Search

We aim to provide a comprehensive review of the current scientific evidence supporting the use of invasive neurostimulation in the treatment of deficits associated with traumatic brain injury (TBI), as well as to identify future directions for research and highlight important questions that remain unaddressed. Neurostimulation is a treatment modality with expanding applications in modern medical practice. Targeted electrical stimulation of specific brain regions has been shown to increase synaptogenesis and enhance structural reorganization of neuronal networks. This underlying therapeutic effect might be of high value for patients suffering from TBI because it could modulate neuronal connectivity and function of areas that are partially or completely spared after injury. The current published literature exploring the application of invasive neurostimulation for the treatment of functional deficits associated with TBI is scarce but promising. Rodent models have shown that targeted stimulation of the hippocampus or connecting structures can result in significant cognitive recovery, while stimulation of the motor cortex and deep cerebellar nuclei is associated with motor improvements. Data from clinical studies are extremely limited; single-patient reports and case series found neurostimulation to be effective in relieving motor symptoms, improving visuospatial memory, and supporting emotional adjustment. Looking forward, it will be important to identify stimulation targets and paradigms that can maximize improvement over multiple functional domains. It will also be important to corroborate the observed behavioral improvements with histological, electrophysiological, and radiological evidence. Finally, the impact of biological variables such as sex and age on the treatment outcomes needs to be explored.

Functional outcome following traumatic brain injury (TBI) varies greatly, with approximately half of those who survive suffering long-term motor and cognitive deficits despite contemporary rehabilitation efforts. We have previously shown that deep brain stimulation (DBS) of the lateral cerebellar nucleus (LCN) enhances rehabilitation of motor deficits that result from brain injury. The objective of the present study was to evaluate the efficacy of LCN DBS on recovery from rodent TBI that uniquely models the injury location, chronicity and resultant cognitive symptoms observed in most human TBI patients. We used controlled cortical impact (CCI) to produce an injury that targeted the medial prefrontal cortex (mPFC-CCI) bilaterally, resulting in cognitive deficits. Unilateral LCN DBS electrode implantation was performed 6 weeks post-injury. Electrical stimulation started at week eight post-injury and continued for an additional 4 weeks. Cognition was evaluated using baited Y-maze, novel object recognition task and Barnes maze. Post-mortem analyses, including Western Blot and immunohistochemistry, were conducted to elucidate the cellular and molecular mechanisms of recovery. We found that mPFC-CCI produced significant cognitive deficits compared to pre-injury and naïve animals. Moreover, LCN DBS treatment significantly enhanced the long-term memory process and executive functions of applying strategy. Analyses of post-mortem tissues showed significantly greater expression of CaMKIIα, BDNF and p75

Stroke is a leading cause of disability in the Western world. Current post-stroke rehabilitation treatments are only effective in approximately half of the patients. Therefore, there is a pressing clinical need for developing new rehabilitation approaches for enhancing the recovery process, which requires the use of appropriate animal models. Here, we demonstrate the use of nonlinear microscopy of calcium sensors in the rat brain to study the effects of ischemic stroke injury on cortical activity patterns. We longitudinally recorded from thousands of neurons labeled with a genetically-encoded calcium indicator before and after an ischemic stroke injury in the primary motor cortex. We show that this injury has an effect on the activity patterns of neurons not only in the motor and somatosensory cortices, but also in the more distant visual cortex, and that these changes include modified firing rates and kinetics of neuronal activity patterns in response to a sensory stimulus. Changes in neuronal population activity provided animal-specific, circuit-level information on the post-stroke cortical reorganization process, which may be essential for evaluating the efficacy of new approaches for enhancing the recovery process.

Stroke is a leading cause of disability in the Western world. Current post-stroke rehabilitation treatments are only effective in approximately half of the patients. Therefore, there is a pressing clinical need for developing new rehabilitation approaches for enhancing the recovery process, which requires the use of appropriate animal models. Here we study the activity patterns of multiple cortical regions in the rat brain using two-photon microscopy. We longitudinally recorded the fluorescence signal from thousands of neurons labeled with a genetically-encoded calcium indicator before and after an ischemic stroke injury, and found substantial functional changes across motor, somatosensory, and visual cortical regions during the post-stroke cortical reorganization period. We show that a stroke injury in the primary motor cortex has an effect on the activity patterns of neurons not only in the motor and somatosensory cortices, but also in the more distant visual cortex, and that these changes include modified firing rates and kinetics of neuronal activity patterns in response to a sensory stimulus. Changes in neuronal population activity provided animal-specific, circuit-level information on the poststroke cortical reorganization process, which may be essential for evaluating the efficacy of new approaches for enhancing the recovery process.

5-bromo-2′-dexoyuridine (BrdU) is often used in neuroscience research as a marker of newly-divided cells. However, several studies suggest that BrdU can produce unwanted side effects, including changes in animal behavior and cellular function. In this study, we investigated the effect of BrdU injections on locomotor behavior in a rodent model of ischemic stroke. Ischemic strokes were induced in adult rats, and 50 mg/kg BrdU was intraperitoneally injected over 5 days beginning 2 weeks post-stroke, while control animals received vehicle. Locomotor activity was evaluated by videotaping the rats in their home cages for 30 min, beginning one hour after BrdU injection. BrdU-injected rats showed a nearly three-fold increase in locomotor activity compared to control animals. These findings suggest that BrdU induces a hyperlocomotor effect in rats following brain injury, pointing to the need for caution when interpreting behavioral results in such studies.

We previously reported that 1,3-bisbenzylimidazolium (DBZIM) bromide was neuroprotective for the dopaminergic system in Parkinson's disease (PD) models of rodent, however the underlying mechanism was unclear. We currently further confirmed that DBZIM ameliorated the Parkinsonian motor deficit and protected the nigrostriatal tract from the neurotoxicity of 1-methyl-4-(2'-methylphenyl)-1,2,3,6-tetrahydropyridine (2'-CH3-MPTP) in C57Bl/6 mice. The dopaminergic degeneration in the substantia nigra par compacta (SNc) and striatum was analyzed by immunohistochemistry while the monoamine oxidase B (MAO-B) inhibition effect of DBZIM was determined by enzyme kinetics. DBZIM was at least as effective as the clinically approved anti-PD drug, l-deprenyl (Selegiline), for both neuroprotection and correction of motor deficits. Mechanistically, DBZIM inhibited the specific activity of MAO-B in the striatum and C6 cells without affecting the protein expression. DBZIM directly inhibited the enzymatic activity of a purified MAO-B protein with an estimated Ki value from 780 to 940nM, in par with that of l-deprenyl (970nM). The physical interaction between DBZIM and MAO-B was proven by NMR analysis, with Kd around 21.5-46.8μM. Our binding and modelling data further illustrated that DBZIM is a mixed inhibitor with its binding to active site partially hindering the substrate binding. Therefore, inhibiting MAO-B is a major mechanism through which DBZIM confers neuroprotection for the dopaminergic neurons against 2'-CH3-MPTP toxicity. Remarkably, the post-lesion treatment with DBZIM provided greater anti-parkinsonian and neuroprotective effects than the l-deprenyl. The current study, together with our previous findings in a 6-OHDA PD model, demonstrated that DBZIM is a promising neuroprotectant for PD with anti-MAO-B property.

Background Chronic deep brain stimulation of the rodent lateral cerebellar nucleus (LCN) has been demonstrated to enhance motor recovery following cortical ischemia. This effect is concurrent with synaptogenesis and expression of long-term potentiation markers in the perilesional cerebral cortex. Objective To further investigate the cellular changes associated with chronic LCN stimulation in the ischemic rodent by examining neurogenesis along the cerebellothalamocortical pathway. Methods Rats were trained on the pasta matrix task, followed by induction of cortical ischemia and electrode implantation in the contralesional LCN. Electrical stimulation was initiated 6 wk after stroke induction and continued for 4 wk prior to sacrifice. Neurogenesis was examined using immunohistochemistry. Results Treated animals showed enhanced performance on the pasta matrix task relative to sham controls. Increased cell proliferation colabeled with 5'-Bromo-2'-deoxyuridine and neurogenic markers (doublecortin) was observed in the perilesional cortex as well as bilateral mediodorsal and ventrolateral thalamic subnuclei in treated vs untreated animals. The neurogenic effect at the level of motor cortex was selective, with stimulation-treated animals showing greater glutamatergic neurogenesis but significantly less GABAergic neurogenesis. Conclusion These findings suggest that LCN deep brain stimulation modulates postinjury neurogenesis, providing a possible mechanistic foundation for the associated enhancement in poststroke motor recovery.

Ischemic stroke is one of the most common causes of death. A brain that has suffered a stroke is significantly damaged due to the transient interruption of blood supply. Stroke survivors suffer from stroke-induced cognitive, motor and psychiatric problems that require increased use of the health care system, which results in a very poor quality of life and a heavy emotional and financial burden for patients and their families. Although rehabilitation can ameliorate some behavioral abnormalities, it does not significantly improve stroke symptoms. Neuroregeneration may be the principle process for replenishing of lost brain functions. Stem cell therapy is a novel approach to restore neurological function following ischemic stroke. Our goal in this review is to first briefly introduce the pathophysiology of stroke then document the progress of stem cell research, including clinical trials, towards the treatment of ischemic stroke. We highlight the promising applications of biomaterials to enhance the therapeutic effect of stem cell therapy in stroke treatment. As biomaterials provide efficient scaffolds for the growth and differentiation of stem cells, they may guarantee the quality and yield of stem cells for grafting. Furthermore, biomaterials also provide a convenient method for delivering the stem cells to be transplanted. This review provides direction for future research to combine tissue engineering approaches with stem cell therapy for the treatment of stroke.

Background Many traumatic brain injury (TBI) survivors live with persistent disability from chronic motor deficits despite contemporary rehabilitation services, underscoring the need for novel treatment. Objective/Hypothesis We have previously shown that deep brain stimulation (DBS) of the lateral cerebellar nucleus (LCN) can enhance post-stroke motor recovery and increase the expression of markers of long-term potentiation in perilesional cerebral cortex. We hypothesize that a similar beneficial effect will be for motor deficits induced by unilateral fluid percussion injury (FPI) in rodents through long-term potentiation- and anti-inflammatory based mechanisms. Methods Male Long Evans rats with a DBS macroelectrode in the LCN underwent FPI over contralateral primary motor cortex. After 4 weeks of spontaneous recovery, DBS treatment was applied for 4 weeks, with the pasta matrix, cylinder, and horizontal ladder tests used to evaluate motor performance. All animals were euthanized and tissue harvested for further analysis by histology, immunohistochemistry, RNA microarray assay and Western Blot. Results LCN DBS-treated animals experienced a significantly greater rate of motor recovery than untreated surgical controls, with treated animals showing enhanced expression of RNA and protein for excitability related genes, suppressed expression of pro-inflammatory genes, suppressed microglial and astrocytic activation, but proliferation of c-fos positive cells. Finally, our data suggest a possible role for anti-apoptotic effects with LCN DBS. Conclusion LCN DBS enhanced the motor recovery following TBI, possibly by elevating the neuronal excitability at the perilesional area and mediating anti-apoptotic and anti-inflammatory effects.

Bis(12)-hupyridone (B12H), derived from the Chinese medicinal component huperzine A, was originally designed as a novel acetylcholinesterase (AChE) inhibitor. In this paper, we report that B12H (24-h pretreatment) effectively blocked glutamate-induced neuronal excitotoxicity in cerebellar granule neurons (CGNs). However, the huge discrepancy between the EC50 value and IC50 value of B12H, to protect against neuronal toxicity (0.09 μM) and to block the NMDA receptor (21.8 μM) respectively, suggests that the neuroprotection of B12H might be not primarily due to the blockade of the NMDA receptor. Pretreatment by specific antagonists of alpha7-nicotinic acetylcholine receptor (α7nAChR), but not muscarinic acetylcholine receptor (mAChR) or α4β2nAChR, decreased the neuroprotection of B12H. The neuroprotection of B12H could also be abolished by the pretreatment of specific PI3-K inhibitors. Furthermore, B12H restored the suppressed activation of the Akt pathway caused by glutamate as evidenced by the decreased expressions of pSer473-Akt and pSer9-GSK3β. All these results suggest that B12H substantially protected CGNs against glutamate-induced neuronal excitotoxicity via activating α7nAChR/PI3-K/Akt cascade.

Crossed cerebellar diaschisis (CCD) is a functional deficit of the cerebellar hemisphere resulting from loss of afferent input consequent to a lesion of the contralateral cerebral hemisphere. It is manifested as a reduction of metabolism and blood flow and, depending on severity and duration, it can result in atrophy, a phenomenon known as crossed cerebellar atrophy (CCA). While CCA has been well-demonstrated in humans, it remains poorly characterized in animal models of stroke. In this study we evaluated the effects of cerebral cortical ischemia on contralateral cerebellar anatomy using an established rodent model of chronic stroke. The effects of cortical ischemia on the cerebellar hemispheres, vermis and deep nuclei were characterized. Intracortical microinjections of endothelin-1 (ET-1) were delivered to the motor cortex of Long Evans rats to induce ischemic stroke, with animals sacrificed 6 weeks later. Naive animals served as controls. Cerebral sections and cerebellar sections including the deep nuclei were prepared for analysis with Nissl staining. Cortical ischemia was associated with significant thickness reduction of the molecular layer at the Crus 1 and parafloccular lobule (PFL), but not in fourth cerebellar lobule (4Cb), as compared to the ipsilesional cerebellar hemisphere. A significant reduction in volume and cell density of the lateral cerebellar nucleus (LCN), the rodent correlate of the dentate nucleus, was also noted. The results highlight the relevance of corticopontocerebellar (CPC) projections for cerebellar metabolism and function, including its direct projections to the LCN.

We have recently identified a class of imidazolium salts (IMSs) with antioxidative property and can function as scavengers for radical oxygen species (ROS) [18]. Here, we investigate one of the IMSs, 1,3-bisbenzylimidazolium bromide (DBZIM), for its possible role in attenuating neurotoxicity and gliosis in the retina and the brain induced by a Parkinsonian neurtoxicant, methyl-4(2 � methylphenyl)-1,2,3,6-tetrahydropyridine (2 � -CH3-MPTP), which is a free radical generating agent. In this study, we employ a molecular retinal imaging method, which we recently developed in a transgenic mouse model expressing green fluorescent protein (GFP) under the control of glial fibrillary acidic protein (GFAP) promoter [14], to assess the efficacy of DBZIM, since currently no in vitro system with a sufficient complexity is available for accurately assessing a compound’s efficacy. The longitudinal imaging results showed DBZIM can effectively suppress the neurotoxicant-induced retinal gliosis. Immunohistochemistry performed on the postmodern mouse brain confirmed that DBZIM also reduced striatal gliosis, and concomitantly attenuated the loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc). These findings suggest that DBZIM could be a useful small molecular compound for studying neurotoxicity and neuroprotection in the retina and the brain.

Clinical trials have demonstrated positive proof of efficacy of dual metabotropic glutamate receptor 2/3 (mGluR2/3) agonists in both anxiety and schizophrenia. Importantly, evidence suggests that these drugs may also be neuroprotective against glutamate excitotoxicity, implicated in the pathogenesis of Parkinson's disease (PD). However, whether this neuroprotection also translates into functional recovery is unclear. In the current study, we examined the neuroprotective efficacy of the dual mGluR2/3 agonist, 2R,4R-4-aminopyrrolidine-2,4-dicarboxylate (2R,4R-APDC), and whether this is accompanied by behavioral recovery in a rodent 6-hydroxydopamine (6-OHDA) model of PD. We now report that delayed post lesion treatment with 2R,4R-APDC (10 nmol), results in robust neuroprotection of the nigrostriatal system, which translated into functional recovery as measured by improved forelimb use asymmetry and reduced (+)-amphetamine-induced rotation compared to vehicle treated animals. Interestingly, these beneficial effects were associated with a decrease in microglial markers in the SNc, which may suggest an antiinflammatory action of this drug.

The excessive activation of the N-methyl-D-aspartate receptor (NMDAR)/nitric oxide (NO) pathway has been proposed to be involved in the neuropathology of various neurodegenerative disorders. In this study, NO was found to mediate glutamate-induced excitotoxicity in primary cultured neurons. Compared with the NO synthase (NOS) inhibitor, N(G)-monomethyl-L-arginine (L-NMMA), and the NMDAR antagonist memantine, bis(7)-tacrine was found to be more potent in reducing NO-mediated excitotoxicity and the release of NO caused by glutamate. Moreover, like L-NMMA but not like 5H-dibenzo[a,d]cyclohepten-5,10-imine (MK-801) and memantine, bis(7)-tacrine showed greater neuroprotection and inhibition on NO release when neurons were pretreated for a prolonged time between 0 and 24 h and remained quite potent even when neurons were post-treated 1 h after the glutamate challenge. Bis(7)-tacrine was additionally found to be as moderately potent as memantine in competing with [(3)H]MK-801, inhibiting NMDA-evoked currents and reducing glutamate-triggered calcium influx, which eventually reduced neuronal NOS activity. More importantly, at neuroprotective concentrations, bis(7)-tacrine substantially reversed the overactivation of neuronal NOS caused by glutamate without interfering with the basal activity of NOS. Furthermore, in vitro pattern analysis demonstrated that bis(7)-tacrine competitively inhibited both purified neuronal and inducible NOS with IC(50) values at 2.9 and 9.3 microM but not endothelial NOS. This result was further supported by molecular docking simulations that showed hydrophobic interactions between bis(7)-tacrine and three NOS isozymes. Taken together, these results strongly suggest that the substantial neuroprotection against glutamate by bis(7)-tacrine might be mediated synergistically through the moderate blockade of NMDAR and selective inhibition of neuronal NOS.

The olivo-cerebellar system has an essential role in the detection and adaptive correction of movement errors. While there is evidence of an error signal in the cerebellar cortex and inferior olivary nucleus, the deep cerebellar nuclei have been less thoroughly investigated. Here, we recorded local field potential activity in the rodent lateral cerebellar nucleus during a skilled reaching task and compared event-related changes in neural activity between unsuccessful and successful attempts. Increased low gamma (40-50 Hz) band power was present throughout the reach and grasp behavior, with no difference between successful and unsuccessful trials. Beta band (12-30 Hz) power, however, was significantly increased in unsuccessful reaches, compared to successful, throughout the trial, including during the epoch preceding knowledge of the trial's outcome. This beta band activity was greater in unsuccessful trials of high-performing days, compared to unsuccessful trials of low-performing days, indicating that this activity may reflect an error prediction signal, developed over the course of motor learning. These findings suggest an error-related discriminatory oscillatory hallmark of movement in the deep cerebellar nuclei.

The design of biomaterials and the sourcing for appropriate cells are two integrated aspects of tissue engineering to construct a tissue implant for clinical applications. During the past decades, many innovative biomaterials with desirable biological and mechanical properties have emerged, while stem cells have been shown to be a promising cell source to differentiate into many cell types. However, the testing of these bioartificial tissue constructs in the clinical trials is far from satisfactory. How microenvironments in the biomaterials regulate cellular signaling pathways and functions, how stem cell-derived target cells respond to extracellular cues presented by the biomaterials, and how implanted tissue constructs interact with host tissues remain to be investigated. The information of the interplay between cell and biomaterials would be helpful to guide us in improving our current strategy to refine the tissue constructs for effective tissue repair in regenerative medicine. In this special issue, we first present a thorough review by M. Ramamoorthi et al. on the efficacy of dental stem cell therapy in bone regeneration in preclinical in vivo and in vitro studies. They suggest that well-designed randomized animal trials are needed before moving into clinical trials. Then we introduce three types of innovative biomaterials: (1) bionanocomposites based on bacterial cellulose and magnetic nanoparticles (magnetite) for efficient chronic wounds healing reported by B. Galateanu et al.; (2) the elastomeric poly(e-caprolatone urethane) (PCLU) scaffolds using a high internal phase emulsion for bone tissue regeneration developed by S. Changotade et al.; and (3) a novel gelatin-alginate-polyacrylamide 3D interpenetrating network with superior performance in promoting chondrogenesis using human adipose-derived stem cells designed by S. Dinescu et al. Most importantly, our special focus will be given to the insight studies on the cell-biomaterial interactions. C.-H. Wang et al. demonstrate that the migration ability of bone marrow stem cells can be regulated by varying the porous structure of the artificial ligaments and this regulation is related to the RhoA/ROCK signaling pathway. M. Deng et al. demonstrate the endothelial differentiation of human adipose-derived stem cells on the polyglycolic acid/polylactic acid (PGA/PLA) mesh and propose the synergistic effect of 3D environments and biochemical signals such as growth factors on the acquisition of mature characteristic endothelial phenotype. S. Fu et al. have investigated the protective effect of neuropeptide Substance P on bone marrow stromal cells against apoptosis induced by serum deprivation through Wnt signaling. J. Michel et al. have reviewed a wide range of hydroxyapatite containing scaffolds and their interactions with mesenchymal stem cells in in vitro and in vivo contexts.

The neuroprotective properties of bis(7)-tacrine, a novel dimeric acetylcholinesterase (AChE) inhibitor, on glutamate-induced excitotoxicity were investigated in primary cultured cerebellar granule neurons (CGNs). Exposure of CGNs to 75 mum glutamate resulted in neuronal apoptosis as demonstrated by Hoechst staining, TUNEL, and DNA fragmentation assays. The bis(7)-tacrine treatment (0.01-1 mum) on CGNs markedly reduced glutamate-induced apoptosis in dose- and time-dependent manners. However, donepezil and other AChE inhibitors, even at concentrations of inhibiting AChE to the similar extents as 1 mum bis(7)-tacrine, failed to prevent glutamate-induced excitotoxicity in CGNs; moreover, both atropine and dihydro-beta-erythroidine, the cholinoreceptor antagonists, did not affect the anti-apoptotic properties of bis(7)-tacrine, suggesting that the neuroprotection of bis(7)-tacrine appears to be independent of inhibiting AChE and cholinergic transmission. In addition, ERK1/2 and p38 pathways, downstream signals of N-methyl-d-aspartate (NMDA) receptors, were rapidly activated after the exposure of glutamate to CGNs. Bis(7)-tacrine inhibited the apoptosis and the activation of these two signals with the same efficacy as the coapplication of PD98059 and SB203580. Furthermore, using fluorescence Ca(2+) imaging, patch clamp, and receptor-ligand binding techniques, bis(7)-tacrine was found effectively to buffer the intracellular Ca(2+) increase triggered by glutamate, to reduce NMDA-activated currents and to compete with [(3)H]MK-801 with an IC(50) value of 0.763 mum in rat cerebellar cortex membranes. These findings strongly suggest that bis(7)-tacrine prevents glutamate-induced neuronal apoptosis through directly blocking NMDA receptors at the MK-801-binding site, which offers a new and clinically significant modality as to how the agent exerts neuroprotective effects.

Minocycline has been shown to have remarkably neuroprotective qualities, but underlying mechanisms remain elusive. We reported here the robust neuroprotection by minocycline against glutamate-induced apoptosis through regulations of p38 and Akt pathways. Pre-treatment of cerebellar granule neurons (CGNs) with minocycline (10-100 microm) elicited a dose-dependent reduction of glutamate excitotoxicity and blocked glutamate-induced nuclear condensation and DNA fragmentations. Using patch-clamping and fluorescence Ca2+ imaging techniques, it was found that minocycline neither blocked NMDA receptors, nor reduced glutamate-caused rises in intracellular Ca2+. Instead, confirmed by immunoblots, minocycline in vivo and in vitro was shown to directly inhibit the activation of p38 caused by glutamate. A p38-specific inhibitor, SB203580, also attenuated glutamate excitotoxicity. Furthermore, the neuroprotective effects of minocycline were blocked by phosphatidylinositol 3-kinase (PI3-K) inhibitors LY294002 and wortmannin, while pharmacologic inhibition of glycogen synthase kinase 3beta (GSK3beta) attenuated glutamate-induced apoptosis. In addition, immunoblots revealed that minocycline reversed the suppression of phosphorylated Akt and GSK3beta caused by glutamate, as were abolished by PI3-K inhibitors. These results demonstrate that minocycline prevents glutamate-induced apoptosis in CGNs by directly inhibiting p38 activity and maintaining the activation of PI3-K/Akt pathway, which offers a novel modality as to how the drug exerts protective effects.

The neuroprotective effect of DBZIM, a novel imidazolium compound, has previously been documented to slow down neurodegeneration in a mouse model of Parkinson's disease. In this study, we conducted behavioural studies and further investigated the neuroprotection in a rat Parkinsonian model induced by 6-hydroxydopamine (6-OHDA). DBZIM was found to significantly reduce the 6-OHDA-induced asymmetrical rotation and preferential usage of contralateral forelimbs. Furthermore, the degeneration of tyrosine hydroxylase immunopositive (TH+) dopaminergic neurones in the substantia nigra par compacta (SNc) was illustrated by immunohistochemistry. The significant loss of TH+ neurones by 6-OHDA administration was ameliorated by three different doses of DBZIM treatment in a bell-shape manner. Such neuroprotection was also observed in the 6-OHDA-lesioned striata. High-performance liquid chromatography (HPLC) analysis of the striatal tissues revealed that DBZIM beneficially maintained the dopamine level by slowing down its metabolism. In addition, DBZIM attenuated the activation of astrocytes and microglia. This suggests that anti-inflammation may be an additional mechanism underlying the DBZIM-mediated neuroprotection. These findings warrant further investigation of DBZIM as a promising and potent agent for the future treatment of Parkinson's disease.

The activation of N-methyl-d-aspartate (NMDA) receptors by excessive release of glutamate is involved in the pathogenesis of ischemic stroke. Thus the NMDA receptor has become an attractive therapeutic target for the development of neuroprotectants, especially from antagonists with moderate to low affinity. In the current study, NMDA receptor blockage and neuroprotective effects of bis(12)-hupyridone (B12H), a novel dimeric acetylcholinesterase inhibitor derived from a naturally occurring monomeric analog huperzine A, were investigated in vitro and in vivo. In primary rat cerebellar granule neurons, B12H (0.1 nM to 1 μM) prevented glutamate-induced apoptosis in a concentration- and time-dependent manner. Receptor-ligand binding analysis showed that B12H competed with [(3)H]MK801 with a K(i) value of 7.7 μM. In the 2-hour middle cerebral artery occlusion rat model, B12H (0.4 and 0.8 mg/kg, 30 min before-ischemia and 15 min post-ischemia, i.p.) significantly attenuated ischemia-induced apoptosis in the penumbra region, improved neurological behavior impairment, and decreased cerebral infarct volume, cerebral edema and neuronal apoptosis in the stroke model. Together, these results showed that B12H moderately blocks NMDA receptors at MK801 site and exerts neuroprotection against excitotoxic and ischemic insults in vitro and in vivo. Combined with our previous publications, we conjecture that B12H might exert neuroprotection via acting on multiple targets.