Biomedical subjects

T Bruhn

Publications and source records attributed to T Bruhn.

At least 19 recordsLinked to original sources

PubMed2002

Thrombin as a survival factor for cancer cells: thrombin activation in malignant effusions in vivo and inhibition of idarubicin-induced cell death in vitro.

OBJECTIVES: The aim of the experiments shown here, is to demonstrate exemplarily that thrombin can be a survival factor for malignant cells. METHODS: Activation of the coagulation system has been examined in patients with acute myeloid leukemia (AML) and non-Hodgkin lymphoma (NHL) before and after chemotherapy as well as in malignant effusions of heavily pretreated patients with solid tumors. Thrombin receptor expression (PAR-I) has been examined on HL-60 cells; the effect ofthrombin on the proliferation of the cells and inhibition of apoptosis induction by idarubicin has been shown. RESULTS: Using fibrinopeptide A as an indirect parameter for thrombin activation, we found elevated levels in patients with AML and NHL before and a significant 2-fold increase after chemotherapy (p < 0.02 for the AML group; p < 0.0006 for the NHL group). Apparently, this does not only affect patients with hematological diseases, but also with solid tumors. In order to find out if the tumor cells directly activate thrombin, we examined malignant effusions of patients with different solid tumors. Comparing prothrombin fragment 1 + 2 in ascites and pleural effusions with the patients' serum levels, we found it significantly increased in all cases (mean of 1.96 +/- 0.5 nmol/l in the serum vs. 12.1 +/- 3.6 nmol/l in effusions; p < 0.001). The majority of patients presented elevated serum levels. Additionally, we incubated HL-60 cells (human promyelocytic leukemia) with thrombin prior to treatment with idarubicin. Expression of thrombin receptor (PAR-1) could be verified by FACS-analysis using a monoclonal antibody. HL-60 cells responded with increased proliferation to thrombin exposure with concentrations between 0.3 and 3 U/ml. This effect could be abolished by the addition of hirudin, demonstrating thrombin specificity. In these concentrations, thrombin was able to abrogate the induction of apoptosis by idarubicin completely (p < 0.005). CONCLUSIONS: Here we give evidence for the role of thrombin as a resistance factor for tumor cells towards chemotherapy. In the light of the fact that thrombin is regularly activated in cancer patients, these findings indicate that thrombin is a clinically relevant cellular resistance factor. A number of pre-clinical and clinical studies imply that inhibition of the coagulation system, e.g. by low-molecular weight heparins or warfarin, increases the effect of chemotherapy.

Apoptosis↗

PubMed2001

In vivo cellular uptake of glutamate is impaired in the rat hippocampus during and after transient cerebral ischemia: a microdialysis extraction study.

Using microdialysis in CA1 of the rat hippocampus, we studied the effect of transient cerebral ischemia on in vivo uptake and on extracellular levels of glutamate during, and at different time points after ischemia. (3)H-D-aspartate (test substance), and (14)C-mannitol (reference substance), were added to the dialysis perfusate, and the cellular extraction of (3)H-D-aspartate was calculated from scintillation analysis of fractionated dialysate samples. The extraction of (3)H-D-aspartate was studied both in a tracer like condition with a perfusate concentration of 0.2 microM, and in a condition of high saturation level, with 1.0 mM D-aspartate added to the perfusate. In between radioisotope perfusions, dialysate was sampled for analysis of amino acid content by HPLC. During ischemia, extraction of (3)H-D-aspartate (0.2 microM) declined to a maximum reduction of 68%. In the hours after ischemia, extraction of (3)H-D-aspartate (0.2 microM) was decreased by 32%. In the days after ischemia, there was a progressive decline in extraction of (3)H-D-aspartate (1.0 mM), reaching a reduction of 89% on Day 4 after ischemia. Extracellular glutamate remained at control levels at all time points after ischemia. The present study is the first to investigate uptake of glutamate in the intact rat brain in relation to cerebral ischemia. Evidence is provided that uptake of Glu is restrained during ischemia, and that in the hours after ischemia, the extracellular turnover of glutamate is decreased. In the course of the days after ischemia, degeneration of CA1 pyramidal cells occurs concomitantly with a progressive decline in glutamate transport ability, possibly of pathogenetic importance to CA1 pyramidal cell loss.

Animals↗

PubMed2001

Isolation and characterisation of five neurotoxic and cardiotoxic polypeptides from the sea anemone Anthopleura elegantissima.

Five toxins (APE 1 to APE 5) of the sea anemone species Anthopleura elegantissima (Brandt) have been isolated from a toxic by-product fraction of its concentrated crude watery-methanolic extract, prepared previously for the isolation of a neuropeptide (the head-activator) by Schaller and Bodenmüller (Proc. Natl. Acad. Sci. USA 78 (1981) 7000) from 200kg sea anemones. Toxin purification was performed by desalting of the starting material by dialysis (MWCO 3500) against distilled water, anion exchange chromatography on QAE-Sephadex A25 at pH 8, twice gel filtration on Sephadex G50 m, repeated chromatography on QAE-Sephadex at pH 10 and chromatography on the cation exchanger Fractogel EMD SO(3)(-)-650 M.Final purification of the toxins was achieved by HPLC on MN SP 250/10 Nucleosil 500-5 C(18) PPN and MN SP 250/21 Nucleosil 300-7 C(18). Each toxin was composed of at least two isotoxins of which APE 1-1, APE 1-2, APE 2-1, APE 2-2 and APE 5-3 were isolated in preparative scale. With exception of APE 5-3 the sequences of the isotoxins have been elucidated. They resemble the 47 residue type-I long polypeptide toxins native to Anemonia sulcata (Pennant). All isotoxins paralyse the shore crab (Carcinus maenas) by tetanic contractions after i.m. application. The toxins modify current passing through the fast Na(+) channel in neuroblastoma cells, leading to delayed and incomplete inactivation. APE 1-1, APE 2-1 and APE 5-3 produce a positive inotropic effect in mammalian heart muscle, although they differ in potency. The order of potency is APE 2-1>APE 1-1>APE 5-3 (i.e. threshold concentrations are 1, 10 and 300nM, respectively). In addition, they enhance the spontaneous beating frequency in isolated right atria (guinea pig). The most potent cardiotoxic isotoxin is APE 2-1, its sequence is identical with that of AP-C, a toxin isolated and characterised previously by Norton et al. (Drugs and Foods from the Sea, 1978, University of Oklahoma Press, p. 37-50).LD50 APE 2-1:1 micro g/kg b.w. C. maenas (i.m.). LD50 APE 1-1:10 microg/kg b.w. C. maenas (i. m.). LD50 APE 5-3:50 microg/kg b.w. C. maenas (i.m.).

Animals↗

PubMed2000

The Microwave Spectrum of m-Tolunitrile: Methyl Internal Rotation and (14)N Nuclear Quadrupole Coupling.

The microwave spectrum of m-tolunitrile (3-methylbenzonitrile, m-C(6)H(4)CH(3)CN) has been investigated in the frequency range from 1 to 4 and 8 to 26.5 GHz. The spectra in the two lowest states of internal methyl rotation (m = 0, +/-1) were recorded by means of pulsed molecular beam Fourier transform microwave (MB-FTMW) spectrometers. The interpretation of the spectra was based on an asymmetric frame-symmetric top Hamiltonian with inclusion of centrifugal distortion terms and first-order contributions from (14)N nuclear quadrupole coupling. A least-squares analysis yielded the rotational constants A = 3295.9103(10) MHz, B = 1199.1188(2) MHz, C = 883.9223(1) MHz, all elements of the nuclear quadrupole coupling tensor chi(aa) = -3.626(1) MHz, chi(bb) = 1.684(1) MHz, chi(cc) = 1.943(1) MHz, and chi(ab) = -1.870(3) MHz, as well as the threefold barrier to internal rotation, V(3) = 14.2 cm(-1), and the angle between the internal rotor axis and the principal moment of inertia a axis, θ = 42.66 degrees, using fixed values for the sixfold barrier term V(6) (-11 cm(-1)) and the moment of inertia of the methyl top I(alpha) (3.16 u Å(2)). Copyright 2000 Academic Press.

Journal Article↗

PubMed2000

The Microwave Spectrum of 1-Chloro-2-methylpropene.

The rotational spectrum of 1-chloro-2-methylpropene has been measured with very high resolution using pulsed molecular-beam Fourier transform microwave (MB-FTMW) spectrometers in the range from 3 to 40 GHz. The analysis yielded accurate rotational constants, centrifugal distortion constants, the complete (35)Cl and (37)Cl quadrupole coupling tensors, and the V(3) hindering barriers of both methyl rotors. These results supplement and improve previous observations made with waveguide microwave and infrared spectroscopy under considerably lower resolution. Moreover, a potential surface depending on the two angles of internal rotation was calculated using ab initio methods. This enabled us to compare theoretical and experimental potential barriers and to obtain information on higher potential terms and top-top interaction constants, which were not available from the experimental data alone. Copyright 2000 Academic Press.

Journal Article↗

PubMed1999

Secretory phospholipase A2 potentiates glutamate-induced rat striatal neuronal cell death in vivo.

The secretory phospholipases A2 (sPLA2) OS2 (10, 20 and 50 pmol) or OS1, (50 pmol) purified from taipan snake Oxyuranus scutellatus scutellatus venom, and the excitatory amino acid glutamate (Glu) (2.5 and 5.0 micromol) were injected into the right striatum of male Wistar rats. Injection of 10 and 20 pmol OS2 caused no neurological abnormalities or tissue damage. OS2 (50 pmol) caused apathy and circling towards the injection side. Histology revealed an infarct at the injection site. Injection of 50 pmol OS1 showed very little or no signs of neurotoxicity. Injection of 2.5 micromol Glu caused no tissue damage or neurological abnormality. After injection of 5.0 micromol Glu, the animals initially circled towards the side of injection, and gradually developed generalized clonic convulsions. These animals showed a well demarcated striatal infarct. When non-toxic concentrations of 20 pmol OS2 and 2.5 micromol Glu were co-injected, a synergistic neurotoxicity was observed. Extensive histological damage occurred in the entire right hemisphere, and in several rats comprising part of the contralateral hemisphere. These animals were apathetic in the immediate hours following injection, with circling towards the side of injection in the following days. Thus, OS2 greatly potentiates glutamate excitoxicity in vivo.

Animals↗

PubMed1999

Double-tracer autoradiographic study of protein synthesis and glucose consumption in rats with focal cerebral ischemia.

A double-tracer autoradiographic method for simultaneous measurement of regional glucose utilization (rCMRglc) and regional protein synthesis (PS) in consecutive brain sections is described and applied to study the metabolism of the ischemic penumbra 2 h after occlusion of the middle cerebral artery (MCAO) in rats. In halothane anesthesia, the left middle cerebral artery was permanently occluded. Two hours after MCAO an i.v. bolus injection of 14C-deoxyglucose and 3H-leucine was given and circulated for 45 min. Two sets of brain sections were processed for quantitative autoradiography. Neighboring brain sections exposed an X-ray film (3H-insensitive), and a 3H-sensitive for determination of rCMRglc and PS, respectively. Sections for PS determination were washed in trichloroacetic acid (TCA) prior to film exposure in order to remove 14C-deoxyglucose and unincorporated 3H-leucine. Regional rates of PS and glucose utilization were measured by densitometric image analysis. Normal rates of metabolism were defined as mean +/- 2 SD of values in the non-ischemic cortex. The volumes of ischemic cortex displaying normal rates of PS and glucose utilization, respectively, were measured. The cortical volume with normal PS was significantly less than that of normal rCMRglc: 142 (127-147) mm3 vs. 203 (184-206) mm3. Treatment with the glutamate antagonists MK-801 (1 mg kg-1) and NBQX (30 mg kg-1 x 2) did not significantly change this, although MK-801 tended to reduce the size of the metabolic penumbra calculated as the difference between ischemic cortex with reduced PS and ischemic cortex with reduced rCMRglc.

Animals↗

PubMed1997

Anticoagulant fucoidan fractions from Fucus vesiculosus induce platelet activation in vitro.

Anticoagulant fucoidan fractions of different molecular weight and sulfate content were prepared and investigated for their effects on platelet function in vitro. The fucoidan fractions were incubated with human platelet rich plasma (PRP) at concentrations of 5, 10 and 50 micrograms/ml. Platelet activation was subsequently studied by a standard aggregation assay and flow cytometric determination of the activation dependent platelet-surface markers CD62p (P-selectin, GMP-140) and CD63 (GP53). All fucoidan fractions induced irreversible platelet aggregation in a dose-dependent manner. Comparing fractions of identical molecular weight (100 kDa) the low sulfate content fucoidan FF5 (S = 7.6%) exerted a significantly greater effect than the highly sulfated fucoidan FF7 (S = 10.2%) over the whole concentration range (n = 5, P < 0.05). Among fractions of identical sulfate content fucoidan-induced platelet aggregation was also found to depend on the molecular weight of the fucoidan. At concentrations of 10 and 50 micrograms/ml the high molecular weight fraction FF7/1 (150 kDa) showed a significantly greater effect than the 50 kDa fraction FF7/3 (24.8 +/- 6.7 vs. 7.0 +/- 3.5 and 54.6 +/- 13.5 vs. 15.0 +/- 9.0%, respectively; mean +/- SD, n = 5, P < 0.05). The molecular weight dependence of the fucoidan effect was also reflected by the flow cytometric data. Coincubation of FF7/1 and FF7/3 (10 micrograms/ml) with PRP increased the number of CD62p and CD63 positive platelets by 9.0 +/- 3.3 vs. 2 +/- 1.9 and 7.1 +/- 2.4 vs. 3.2 +/- 2.6% over control values, respectively (n = 5, P < 0.05). In conclusion, our results show that the low molecular weight fucoidan FF7/3 combines potent anticoagulant and fibrinolytic properties with only minor platelet activating effects and is therefore a suitable substance for further pharmacological studies.

Anticoagulants↗

PubMed1997

Evidence for increased cellular uptake of glutamate and aspartate in the rat hippocampus during kainic acid seizures. A microdialysis study using the "indicator diffusion' method.

Using a newly developed technique, based on microdialysis, which allows cellular uptake of glutamate and aspartate to be studied in awake animals, we investigated uptake of glutamate and aspartate in the hippocampal formation of rats during limbic seizures induced by systemical administration of kainic acid (KA). With [14C]mannitol as an extracellular reference substance, the cellular extraction of the test substance [3H]D-aspartate was measured at different stages of seizure-activity. The results were compared to those obtained in a sham operated control group. During severe generalized clonic seizures, the extraction of [3H]D-aspartate was increased by 17%. The increase in uptake of [3H]D-aspartate was accompanied by a 24% increase in the extracellular level of aspartate, as obtained by conventional microdialysis. No significant changes were observed in the extracellular level of glutamate. The results indicate that during KA-induced seizures, uptake of glutamate and aspartate is increased, possibly aimed at maintaining the extracellular homeostasis of these two excitatory amino acids.

Animals↗

PubMed1997

Effects of the AMPA-receptor antagonist, NBQX, on neuron loss in dentate hilus of the hippocampal formation after 8, 10, or 12 min of cerebral ischemia in the rat.

The alpha-amino-3-hydroxy-5-methyl-4-isoxazole (AMPA) receptor antagonist, 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline (NBQX), offers protection to hippocampal CA1 pyramidal cells after short episodes of transient cerebral ischemia. Besides CA1 pyramidal cells, neurons containing somatostatin (SS) and located in the dentate hilus of the hippocampal formation are lost after cerebral ischemia. We studied the protective effects of NBQX on SS neurons in the hilus and on hippocampal CA1 pyramidal cells following 8, 10, or 12 min of four-vessel occlusion ischemia during systemic hypotension. NBQX was administered 3 x 30 mg/kg at 0, 10, and 25 after induction of ischemia or sham, and all rats survived for 7 days. NBQX given to control rats without ischemia had no influence on number or morphology of hilar SS neurons and CA1 pyramidal cells. After 8 min of ischemia, NBQX prevented loss of hilar SS neurons. After 10 and 12 min of ischemia, NBQX had no significant effects on loss of SS neurons in the dentate hilus. However, in all ischemic groups, NBQX significantly reduced loss of CA1 pyramidal cells as compared to control rats. This neuroprotective effect decreased gradually and significantly as the time of ischemia increased. Our results support the observation that SS neurons in hilus are among the most ischemia-vulnerable neurons in the brain. We found that administration of NBQX in generally accepted dosages can protect the rapidly dying SS neurons in hilus from only brief episodes of ischemia.

Animals↗

PubMed1996

Effects of phenylsuccinate on potassium-stimulated taurine release in cultured neurons and astrocytes and in rat hippocampus in vivo.

Swelling-induced release of taurine was investigated in vivo in hippocampus by microdialysis or in vitro in cultured neocortical neurons or astrocytes. Swelling was induced either by increasing the extracellular K+ concentration or by exposing the cells to hyposmotic conditions. It was found that the drug phenylsuccinate, which inhibits the mitochondrial dicarboxylate carrier as well as biosynthesis of neurotransmitter glutamate, inhibits swelling-induced taurine release both in vivo and in cultured cells. Thus, phenylsuccinate may be used to investigate the mechanism involved in taurine release associated with regulatory volume decrease in cells.

Amino Acids↗

PubMed1996

Selective depletion of spinal monoamines changes the rat soleus EMG from a tonic to a more phasic pattern.

1. To assess the role of descending monoaminergic pathways for motor activity long-lasting EMG recordings were performed from the adult soleus muscle before and after selective depletion of spinal monoamines. 2. Rats were chronically implanted with an intrathecal catheter placed in the lumbar subarachnoid space and gross-EMG recording electrodes in the soleus muscle. EMG recordings were performed in control conditions and at different times after intrathecal administration of either 40-55 micrograms 5,6-dihydroxytryptamine (5,6-DHT) and 40-55 micrograms 6-hydroxydopamine (6-OHDA) or 80 micrograms 5,7-dihydroxytryptamine (5,7-DHT) alone. The depletions were evaluated biochemically in brains and spinal cords after recordings. 3. In agreement with previous studies the intrathecal administration of neurotoxins caused a reduction of the noradrenaline (NA) and serotonin (5-HT) content of the lumbar spinal cord to about 2-3% of control, with little or no changes in the monoamine content of the cortex. 4. In non-treated chronically catheterized rats the integrated rectified gross EMG displayed long-lasting EMG episodes composed of phasic high-amplitude events and tonic segments of varying duration and amplitude. 5. After intrathecal administration of neurotoxins the number of long-lasting gross-EMG episodes, the mean episode duration, and the total EMG activity per 24 h, were reduced. These changes were accompanied by a simultaneous increase both in the number of short-lasting EMG episodes and the total number of EMG episodes per 24 h period. The changes were apparent 5-6 days after drug administration and fully developed after 2-3 weeks. 6. No changes in general movement ability were observed, except that the denervated animals had a tendency to a less errect posture. 7. These results indicate that descending monoaminergic pathways are important for the maintained motor output in tonic hindlimb muscles.

Animals↗

PubMed1996

Extended studies on the effect of glutamate antagonists on ischemic CA-1 damage.

Glutamate receptors are numerous on the ischemia vulnerable CA-1 pyramidal cells. Postischemic use of the AMPA antagonist NBQX has shown up to 80% protection against cell death. Three aspects of this were studied: In the first study, male Wistar rats were given NBQX (30 mg/kg x 3) either 20 hours or immediately (0 h) before 12 min of 4-vessel occlusion with hypotension. After six days of reperfusion comparison with an untreated group showed almost full protection in the 0 h group (4% cell loss, p < 0.001) but only slight protection in the 20 h group (62% cell loss, p < 0.05). After 12 min of ischemia in the present model, eosinophilic CA-1 cells are seen from day 2 on. Since there could be a late, deleterious calcium influx via NMDA receptors, one group of ischemic rats was given MK-801 (5 mg/kg i.p.) 24 hours after ischemia. However, quantitation 6 days later of remaining CA-1 cells showed no protection. In the third study referred here, two groups of ischemic rats were given NBQX (30 mg/kg x 3) immediately after ischemia. The groups survive for six and 21 days, respectively. Counting of CA-1 pyramidal cells showed an equal, significant protection in both groups (approx 20% cell loss).

Animals↗

PubMed1995

Kalicludines and kaliseptine. Two different classes of sea anemone toxins for voltage sensitive K+ channels.

New peptides have been isolated from the sea anemone Anemonia sulcata which inhibit competitively the binding of 125I-dendrotoxin I (a classical ligand for K+ channel) to rat brain membranes and behave as blockers of voltage-sensitive K+ channels. Sea anemone kalicludines are 58-59-amino acid peptides cross-linked with three disulfide bridges. They are structurally homologous both to dendrotoxins which are snake venom toxins and to the basic pancreatic trypsin inhibitor (Kunitz inhibitor) and have the unique property of expressing both the function of dendrotoxins in blocking voltage-sensitive K+ channels and the function of the Kunitz inhibitor in inhibiting trypsin. Kaliseptine is another structural class of peptide comprising 36 amino acids with no sequence homology with kalicludines or with dendrotoxins. In spite of this structural difference, it binds to the same receptor site as dendrotoxin and kalicludines and is as efficient as a K+ channel inhibitor as the most potent kalicludine.

Amino Acid Sequence↗

PubMed1995

Kainic acid-induced seizures and brain damage in the rat: role of calcium homeostasis.

Seizure activity induced by kainic acid (KA) and subsequent neuronal death are thought to be associated with an increase in cytoplasmic free calcium ([Ca2+]i) and can be prevented by N-methyl-D-aspartate (NMDA) antagonists. In addition to influx through receptor operated Ca2+ channels the increase in [Ca2+]i may be the result of an increased influx through voltage-operated calcium channels and/or release from intracellular deposits. It was therefore investigated whether compounds other than NMDA antagonists with known actions on the intracellular Ca2+ homeostasis had any protective effect against KA-induced neuronal death. Voltage-operated calcium channels in the cell membrane were blocked with the L-type ion channel antagonist, Nimodipine (1.0 mg/kg), and release of Ca2+ from internal stores was prevented with Dantrolene (10 mg/kg). Animals from two control groups injected with kainate (8 mg/kg) exhibited a survival rate of 67 and 53%, respectively. Countings of neurons in dorsal hippocampus showed subtotal or total loss in the CA1 and CA3 subregions. There were no significant differences concerning seizure and survival rates in the groups injected with kainate and treated with Dantrolene or Nimodipine and the control groups. The group treated with Dantrolene showed no neuropathological changes in the hippocampal CA3 region and only slight changes in the Ca1 region, while the neuron loss in the Nimodipine group did not differ from that of its control group. The results emphasize the importance of Dantrolene-sensitive Ca2+ release from intracellular stores for the development of seizure-induced neuronal death.

Animals↗

PubMed1995

Microdialysis as a tool for in vivo investigation of glutamate transport capacity in rat brain.

The role of glutamate as a possible mediator of neurodegeneration is well described, and the homeostasis of extracellular glutamate is considered of major importance when addressing the pathogenesis of excitatory neurodegeneration. Applying the 'indicator diffusion' method to the microdialysis technique, we present a method that is suitable for the in vivo investigation of the capacity of cellular uptake of glutamate. Using 14C-mannitol as reference, we measured the cellular extraction and the cell membrane permeability of the test substance 3H-D-aspartate in the corpus striatum of the rat brain. The cellular extraction fraction of 3H-D-aspartate was 0.29, and the cell membrane permeability 2.24 x 10(-4) cm/s. In the presence of the glutamate-uptake blocker DL-threo-beta-hydroxyaspartate (THA) the extraction of 3H-D-aspartate was completely abolished, indicating that extraction of 3H-D-aspartate was due to cellular uptake by glutamate transporters. The cell membrane permeability towards 3H-D-aspartate was reduced by approximately 98% due to THA, indicating that the cell membranes per se are highly resistant to diffusion of 3H-D-aspartate. It is concluded that the present method can be used in studying the capacity of the glutamate transporters in vivo.

ATP-Binding Cassette Transporters↗

PubMed1995

Glucocorticoids stimulate thyrotropin-releasing hormone gene expression in cultured hypothalamic neurons.

Although there is much evidence indicating that glucocorticoids (GC) inhibit the hypothalamic-pituitary-thyroid axis in both rat and man in vivo, there have been no previous studies on the direct effect of GC on hypothalamic TRH neurons in vitro. In this laboratory, we developed fetal rat (day 17) diencephalic neuronal cultures in the presence of 5'-bromo-2-deoxyuridine, a cell-differentiating agent that stimulates TRH gene expression. In 12 separate experiments, dexamethasone (Dex) induced a 2.2-fold increase in TRH content vs. the control value (P < 0.01). Dex (10(-8)M) enhanced TRH messenger RNA (mRNA) 1.6-fold (n = 75 wells; P < 0.01) by nonisotopic in situ hybridization. On Northern blot analysis using a 32P-labeled complementary RNA probe, TRH mRNA was enhanced 3-fold (n = 4; P < 0.01). Nuclear run-on analysis revealed that Dex enhanced transcription 7.7 fold (n = 3; P < 0.01). We conclude that 1) Dex stimulates the expression of TRH peptide and TRH mRNA in cultured hypothalamic neurons; 2) the increase in TRH mRNA results (at least in part) from enhanced transcription; and 3) the reported in vivo depression of TRH in the paraventricular nucleus after GC stimulation suggests that this effect must be mediated indirectly on the TRH neuron.

Animals↗

PubMed1994

Evidence for formation of hydroxyl radicals during reperfusion after global cerebral ischaemia in rats using salicylate trapping and microdialysis.

Systemic administration of salicylate (SA) to rats (100 mg kg-1 i.p. ) was used as an in vivo trap of hydroxyl radicals (.OH). In the brain SA reacts with hydroxyl radicals to form the stable adducts 2, 3- and 2,5 dihydroxybenzoic acid (DHBAs) which can thus be taken as an index of .OH formation. The DHBAs were recovered by intracerebral microdialysis in hippocampus or striatum and quantified by high pressure liquid chromatography (HPLC) with electrochemical detection. There were no peaks corresponding to 2,5-DHBA or 2,3-DHBA in the chromatograms from rats not receiving SA. A basal level of 2,5-DHBA was seen in the dialysates from all animals given SA whereas 2, 3-DHBA was not detected. In one group of rats generation of free oxygen radicals was induced in the striatum by adding Fe2+ and ascorbate to the perfusion fluid to test the sensitivity of the system. Addition of Fe2+ ascorbate to the perfusion fluid induced a significant 7-fold increase in 2,5-DHBA that gradually returned to baseline after removal of Fe2+/ascorbate. In two other groups the microdialysis probes were implanted in either the striatum or the hippocampus and the animals were subjected to 20 min of four-vessel occlusion + hypotension (4-VOH). Significant reductions in 2,5-DHBA were detected during ischaemia followed by significant increases of 5-fold and 3-fold in the striatum and hippocampus, respectively, beginning immediately upon reperfusion and lasting for the remainder of the observation period (160 min).

Animals↗