minocycline and Ischemic-Attack--Transient

Memory deficit is the most visible symptom of cerebral ischemia that is associated with loss of pyramidal cells in CA1 region of the hippocampus. Oxidative stress and inflammation may be involved in the pathogenesis of ischemia/reperfusion (I/R) damage. Minocycline, a semi-synthetic tetracycline derived antibiotic, has anti-inflammatory and antioxidant properties. We evaluated the neuroprotective effect of minocycline on memory deficit induced by cerebral I/R in rat. I/R was induced by occlusion of common carotid arteries for 20min. Minocycline (40mg/kg, i.p.) was administered once daily for 7days after I/R. Learning and memory were assessed using the Morris water maze test. Nissl staining was used to evaluate the viability of CA1 pyramidal cells. The effects of minocycline on the microglial activation was also investigated by Iba1 (Ionized calcium binding adapter molecule 1) immunostaining. The content of malondialdehyde (MDA) and pro-inflammatory cytokines (IL-1β and TNF-α) in the hippocampus were measured by thiobarbituric acid reaction substances method and ELISA, respectively. Minocycline reduced the increase in escape latency time and in swimming path length induced by cerebral I/R. Furthermore, the ischemia-induced reduction in time spent in the target quadrant during the probe trial was increased by treatment with minocycline. Histopathological results indicated that minocycline prevented pyramidal cells death and microglial activation induced by I/R. Minocycline also reduced the levels of MDA and pro-inflammatory cytokines in the hippocampus in rats subjected to I/R. Minocycline has neuroprotective effects on memory deficit induced by cerebral I/R in rat, probably via its anti-inflammatory and antioxidant properties.

Topics: Animals; Anti-Inflammatory Agents; Antioxidants; Brain Ischemia; Cognitive Dysfunction; Disease Models, Animal; Hippocampus; Inflammation; Ischemic Attack, Transient; Learning; Male; Memory; Memory Disorders; Minocycline; Neuroprotective Agents; Oxidants; Oxidative Stress; Pyramidal Cells; Rats; Rats, Wistar; Reperfusion; Reperfusion Injury; Tumor Necrosis Factor-alpha

We investigated the neuroprotective properties of single doses of minocycline and ozagrel when administered prior to stroke. Male Sprague-Dawley rats were assigned randomly to one of the following groups: (1) control (Con) group (n=10), (2) minocycline (Mino) group (n=10), (3) sodium ozagrel (SO) group (n=10). Rats were treated with a single dose of minocycline or ozagrel at 30min before stroke. A middle cerebral artery occlusion (MCAO) was made at 30min after drug administration and reperfusion was done. The rats were subjected to a neurobehavioral test at days 1, 3 and 7 after MCAO. The cerebral ischemic volume was quantified by MetaMorph imaging software after TTC staining. The neuronal cell survival and astrocytes expansion were assessed by the NeuN and GFAP immunohistofluorescence staining. Apoptosis was detected by the TUNEL assay. We statistically analyzed and compared the results with each other. Mino and SO groups had neuroprotective effect and showed a better behavioral performance of adhesive removal and treadmill test at 7 days after stroke. Mino and SO groups also showed a smaller infarct volume than control group at 7 days after stroke. Immunohistofluorescence staining showed a higher number of NeuN positive cells, lower activated astrocytes in GFAP and a lower apoptosis in TUNEL staining. This study showed that single doses of minocycline and ozagrel prior to stroke had neuroprotective effects. These agents will be useful not only in post-stroke therapy but also in stroke prevention in several cerebrovascular procedures like carotid endarterectomy, bypass procedure, endovascular angioplasty, thromboembolectomy or thrombolysis.

Topics: Animals; Apoptosis; Astrocytes; Brain; Cell Survival; Disease Models, Animal; Fibrinolytic Agents; Infarction, Middle Cerebral Artery; Ischemic Attack, Transient; Male; Methacrylates; Minocycline; Neurons; Neuroprotective Agents; Neuropsychological Tests; Random Allocation; Rats, Sprague-Dawley; Stroke; Treatment Outcome

Cerebrovascular disease is a major cause of mortality and disability in adults. Diabetes mellitus increases the risk of cerebral ischaemia and is associated with worse clinical outcome following an event. Upregulation of matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9) in diabetes appears to play a role in vascular complications of diabetes. We hypothesised that inhibition of MMP-2 and MMP-9 by minocycline can be potentiated by aspirin through inhibition of cyclooxygenase-2 and tissue plasminogen activator, resulting in amelioration of clinical cerebral ischaemia in diabetes. In the present study, cerebral ischaemia/reperfusion injury was induced in streptozotocin diabetic rats by 1 h middle cerebral artery occlusion and 24 h reperfusion. Infarct volume, cerebral oedema, neurological severity score and blood-brain barrier disruption were significantly increased in diabetic animals compared with the normoglycemic control group. The combination of aspirin and minocycline treatment significantly improved these parameters in diabetic animals. Moreover, this therapy was associated with significantly lower mortality and reduction in MMP-2 and MMP-9 levels. Our data indicate that combination of aspirin and minocycline therapy protects from the consequences of cerebral ischaemia in animal models of diabetes and is associated with inhibition of MMP-2 and MMP-9. Therefore, this combination therapy may represent a novel strategy to reduce the neurological complications of cerebral ischaemia in diabetes.

Topics: Animals; Aspirin; Blood Glucose; Blood-Brain Barrier; Body Weight; Brain; Brain Edema; Capillary Permeability; Cerebral Infarction; Cyclooxygenase 2 Inhibitors; Cytoprotection; Diabetes Complications; Diabetes Mellitus, Experimental; Drug Synergism; Drug Therapy, Combination; Ischemic Attack, Transient; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Matrix Metalloproteinase Inhibitors; Minocycline; Protease Inhibitors; Rats; Rats, Wistar; Severity of Illness Index; Time Factors; Tissue Plasminogen Activator

Although it has been well established that ischemic insults promote cell proliferation in the subgranular zone (SGZ) of the hippocampal dentate gyrus (DG), the mechanisms by which this occurs remain unclear. The present study demonstrates that early-activated microglia in the hilus of the DG play an important role in ischemia-induced cell proliferation. Transient forebrain ischemia induced by 20min of bilateral common carotid artery occlusion (BCCAO) significantly increased cell proliferation in the SGZ of the DG beginning 4 days post-reperfusion. Moreover, BCCAO increased microglial activation in the hilus of the DG from 1 day post-reperfusion and in the CA1 layer from 4 days post-reperfusion. An injection of minocycline (10 or 100nmol in 0.5microl) into the DG immediately after reperfusion decreased microglial activation in the hilus of the DG 1 day post-reperfusion, but only a high dose of minocycline (100nmol) significantly decreased microglial activation in the CA1 layer. Both high and low doses of minocycline significantly decreased the number of BrdU-positive cells at 7 days post-reperfusion. These results suggest that early-activated microglia in the hilus of the DG take part in the cell proliferation induced by transient forebrain ischemia.

Topics: Animals; Carotid Artery, Common; Carotid Stenosis; Cell Proliferation; Ischemic Attack, Transient; Male; Mice; Mice, Inbred C57BL; Microglia; Minocycline; Neurons; Prosencephalon; Stem Cells

Recently, the beneficial role of minocycline on endogenous neurogenesis after cerebral ischemia has been contradicted by many reports. We examined whether minocycline influences post-ischemic neurogenesis in the subventricular zone. Adult male Sprague-Dawley rats were subjected to focal cerebral ischemia for 2 h, and divided into a minocycline-treated (90 mg/Kg on reperfusion and 45 mg/Kg daily for maintenance) and a saline-treated group. Bromodeoxyuridine was injected to determine levels of cell proliferation. Inflammation was assessed by counting polymorphonuclear cell and activated microglia and by measuring myeloperoxidase activity. Endogenous neurogenesis was quantified by immunohistochemical staining and functional outcome was measured by infarct size and behavioral tests. Minocycline treatment decreased inflammation on 1st and 4th days after ischemia. BrdU-positive cells on 7th day (saline vs. minocycline: 602.80+/-146.96 vs. 399.40+/-109.69) and the number of double labeling cells of BrdU/NeuN on 7th day (13.00+/-4.36 vs. 6.40+/-2.07) and BrdU/DCx on 4th day (17.00+/-5.00 vs. 7.50+/-1.91) were significantly decreased in minocycline-treated rats. Infarct size and behavioral tests were not different. Our results indicate that minocycline may reduce immediate post-ischemic neurogenesis despite adequately suppressed inflammation.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Brain; Brain Ischemia; Bromodeoxyuridine; Cell Count; Doublecortin Domain Proteins; Doublecortin Protein; Enzyme Activation; Ischemic Attack, Transient; Male; Microglia; Microtubule-Associated Proteins; Minocycline; Motor Activity; Neurogenesis; Neuroimmunomodulation; Neurons; Neuropeptides; Neutrophils; Peroxidase; Rats; Rats, Sprague-Dawley; Treatment Outcome

Considering that several pathways leading to cell death are activated in cerebral ischemia, we tested in mouse models of transient and permanent ischemia a drug cocktail aiming at distinct pharmacological targets during the evolution of ischemic injury. It consists of minocycline--an antibiotic with anti-inflammatory properties, riluzole--a glutamate antagonist, and nimodipine--a blocker of voltage-gated calcium channels. Administered 2 h after transient or permanent MCAO, it significantly decreased the size of infarction, by approximately 65% after transient and approximately 35% after permanent ischemia and markedly improve clinical recovery of mice. In both experimental models a three-drug cocktail achieved significantly more efficient neuroprotection than any of the components tested alone. However, some interesting observation emerged from the single-drug studies. Treatment with minocycline alone was efficient in both experimental models while treatment with glutamate antagonist riluzole conferred neuroprotection only after transient MCAO. Immunohistochemical analysis following three-drug treatment revealed reduced microglia/macrophages and caspase-3 activation as well as preserved GFAP immunoreactivity following transient ischemia. No detectable differences in the levels of Mac-2, GFAP and caspase-3 immunoreactivities were observed 72 h after permanent MCAO. These marked differences in the brain tissue responses to ischemic injury and to treatments suggest that different pathological mechanisms may be operating in transient and permanent ischemia. However, the three-drug cocktail exerted significant neuroprotection in both experimental models thus demonstrating that simultaneous targeting of several pathophysiological pathways involved in the evolution of ischemic injury may represent a rational therapeutic strategy for stroke.

Topics: Animals; Brain; Brain Ischemia; Caspase 3; Drug Combinations; Enzyme Activation; Glial Fibrillary Acidic Protein; Immunohistochemistry; Ischemic Attack, Transient; Macrophages; Male; Mice; Mice, Inbred C57BL; Microglia; Minocycline; Myocardial Infarction; Neuroprotective Agents; Nimodipine; Riluzole; Time Factors

We investigated the effect of minocycline on neuronal damage in the hippocampus and striatum in a mouse model of transient global forebrain ischemia. Male C57BL/6 mice were anesthetized with halothane and subjected to bilateral occlusion of the common carotid artery (BCCAO) for 30 min. Minocycline (90 mg/kg, i.p., qd) or saline was injected immediately after BCCAO and daily for the next two days (45 mg/kg, i.p., bid). In order to reduce the variability in ischemic neuronal damage, we applied selection criteria based on regional cerebral blood flow (rCBF), evaluated using laser Doppler flowmetry, and the plasticity of the posterior communicating artery (PcomA), evaluated using India ink solution. In animals with rCBF that was less than 15% of the baseline value and with a smaller PcomA, of diameter less than one-third that of the basilar artery, we consistently observed neuronal damage in the striatum and hippocampal subfields, including medial CA1, CA2, and CA4. When the effect of minocycline was assessed with cresyl violet staining, neuronal damage in the medial part of the CA1 subfield and the striatum was found to be significantly attenuated, although minocycline did not protect against neuronal damage in the remaining hippocampal subfields. Immunohistochemistry for NeuN, adenosine A1 receptor, and SCIP/Oct-6 confirmed the region-specific effect of minocycline in the hippocampus. In summary, our results suggest that minocycline protects neurons against global forebrain ischemia in a subregion-specific manner.

Topics: Animals; Cerebrovascular Circulation; Corpus Striatum; Disease Models, Animal; Hippocampus; Immunohistochemistry; Ischemic Attack, Transient; Laser-Doppler Flowmetry; Male; Mice; Mice, Inbred C57BL; Minocycline; Nerve Degeneration; Neurons; Neuroprotective Agents; Prosencephalon

Minocycline is an antibiotic now recognized to have antiapoptotic and antiinflammatory properties. Because of these properties, minocycline may be of benefit in reducing neuronal apoptosis from ischemia and subsequent postischemic inflammation if administered soon after a stroke. We now explore the feasibility of using (99m)Tc-annexin V, an in vivo marker of apoptosis, with SPECT to monitor the antiapoptotic effects of minocycline therapy.. CB6/F1 adult male mice underwent unilateral distal middle cerebral artery occlusion (dMCA) occlusion and were imaged and sacrificed at 1, 3, 7, or 30 d after injury. Animals were given minocycline (or vehicle) 30 min and 12 h after dMCA occlusion and then given 22.5 mg/kg twice daily for up to 7 d. Before imaging, behavioral tests were performed to evaluate the neurologic function. After imaging, brains were collected for histology and assessed for the degree of apoptosis and microglial activation.. (99m)Tc-Annexin V uptake in injured hemispheres was significantly decreased 2- to 3-fold by minocycline at all time points. Minocyline reduced infarct size as seen histologically and improved behavioral indices as late as 30 d. Infarct volume as seen histologically correlated with radiolabeled annexin V uptake seen by SPECT. In situ fluorescent microscopy demonstrated that annexin V bound primarily to neurons at 1 and 3 d, with a shift toward microglia by 7 and 30 d.. We found that minocycline significantly reduces neuronal apoptosis and infarct size and improves neurologic outcome in mice after acute focal cortical ischemia.

Topics: Animals; Annexin A5; Apoptosis; Hydrazines; Infarction, Middle Cerebral Artery; Ischemic Attack, Transient; Male; Mice; Microglia; Minocycline; Neuroprotective Agents; Nicotinic Acids; Organotechnetium Compounds; Radiopharmaceuticals; Tomography, Emission-Computed, Single-Photon

Minocycline, a semi-synthetic tetracycline antibiotic, is an effective neuroprotective agent in animal models of cerebral ischemia when given in high doses intraperitoneally. The aim of this study was to determine if minocycline was effective at reducing infarct size in a Temporary Middle Cerebral Artery Occlusion model (TMCAO) when given at lower intravenous (IV) doses that correspond to human clinical exposure regimens.. Rats underwent 90 minutes of TMCAO. Minocycline or saline placebo was administered IV starting at 4, 5, or 6 hours post TMCAO. Infarct volume and neurofunctional tests were carried out at 24 hr after TMCAO using 2,3,5-triphenyltetrazolium chloride (TTC) brain staining and Neurological Score evaluation. Pharmacokinetic studies and hemodynamic monitoring were performed on minocycline-treated rats.. Minocycline at doses of 3 mg/kg and 10 mg/kg IV was effective at reducing infarct size when administered at 4 hours post TMCAO. At doses of 3 mg/kg, minocycline reduced infarct size by 42% while 10 mg/kg reduced infarct size by 56%. Minocycline at a dose of 10 mg/kg significantly reduced infarct size at 5 hours by 40% and the 3 mg/kg dose significantly reduced infarct size by 34%. With a 6 hour time window there was a non-significant trend in infarct reduction. There was a significant difference in neurological scores favoring minocycline in both the 3 mg/kg and 10 mg/kg doses at 4 hours and at the 10 mg/kg dose at 5 hours. Minocycline did not significantly affect hemodynamic and physiological variables. A 3 mg/kg IV dose of minocycline resulted in serum levels similar to that achieved in humans after a standard 200 mg dose.. The neuroprotective action of minocycline at clinically suitable dosing regimens and at a therapeutic time window of at least 4-5 hours merits consideration of phase I trials in humans in view of developing this drug for treatment of stroke.

Topics: Animals; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Evaluation, Preclinical; Infarction, Middle Cerebral Artery; Injections, Intravenous; Ischemic Attack, Transient; Male; Minocycline; Neurologic Examination; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Time Factors; Treatment Outcome

Ischemic stroke is the most common life-threatening neurological disease and has limited therapeutic options. One component of ischemic neuronal death is inflammation. Here we show that doxycycline and minocycline, which are broad-spectrum antibiotics and have antiinflammatory effects independent of their antimicrobial activity, protect hippocampal neurons against global ischemia in gerbils. Minocycline increased the survival of CA1 pyramidal neurons from 10.5% to 77% when the treatment was started 12 h before ischemia and to 71% when the treatment was started 30 min after ischemia. The survival with corresponding pre- and posttreatment with doxycycline was 57% and 47%, respectively. Minocycline prevented completely the ischemia-induced activation of microglia and the appearance of NADPH-diaphorase reactive cells, but did not affect induction of glial acidic fibrillary protein, a marker of astrogliosis. Minocycline treatment for 4 days resulted in a 70% reduction in mRNA induction of interleukin-1beta-converting enzyme, a caspase that is induced in microglia after ischemia. Likewise, expression of inducible nitric oxide synthase mRNA was attenuated by 30% in minocycline-treated animals. Our results suggest that lipid-soluble tetracyclines, doxycycline and minocycline, inhibit inflammation and are neuroprotective against ischemic stroke, even when administered after the insult. Tetracycline derivatives may have a potential use also as antiischemic compounds in humans.

Topics: Animals; Doxycycline; Gerbillinae; Glial Fibrillary Acidic Protein; Hippocampus; Humans; Ischemic Attack, Transient; Male; Microglia; Minocycline; Neuroprotective Agents; Pyramidal Cells; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tetracyclines; Transcription, Genetic