F. Block

Evaluation of R6/2 HD transgenic mice for therapeutic studies in Huntington's disease: behavioral testing and impact of diabetes mellitus.

R6/2 transgenic mice express exon 1 of the human Huntingtons disease (HD) gene with an increased CAG repeat length. They develop a progressive neurological phenotype, die within 12-14 weeks of age and were also found to develop diabetes mellitus. Since R6/2 mice are broadly used to screen for potential therapies in HD, the aim of this study was (a) to search for behavioral tests that are best applicable to monitor the behavioral abnormalities in therapy studies and (b) to investigate the extent to which diabetes influences the disease phenotype. We found that the rotarod test for motor coordination and the open field test for spontaneous explorative behavior were useful to monitor the progressive behavioral deterioration of R6/2 mice. An accelerating rotarod paradigm was superior over testing with a rotarod at various fixed speeds since it leads to similar results with less repetitive daily trials so that exhaustion cannot contribute substantially to their decline in performance. With the Morris water maze, however, it was only possible to monitor cognitive decline in visuo-spatial learning in the first weeks of disease since, at later stages, mice were not able to learn the task adequately. A latent diabetes mellitus was found in all transgenic mice demonstrated by a pathological glucose tolerance. Only 26% of the mice, however, were found to develop a manifest diabetes with increasing blood glucose levels on normal diet over the disease period. R6/2 mice with manifest and latent diabetes showed no significant differences in survival, weight loss, motor coordination, or spontaneous explorative behavior. These results suggest that diabetes mellitus is not a major contributing factor to the disease phenotype.

Time course of glial proliferation and glial apoptosis following excitotoxic CNS injury.

Activation of microglial cells and astrocytes after CNS injury results in changes in their morphology, immunophenotype and proliferative activity and has neurotrophic as well as neurotoxic consequences. However, little is known about the exact time course of glial activation as regards their proliferative activity and their fate. In this study, quantification of the densities of proliferating and non-proliferating microglial cells and astrocytes was carried out over 30 days by counting differentially labeled cells in the striatum and substantia nigra pars reticulata (SNr) after injection of quinolinic acid into the rat striatum. The TdT-mediated dUTP nick end labeling (TUNEL)-reaction was used to detect possible apoptotic mechanisms which limit the glial reaction. At 1 day post injection (p.i.) non-proliferating ameboid microglia/macrophages were seen in the striatum, but at 3 and 5 days p.i. many proliferating, ameboid microglia/macrophages and hypertrophic microglia were detected. At 10 days p.i., the time point with the highest density of hypertrophic microglia, TUNEL-positive microglial cells were observed indicating that apoptotic processes play a role in restricting this reaction. In contrast to this, at early time points, a reduction in the density and glial fibrillary acidic protein (GFAP)-immunoreactivity of astrocytes in the striatum was detected. At later time points, a dense astrogliosis with proliferating astrocytes developed in the dorsal and medial striatum. At 30 days p.i., in the entire striatum a dense astrogliosis was detected. The SNr showed a short period of microglial activation and proliferation and a long lasting astrogliosis without proliferation

Focal ischemia induces transient expression of IL-6 in the substantia nigra pars reticulata

We examined the expression of IL-6 in the substantia nigra pars reticulata (SNr) at various time points after transient (3 h) middle cerebral artery occlusion (MCAO) in rats. The animals were killed at 1, 3, 7 or 14 days following operation. Coronal brain sections were processed for immunohistochemistry with antibodies against GFAP, OX-42 and IL-6 and for Nissl staining. Microglial activation was detected 3 and 7 days after ischemia. Reactive astrocytes have been found 7 and 14 days after ischemia. IL-6 expression was detected 3 and 7 days after ischemia. IL-6-positive cells beared the typical morphology of neurons. Distribution of IL-6-positive cells within the SNr was not homogenous. The lateral area of the SNr bears the highest number of IL-6-positive neurons while the central core bears the lowest. Quantification of intact neurons in the SNr 14 days after reperfusion shows that the highest amount of cell loss was found in the central core of the SNr and less neuronal cell loss was observed in the lateral area of the SNr. Thus, the SNr area with the highest IL-6 expression 3 and 7 days after ischemia bears the highest number of intact neurons 14 days after ischemia. This finding could be a clue for the neuroprotective role of IL-6 in the remote region SNr after focal cerebral ischemia.

Immunohistochemical evidence for flupirtine acting as an antagonist on theN-methyl-d-aspartate and homocysteic acid-induced release of GABA in the rabbit retina

When rabbit retinas are exposed in vitro to specific excitatory amino acid receptor agonists certain GABAergic amacrine cells are activated to cause a release of GABA. The GABA that is not released can be detected by immunohistochemistry. Exposure of tissues to kainate or NMDA each caused a characteristic change in the GABA immunoreactivity. CNQX antagonised the kainate effect specifically while MK-801 counteracted the influence of NMDA. The effect produced by kainate was mimicked by domoic acid while the influence of homocysteic acid was identical with NMDA. Flupirtine alone did not influence the nature of the GABA immunoreactivity and so did not act as a kainate or NMDA agonist. However, flupirtine counteracted the influence produced by NMDA and homocysteic acid but had no effect on the kainate and domoic acid responses. Thus in this system flupirtine acts as an NMDA antagonist.

Flupirtine reduces functional deficits and neuronal damage after global ischemia in rats

Global cerebral ischemia leads to selective neuronal damage in the CA1 sector of the hippocampus and in the dorsolateral striatum. In addition, it results in deficits in spatial learning and memory as shown by an increase in escape latency and swim distance during the escape trials and a reduction of time spent in the quadrant of the former platform position during the probe trial of the water maze. Flupirtine is a non-opioid, centrally acting analgesic which has been shown to be neuroprotective against N-methyl-D-aspartate (NMDA)-mediated toxicity in vitro. The purpose of the present study was to investigate the potential protective effect of flupirtine in vivo with both behavioural and histological measures of global cerebral ischemia. Global ischemia was induced by four-vessel-occlusion (4VO) for 20 min in rats. Flupirtine was administered at a dose of 5 mg/kg i.p. either 20 min before and 50 min after occlusion (pre-treatment) or directly and 70 min after occlusion (post-treatment). 1 week after surgery, spatial learning and memory was tested in the Morris water maze. Pre-treatment with flupirtine reduced the increase in escape latency and in swim distance induced by 4VO. It also diminished the deficit in spatial memory as revealed by an increase in time spent in the quadrant of the former platform position during the probe trial which was reduced by 4VO. Post-treatment with flupirtine had no effect on the deficits in spatial learning and memory induced by 4VO. Neuronal damage in the CA1 sector of the hippocampus and in the striatum produced by 4VO was significantly attenuated with pre-treatment of flupirtine whereas post-treatment did not affect this neuronal damage. The present data demonstrate that pre-treatment with flupirtine exerts a protective effect on hippocampal and striatal neuronal damage and on deficits in spatial learning induced by 4VO.

N-Methyl-d-aspartate and α2-adrenergic mechanisms are involved in the depressent action of flupirtine on spinal reflexes in rats

In urethane-chloralose anesthetised rats the muscle relaxant activity of flupirtine was investigated on the monosynaptic Hoffmann reflex recorded from plantar foot muscles and on the polysynaptic flexor reflex recorded from tibialis muscle. Intraperitoneal (i.p.; 2.5-25 mumol/kg) and intrathecal (i.t.; 33-330 nmol) administration of flupirtine depressed the polysynaptic flexor reflex in anesthetised rats in a dose-dependent manner without affecting the monosynaptic Hoffmann reflex. Flupirtine produced a similar pattern on spinal reflexes as NMDA receptor antagonists, such as (-)-2-amino-7-phosphonoheptanoic acid (500 nmol i.t.) and memantine (125 mumol/kg i.p.), the benzodiazepines diazepam (18 mumol/kg i.p.) and midazolam (80 nmol i.t.), and the alpha 2-adrenoceptor agonist tizanidine (2 mumol/kg). In contrast, the GABAA receptor agonist muscimol (21 mumol/kg i.p.; 20 nmol i.t.) and the GABAB receptor agonist baclofen (47 mumol/kg i.p.; 2 nmol i.t.) reduced the magnitude of both the flexor and the Hoffmann reflex, whereas the non-NMDA receptor antagonist 6,7-dinitroquinoxaline-2,3-dione (DNQX; 10 nmol i.t.) depressed the Hoffmann reflex without affecting the flexor reflex. The effect of i.t. injection of flupirtine was prevented by coadministration of the mixed alpha 1/alpha 2-adrenoceptor antagonist yohimbine (10 nmol) and the excitatory amino acid N-methyl-D-aspartate (NMDA; 0.1 nmol), but neither by coadministration of the alpha 1-adrenoceptor antagonist prazosine (10 nmol), the GABAA receptor antagonist bicuculline (1 nmol), the GABAB receptor antagonist phaclofen (100 nmol), the non-NMDA receptor agonist alpha-amino-3-hydroxy-5-tertbutyl-4-isoxazolepropionic acid (ATPA; 0.1 pmol) nor by pre-treatment with the benzodiazepine receptor antagonist flumazenil (16 mumol/kg).(ABSTRACT TRUNCATED AT 250 WORDS)

Pretreatment but not posttreatment with GYKI 52466 reduces functional deficits and neuronal damage after global ischemia in rats

Glutamate antagonists have been shown to be neuroprotective in animal models of cerebral ischemia. Global cerebral ischemia in rats leads to selective neuronal damage in the hippocampus and striatum. Following ischemia a deficit in spatial learning and memory occurs. The aim of the present study was to investigate the potential neuroprotective effect of GYKI 52466, an antagonist at the non-N-methyl-D-aspartate receptor, with behavioural and histological measures of global ischemia in rats. Global ischemia was induced by four-vessel-occlusion (4VO) for 20 min in rats. GYKI 52466 (30 mg/kg i.p.) was administered either 20 min before induction of ischemia or immediately after onset of reperfusion. One week after surgery spatial learning was tested in the Morris water maze. After behavioural testing the animals were sacrificed and the neuronal damaged was assessed. GYKI 52466 reduced the increase in escape latency and in swim distance induced by 4VO when given before ischemia but not when applied after ischemia. Neuronal damage in the CA1 sector of the hippocampus produced by 4VO was significantly attenuated by pretreatment but not by posttreatment with GYKI 52466. Striatal neuronal damage was not affected by either treatment with GYKI 52466. GYKI 52466 had neuroprotective effects in a rat model of global cerebral ischemia. Pretreatment with GYKI 52466 protected rats against behavioural deficits and hippocampal neuronal damage induced by 4VO.

The antioxidant LY 231617 ameliorates functional and morphological sequelate induced by global ischemia in rats

In this study the effect of LY 231617, an antioxidant, on spatial learning deficit and on neuronal damage following transient cerebral ischemia was evaluated. Global ischemia was induced by four-vessel-occlusion (4VO) for 20 min in rats. LY 231617 (20 mg/kg i.p.) was administered after onset of reperfusion. One week after surgery spatial learning was tested in the Morris water maze. LY 231617 reduced the increase in escape latency and in swim distance induced by 4VO. Neuronal damage in the CAI sector of the hippocampus produced by 4VO was significantly attenuated by LY 231617. The present data demonstrate that posttreatment with LY 231617 exerts a protective effect on hippocampal neuronal damage and deficits in spatial learning induced by 4VO.

Unilateral Thalamic Edema in Internal Cerebral Venous Thrombosis: Is it Mostly Left?

Thrombotic occlusion of the internal cerebral veins is a particularly dangerous form of cerebral venous thrombosis (CVT) as it causes venous infarction of the thalami. Because both thalami drain into the vein of Galen and straight sinus, bilateral thalamic involvement is frequently encountered in internal CVT. However, unilateral thalamic edema may also occur, even if all internal cerebral veins are occluded. This suggests collateral venous drainage of the thalami, which is commonly insufficient in internal CVT. Patients with unilateral congestion of the thalamus, including 3 patients reported here, had mostly left-sided involvement, indicating that right-sided unilateral thalamic involvement in CVT may be clinically silent.

Zerebrale Vaskulitis als neurologische Begleiterkrankung bei Morbus Crohn

ZusammenfassungFür den Morbus Crohn sind einige neurologische Begleiterkrankungen wie die funikuläre Spinalerkrankung, Sehnervenschädigungen, Polyneuropathien, zerebrale Blutungen und Ischämien beschrieben worden. Eine zerebrale Vaskulitis als Ursache zerebraler Ischämien bei Patienten mit einem Morbus Crohn scheint eine Rarität zu sein. Wir berichten über 3 Fälle von zerebraler Vaskulitis bei Morbus Crohn bei Patientinnen im Alter von 26, 29 und 61 Jahren, die jeweils mit einer Hemiparese symptomatisch wurden; zudem beklagte eine Patientin Wortfindungsstörungen, Kopfschmerzen und Desorientierung. In der CCT- bzw. MRT-Untersuchung ließen sich multiple Hirnparenchymläsionen nachweisen. Die zerebrale Angiographie ergab in allen Fällen jeweils multiple Kalibersprünge der mittleren und großen intrakraniellen Gefäße im Sinne einer zerebralen Vaskulitis bei negativer Vaskulitisserologie. Die jeweilige Symptomatik bildete sich unter einer Immunsuppression und einer bei 2 Patientinnen zusätzlichen Antikoagulation zurück. Im Beobachtungszeitraum von 6–12 Monaten traten keine neuen neurologischen Symptome auf. Die dopplersonographischen Kontrolluntersuchungen zeigten in 2 Fällen unveränderte und in einem Fall progrediente Stenosen der basalen Hirnarterien.Da die autoimmunologisch bedingten entzündlichen Darmerkrankungen auch mit Vaskulitiden anderer Organe vergesellschaftet sein können, stellt das Auftreten einer zerebralen Vaskulitis im Zusammenhang mit dem Morbus Crohn wie bei unseren Patientinnen eine Möglichkeit einer vaskulitischen Organmanifestation dar.SummaryIn Crohns disease, some concomitant neurological illnesses such as cerebral ischemia following arterial or venous thrombosis, subacute combined degeneration of the spinal cord following malabsorption of vitamin B12 or folic acid, opticus neuropathy, and polyneuropathy have been described. Cerebral vasculitis secondary to Crohns disease seems to be a very rare phenomenon. We report on three such cases in three female patients (aged 26, 29, and 61 years). All patients became symptomatic with a hemiparesis; one complained additionally of a speech disorder, headache, and intermittent loss of orientation. In CT and MRI scans, multiple lesions were detected; cerebral angiography showed multiple stenoses of middle- and large-sized vessels that were compatible with cerebral vasculitis. Serologic tests concerning vasculitis were inconspicuous at that time. Under anticoagulation (in two cases) and immunosuppressive therapy, neurologic symptoms disappeared. In the following 6 to 12 months, no new neurological symptoms appeared. In two cases, Doppler sonographic controls showed stationary and, in one case, progressive intracranial stenoses.Since autoimmunologically caused inflammatory bowel diseases might be associated with vasculitis of other organs, the appearance of cerebral vasculitis secondary to Crohns disease is a possible organ manifestation by inflamed vessels.