density (PSD)-95, the NMDA receptor subunits, chapsyn-110, and the CaMKII alpha subunit. However, synaptophysin, SAP-102, and the GABA(A) receptor beta subunit exhibited lower enrichment levels compared to PSD-95. Additionally, cerebellar but not cortical PSDs exhibited significantly lower enrichment of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) GluR1. Thus, although postsynaptic densities are highly enriched in synaptic proteins, there appears to be no selective incorporation of specific NR1 splice variants or NR2 subunits into this structure.
Subject(s)
Nerve Tissue Proteins/metabolism , Receptors, N-Methyl-D-Aspartate/metabolism , Synapses/metabolism , Alternative Splicing , Animals , Antibody Specificity , Blotting, Western , Calcium-Calmodulin-Dependent Protein Kinase Type 2 , Calcium-Calmodulin-Dependent Protein Kinases/metabolism , Cell Line , Cell Membrane/chemistry , Cell Membrane/metabolism , Cerebellum/chemistry , Cerebellum/metabolism , Cerebral Cortex/chemistry , Cerebral Cortex/metabolism , Disks Large Homolog 4 Protein , Humans , Intracellular Signaling Peptides and Proteins , Kidney/cytology , Kidney/metabolism , Male , Membrane Proteins , Nerve Tissue Proteins/analysis , Organ Specificity , Rats , Rats, Sprague-Dawley , Receptors, N-Methyl-D-Aspartate/analysis , Receptors, N-Methyl-D-Aspartate/genetics , Synapses/chemistry , TransfectionABSTRACT
A synthetic peptide corresponding to the C-terminus of the alpha 3 subunit of the rat neuronal nicotinic acetylcholine receptor (nAChR) was used to generate a rabbit polyclonal alpha 3 antibody. The specificity of this antibody was characterized by immunoblotting, immunohistochemical and immunoprecipitation techniques. Using this antibody, the relative densities of the alpha 3 subunit were quantitatively determined in different brain regions and in superior cervical ganglion (SCG). Among these regions, SCG, interpeduncular nucleus (IPN) and pineal gland showed the highest levels of alpha 3 protein expression. Habenula and superior colliculi had intermediate levels of expression. Low levels were found in cerebral cortex, hippocampus and cerebellum. The ontogenic profile of the alpha 3 subunit in the SCG was also determined. The alpha 3 protein level is low at postnatal day (P 1), but increases rapidly during the first seven postnatal days. This level then plateaus and remains stable through postnatal day 35. These findings suggest that neuronal nAChRs containing the alpha 3 subunit participate in important roles in specific regions of the rat brain and the SCG.
Subject(s)
Brain/metabolism , Neurons/metabolism , Receptors, Nicotinic/biosynthesis , Superior Cervical Ganglion/metabolism , Aging/metabolism , Animals , Antibody Specificity/immunology , Brain/cytology , Cell Line , Female , Habenula/metabolism , Humans , Kidney/cytology , Kidney/metabolism , Mesencephalon/metabolism , Organ Specificity , Pineal Gland/metabolism , Rats , Rats, Sprague-Dawley , Receptors, Nicotinic/analysis , Receptors, Nicotinic/genetics , Receptors, Nicotinic/immunology , Receptors, Nicotinic/metabolism , Superior Cervical Ganglion/cytology , Superior Colliculi/metabolism , TransfectionABSTRACT
In vitro autoradiography using 125I-alpha-bungarotoxin (alpha-BGTx) and anti-alpha7 immunohistochemistry were performed on the dorsal motor nucleus of the vagus (DMV) of sham and chronically vagotomized rats to determine whether the alpha7-nicotinic acetylcholine receptor (nAChR) is located postsynaptically on DMV neurons whose axons contribute to the vagus nerve. Intense bilateral 125I-alpha-BGTx binding and anti-alpha7 immunostaining were observed in coronal brain sections containing the DMV of sham-vagotomized animals. Unilateral cervical vagotomy resulted in ipsilateral losses of 125I-alpha-BGTx binding and anti-alpha7 immunostaining from the DMV. Simultaneous staining of rat brainstem sections with anti-alpha7 and anti-choline acetyltransferase (ChAT) antibodies (to identify cholinergic DMV neurons that project into the vagus nerve) revealed that every DMV neuron that was stained for ChAT showed alpha7-staining as well. In vagotomized animals, no ChAT-positive neurons expressing alpha7-nAChRs remained in the ipsilateral DMV. We conclude that the alpha7-nAChR subtype is located postsynaptically on DMV neurons. To test whether the alpha7-nAChR is similar to the alpha7-homomeric nAChR, experiments were performed in anesthetized rats, and compounds were microinjected into the DMV while monitoring intragastric pressure (IGP). alpha-BGTx and strychnine antagonized nicotine-induced increases in IGP; no antagonism was observed with methyllycaconitine, a compound known to block the homomeric alpha7-nAChR subtype. Recovery from alpha-BGTx-induced antagonism of the nicotine response was observed. We conclude that there is a nAChR containing the alpha7-subunit in the DMV that is different from the homomeric alpha7-nAChR subtype.
Subject(s)
Aconitine/analogs & derivatives , Motor Neurons/metabolism , Receptors, Nicotinic/metabolism , Vagus Nerve/metabolism , Aconitine/pharmacology , Animals , Autoradiography , Bungarotoxins/pharmacokinetics , Bungarotoxins/pharmacology , Fluorescein-5-isothiocyanate , Fluorescent Antibody Technique, Indirect , Male , Microinjections , Nicotinic Agonists/pharmacology , Nicotinic Antagonists/pharmacology , Rats , Rats, Sprague-Dawley , Receptors, Nicotinic/drug effects , Stomach/drug effects , Vagotomy , Vagus Nerve/cytology , alpha7 Nicotinic Acetylcholine ReceptorABSTRACT
Primary culture of rat hepatocyte was performed in an oxygen-permeable film dish (F-dish), which would be expected to give an oxygen-rich culture condition. In the conventional culture dish in which the depth of medium was 2 mm, the oxygen tension (pO(2)) in the medium decreased from 19% (144 mmHg) to 0.3% (2.3 mmHg) within 2 hr, while the pO(2) in the F-dish maintained 8.5% (64.6 mmHg) even after 2 hr. The adverse effect of the oxygen-deficiency appeared in the albumin secretion activity of the hepatocytes and it was more remarkable in the early period of culture. The average rate of albumin secretion for the initial 48 hr was 2.0 mug ml(-1) hr(-1) or 96 mug 10(6) cells(-1) day(-1) in the F-dish. The average rate of albumin secretion for the initial 12 hr was only 0.36 mug ml(-1) hr(-1) in the conventional culture dish. The activity of ammonia elimination in the F-dish was 20-50% higher than the conventional culture dish. Three-dimensional aggregate was formed only in the F-dish. The advantage of three-dimensional aggregate for albumin secretion was not clear compared with two-dimensional monolayer.
ABSTRACT
OBJECTIVE: To determine patterns of methicillin-resistant staphylococci isolated from apparently healthy horses. SAMPLE POPULATION: 44 horses from 8 riding clubs in Japan. PROCEDURE: Methicill in-resistant staphylococci were isolated from the skin or nares, using a selective medium containing a beta-(symboric) lactam antibiotic, ceftizoxime. Clonality of isolates was determined by use of pulsed-field gel electrophoresis. Detection of mecA, mecl, and mecR1 genes was accomplished by use of polymerase chain reactions. RESULT: Of the 44 horses, 13 (29.5%) yielded 15 isolates of methicillin-resistant staphylococci. The 15 isolates were identified as 6 species (Staphylococcus epidermidis, S lentus, S saprophyticus, S xylosus, S sciuri, and S haemolyticus). However, methicillin-resistant S aureus was seldom isolated. Each isolate contained the mecA gene and had a high resistance to beta-lactam antibiotics. Some isolates also were resistant to other antibiotics such as erythromycin and kanamycin. CONCLUSIONS AND CLINICAL RELEVANCE: Methicillin-resistant coagulase-negative staphylococci that were highly resistant to various antibiotics were isolated from apparently healthy horses in Japan. These organisms must be considered a potential threat to horses and veterinarians who care for them.
Subject(s)
Coagulase/analysis , Hexosyltransferases , Horses/microbiology , Methicillin Resistance , Peptidyl Transferases , Staphylococcus/isolation & purification , Animals , Bacterial Proteins/genetics , Bacteriological Techniques/methods , Bacteriological Techniques/veterinary , Carrier Proteins/genetics , DNA Primers , Electrophoresis, Agar Gel/veterinary , Electrophoresis, Gel, Pulsed-Field/veterinary , Japan , Microbial Sensitivity Tests/veterinary , Muramoylpentapeptide Carboxypeptidase/genetics , Penicillin-Binding Proteins , Polymerase Chain Reaction/veterinary , Repressor Proteins/genetics , Skin/microbiology , Staphylococcus/drug effects , Staphylococcus/enzymologyABSTRACT
The effect of post-mortem delay on the stability of the protein subunits that combine to form NMDA and AMPA type glutamate receptors has been assessed in samples of human brain tissue. While most of the subunits (i.e. GluR1, GluR2/3, GluR4, NR1) appear to be stable for up to 18 h post-mortem, the NR2A and NR2B subunits appear to be proteolyzed rapidly following death. These results are consistent with the concept that the proteolytic products of NR2A and NR2B, although at smaller molecular sizes than the full-length protein, are all identifiable on Western blots. Thus, a method is proposed that allows for the estimation of the levels of these labile proteins even in samples obtained up to 18 h post-mortem. Using this method we have estimated the levels of all AMPA and NMDA receptor subunits in selected (i.e. hippocampus, frontal and entorhinal cortex) brain tissue samples obtained from control patients and patients who have died with Alzheimer's disease. Modest decreases in NMDA receptor subunits NR1, NR2A, and NR2B were found in the hippocampus and in frontal cortex while little or no change in any of these subunits were documented in entorhinal cortex. Subunits for AMPA receptors (GluR1, GluR2/3, and GluR4) appeared to show a generalized decrease in all these tissues. As a surrogate marker for overall decreases due to generalized neuronal cell death, levels of neuron-specific enolase were measured in all tissues and were found to be nearly identical in control and Alzheimer's brains.
Subject(s)
Alzheimer Disease/pathology , Brain Chemistry , Brain/pathology , Receptors, AMPA/analysis , Receptors, N-Methyl-D-Aspartate/analysis , Aged , Antibody Specificity , Blotting, Western , Female , Humans , Male , Middle Aged , Receptors, AMPA/immunology , Receptors, N-Methyl-D-Aspartate/immunology , Time FactorsABSTRACT
To determine the genetic causes and molecular mechanisms responsible for neurobehavioral differences in mice, we used highly parallel gene expression profiling to detect genes that are differentially expressed between the 129SvEv and C57BL/6 mouse strains at baseline and in response to seizure. In addition, we identified genes that are differentially expressed in specific brain regions. We found that approximately 1% of expressed genes are differentially expressed between strains in at least one region of the brain and that the gene expression response to seizure is significantly different between the two inbred strains. The results lead to the identification of differences in gene expression that may account for distinct phenotypes in inbred strains and the unique functions of specific brain regions.