The GABA Synapse as a Target for Antiepileptic Drugs: A Historical Overview Focused on GABA Transporters

It is clear that normal neuronal function relies on a tight balance between excitatory and inhibitory neurotransmission. Inhibitory signaling through the GABAergic system can be tightly regulated at the level of GABA uptake via GABA transporters (GAT). As such, selectively modulating the GABA uptake process through pharmacological agents has been an area of active investigation over several decades. These studies have demonstrated that inhibition of astroglial, but not neuronal, GATs may be preferred for anticonvulsant action. To date, four distinct GAT subtypes have been identified and efforts to selectively target these transporters have led to the proliferation of pharmacological agents aimed at augmenting extrasynaptic GABA levels. These pharmacological tools have provided novel and informative insight into the role of GABA and GABAergic signaling in the brain, but have also provided critical information concerning the regulation of CNS disorders associated with an imbalance in inhibitory tone, such as epilepsy. One such compound with notable inhibitory effects at GATs, tiagabine, has demonstrated clinical anticonvulsant efficacy, and is, to date, the only approved GAT inhibitor for clinical use. Thus, efforts to identify and develop GAT subtype-specific compounds continue to be an area of active investigation for the management of epilepsy and other CNS disorders. Herein, the historical efforts to elucidate the role of GABA in the synapse, as well as the role of GAT inhibitors as anticonvulsants, are described.     

Brain-derived neurotrofic factor (BDNF) secretion of human mesenchymal stem cells isolated from bone marrow,endometrium and adipose tissue

The ability of mesenchymal stem cells (MSCs) to differentiate into neuronal lineage determines the potential of these cells as a substrate for a cell replacement therapy. In this paper we compare the neurogenic potential of the MSCs from different donors, isolated from the bone marrow (BMSC), subcutaneous adipose tissue (AD MSC) and menstrual blood (eMSC). It was established that the native eMCSs, BMSCs and AD MSCs express neuronal marker β-III-tubulin with a frequency of 90, 50 and 14%, respectively. Also we showed that the eMSCs have a high endogenous level of brain-derived neurotrophic factor (BDNF), whereas the BMSCs and the AD MSCs are characterized by low basal BDNF levels. An induction of neuronal differentiation in the studied MSCs using differentiation medium containing B27 and N2 supplements, 5-azacytidine, retinoic acid, IBMX and dbcAMP induced changes in the cells morphology, the increase of β-III-tubulin expression, and the appearance of neuronal markers GFAP, NF-H, NeuN and MAP2. During the differentiation the BDNF secretion was significantly enhanced in the BMSCs and decreased in the eMSCs cultures. However, no correlation between the basal and induced levels of the neuronal markers expression in the studied MSCs has been established.     

Neural Stem Cell Transplants Improve Cognitive Function Without Altering Amyloid Pathology in an APP/PS1 Double Transgenic Model of Alzheimer’s Disease

Neural stem cells (NSCs) are capable of self-renewal and are multipotent. Transplantation of NSCs may represent a promising approach for treating neurodegenerative disorders associated with cognitive decline, such as Alzheimer disease (AD) characterized by extensive loss of neurons. In this study, we investigated the effect of NSC transplantation on cognitive function in the amyloid precursor protein/presenilin-1 (APP/PS1) transgenic mouse, an AD mouse model with age-dependent cognitive deficits. We found that NSCs bilaterally transplanted into hippocampal regions improved spatial learning and memory function in these mice, but did not alter Aβ pathology. Immunohistochemical analyses determined that NSCs proliferated, migrated, and differentiated into three neuronal cell types. The improvement in cognitive function was correlated with enhanced long-term potentiation (LTP) and an increase in the neuron expression of proteins related to cognitive function: N-methyl-d-aspartate (NMDA) 2B unit, synaptophysin (SYP), protein kinase C ζ subtypes (PKCζ), tyrosine receptor kinase B (TrkB), and brain-derived neurotrophic factor (BDNF). Taken together, our data indicated that injected NSCs can rescue cognitive deficits in APP/PS1 transgenic mice by replacing neuronal cell types expressing multiple cognition-related proteins that enhance LTP.     

Spatiotemporal pattern of TRAF3 expression after rat spinal cord injury

TNF receptor associated factor 3 (TRAF3), a member of the TRAF family of intracellular signaling proteins, can directly influence the phosphorylation status and activation of c-Jun N-terminal kinase, participating in CD40-induced apoptosis in carcinoma. However, its expression profile and function are still unclear in spinal cord injury (SCI). In this study, we performed an acute spinal cord contusion injury model in adult rats and detected the dynamic change patterns of TRAF3 expression in spinal cord. Western blot and immunohistochemistry revealed a striking upregulation of TRAF3 after SCI. Double immunofluorescence staining prompted that TRAF3 immunoreactivity was found in neurons rather than astrocytes. Moreover, co-localization of TRAF3/active caspase-3 was detected in neuronal nuclei. To further investigate the function of TRAF3, a neuronal cell line PC12 was employed to establish an apoptosis model in vitro. We analyzed the association of TRAF3 with active caspase-3 on PC12 cells by western blot and immunofluorescent labeling, which was parallel with the data in vivo. Additionally, knocking TRAF3 down with siRNA demonstrated the probable pro-apoptotic role of TRAF3 in the process of neuronal apoptosis. To summarize, we firstly uncover the temporal and spatial expression changes of TRAF3 in SCI. Our data suggest that TRAF3 might be implicated in central nervous system pathophysiology after SCI.     

Protective Effects of Humanin on Okadaic Acid-Induced Neurotoxicities in Cultured Cortical Neurons

Neurofibrillary tangles are pathological hallmarks of Alzheimer’s disease (AD), which are mostly composed of hyperphosphorylated tau and directly correlate with dementia in AD patients. Okadaic acid (OA), a toxin extracted from marine life, can specifically inhibit protein phosphatases (PPs), including PP1 and Protein phosphatase 2A (PP2A), resulting in tau hyperphosphorylation. Humanin (HN), a peptide of 24 amino acids, was initially reported to protect neurons from AD-related cell toxicities. The present study was designed to test if HN could attenuate OA-induced neurotoxicities, including neural insults, apoptosis, autophagy, and tau hyperphosphorylation. We found that administration of OA for 24 h induced neuronal insults, including lactate dehydrogenase released, decreased of cell viability and numbers of living cells, neuronal apoptosis, cells autophagy and tau protein hyperphosphorylation. Pretreatment of cells with HN produced significant protective effects against OA-induced neural insults, apoptosis, autophagy and tau hyperphosphorylation. We also found that OA treatment inhibited PP2A activity and HN pretreatment significantly attenuated the inhibitory effects of OA. This study demonstrated for the first time that HN protected cortical neurons against OA-induced neurotoxicities, including neuronal insults, apoptosis, autophagy, and tau hyperphosphorylation. The mechanisms underlying the protections of HN may involve restoration of PP2A activity.     

Morphological approach to assess the involvement of astrocytes in prion propagation

Transmissible Spongiform Encephalopathies (TSEs) are a group of neurodegenerative disorders affecting animals and humans and for which no effective treatment is available to date. Vacuolation, neuronal/neurite degeneration, deposition of pathological prion protein (PrPsc) and gliosis are changes typically found in brains from TSE affected individuals. However, the actual role of this last feature, microgliosis and astrocytosis, has not been precisely determined. The overall objective of this work is to assess the involvement of glial cells as components of the host protective system in prion propagation; specifically, to analyze the behavior of astroglial cells in prion progression. To achieve this aim, histopathological and immunohistochemical techniques were carried out on samples from cerebella using Scrapie as the prototype of natural TSEs as this made it possible to assess different stages of the disease; specifically, ages and genotypes from Scrapie-affected animals corresponding to different sources, by using optical, confocal and electron microscopy. The results provided in the present study demonstrate the indisputable involvement of astroglia in prion progression by showing specific changes of this glial population matching up to the evolution of the disease. Moreover, cerebellar lesions mainly associated to Purkinje cells that have not previously been reported in animal prion diseases in natural transmission are described here. The close relationship between PrPsc and GFAP hiperimmunoreactivity and Purkinje cells, alongside the evident thickening of their neurites at terminal stages demonstrated in this study, suggest that these neurons are the main target of this neurodegenerative disease.     

CIIA negatively regulates neuronal cell death induced by oxygen–glucose deprivation and reoxygenation

Brain ischemia causes neuronal injury leading to stroke and other related brain diseases. However, the precise mechanism of the ischemia-induced neuronal death remains unclear yet. In this study, we showed that CIIA suppressed neuronal cell death induced by oxygen and glucose deprivation followed by reoxygenation (OGD/R), which mimics ischemia and reperfusion in vivo, in neuroblastoma cell lines as well as primary cortical neurons. Furthermore, CIIA inhibited the OGD/R-induced stimulation of apoptosis signal-regulating kinase 1 (ASK1) and its downstream kinases including c-Jun amino-terminal kinase and p38 kinase, concomitantly blocking ASK1 homo-oligomerization and the binding between ASK1 and TRAF2. CIIA also repressed the OGD/R-induced activation of caspase-3 in neuronal cells. Taken together, our results suggest that CIIA attenuates neurotoxicity caused by OGD/R through inhibiting ASK1-dependent signaling events.     

Rutin Alleviates Prion Peptide-Induced Cell Death Through Inhibiting Apoptotic Pathway Activation in Dopaminergic Neuronal Cells

Prion disorders are progressive neurodegenerative diseases characterized by extensive neuronal loss and accumulation of the abnormal form of the scrapie prion protein (PrP). Rutin is a flavonoid that occurs naturally in plant-derived beverages and foods and is used in traditional and folkloric medicine worldwide. In the present study, we evaluated the protective effects of rutin against PrP fragment (106–126)-induced neuronal cell death. Rutin treatment blocked PrP(106–126)-mediated increases in reactive oxygen species production and nitric oxide release and helped slowing the decrease of neurotrophic factors that results from PrP accumulation. Rutin attenuated PrP(106–126)-associated mitochondrial apoptotic events by inhibiting mitochondrial permeability transition and caspase-3 activity and blocking expression of the apoptotic signals Bax and PARP. Additionally, rutin treatment significantly decreased the expression of the death receptor Fas and its ligand Fas-L. Overall, our results demonstrated that rutin protects against the neurodegenerative effects of prion accumulation by increasing production of neurotropic factors and inhibiting apoptotic pathway activation in neuronal cells. These results suggested that rutin may have clinical benefits for prion diseases and other neurodegenerative disorders.     

Effect of Transient Cerebral Ischemia on the Expression of Receptor for Advanced Glycation End Products (RAGE) in the Gerbil Hippocampus Proper

The receptor for advanced glycation end products (RAGE) is a multi-ligand receptor of the immunoglobulin superfamily that has been implicated in multiple neuronal and inflammatory stress processes. In this study, we examined changes in RAGE immunoreactivity and its protein levels in the gerbil hippocampus (CA1-3 regions) after 5 min of transient global cerebral ischemia. The ischemic hippocampus was stained with cresyl violet, neuronal nuclei (a neuron-specific soluble nuclear antigen) antibody and Fluoro-Jade B (a marker for neuronal degeneration). 5 days after ischemia–reperfusion, delayed neuronal death occurred in the stratum pyramidale of the CA1 region. RAGE immunoreactivity was not detected in any regions of the CA1-3 regions of the sham-group; the immunoreactivity was markedly increased only in the CA1 region from 3 days after ischemia–reperfusion. On the other hand, RAGE immunoreactivity was newly expressed in astrocytes, not in microglia. Western blot analysis showed that RAGE protein level was highest at 5 days post-ischemia. In brief, both the RAGE immunoreactivity and protein level were distinctively increased in astrocytes in the ischemic CA1 region from 3 days after transient cerebral ischemia. These results indicate that the increase of RAGE expression in astrocytes after ischemia–reperfusion may be related to the ischemia-caused activation of astrocytes in the ischemic CA1 region.     

Intromitochondrial IκB/NF-κB signaling pathway is involved in amyloid β peptide-induced mitochondrial dysfunction

Mitochondrial dysfunction is a hallmark of amyloid β peptide (Aβ)-induced neuronal toxicity in Alzheimer’s disease (AD). However, the underlying mechanism (s) of Aβ-induced mitochondrial dysfunction is still not fully understood. There is evidence that nuclear factor-κB (NF-κB) is involved in Aβ-induced neurotoxicity and is present in mitochondria. Using HT22 murine hippocampal neuronal cells and isolated mitochondria, the present study investigated whether intramitochondrial inhibitor of NF-κB (IκB)/NF-κB signaling pathway was involved in mitochondrial dysfunction induced by Aβ. It was found that Aβ impaired mitochondrial function through a NF-κB-dependent signaling pathway. Intramitochondrial IκBα/NF-κB pathway, induced by Aβ, decreased the expression of cytochrome c oxidase subunit (COXIII) and inhibited COX activity. These results provide new insights into the mechanism underlying the neurotoxic effect of Aβ and open up new therapeutic perspectives for AD.     

Up-regulation of SKIP relates to retinal ganglion cells apoptosis after optic nerve crush in vivo

Cell cycle re-entry is one of the key processes in neuronal apoptosis. Previous studies have shown that Ski-interacting protein (SKIP) played an important role in cell cycle re-entry. However, its expression and function in optic nerve injury are still with limited acquaintance. To investigate whether SKIP is involved in retinal ganglion cells (RGCs) death, we performed an optic nerve crush (ONC) model in adult rats. Western blot analysis revealed that up-regulation of SKIP was present in retina at 5 days after ONC. Immunofluorescent labeling indicated that up-regulated SKIP was found mainly in RGCs. We also investigated co-localization of SKIP with active-caspase-3 and TUNEL (apoptotic markers) -positive cells in the retina after ONC. In addition, the expression of SKIP was increased in parallel with P53 and P21 in retina after ONC. All these results suggested that up-regulation of SKIP in the retina was associated with RGCs death after ONC.     

Progression of O6-methylguanine-DNA methyltransferase and temozolomide resistance in cancer research

Temozolomide (TMZ) is an alkylating agent that is widely used in chemotherapy for cancer. A key mechanism of resistance to TMZ is the overexpression of O⁶-methylguanine-DNA methyltransferase (MGMT). MGMT specifically repairs the DNA O⁶-methylation damage induced by TMZ and irreversibly inactivates TMZ. Regulation of MGMT expression and research regarding the mechanism of TMZ resistance will help rationalize the clinical use of TMZ. In this review, we provide an overview of recent advances in the field, with particular emphasis on MGMT structure, function, expression regulation, and the association between MGMT and resistance to TMZ.     

Dynamin-related protein 1 is required for normal mitochondrial bioenergetic and synaptic function in CA1 hippocampal neurons

Disrupting particular mitochondrial fission and fusion proteins leads to the death of specific neuronal populations; however, the normal functions of mitochondrial fission in neurons are poorly understood, especially in vivo, which limits the understanding of mitochondrial changes in disease. Altered activity of the central mitochondrial fission protein dynamin-related protein 1 (Drp1) may contribute to the pathophysiology of several neurologic diseases. To study Drp1 in a neuronal population affected by Alzheimer''s disease (AD), stroke, and seizure disorders, we postnatally deleted Drp1 from CA1 and other forebrain neurons in mice (CamKII-Cre, Drp1^lox/lox (Drp1cKO)). Although most CA1 neurons survived for more than 1 year, their synaptic transmission was impaired, and Drp1cKO mice had impaired memory. In Drp1cKO cell bodies, we observed marked mitochondrial swelling but no change in the number of mitochondria in individual synaptic terminals. Using ATP FRET sensors, we found that cultured neurons lacking Drp1 (Drp1KO) could not maintain normal levels of mitochondrial-derived ATP when energy consumption was increased by neural activity. These deficits occurred specifically at the nerve terminal, but not the cell body, and were sufficient to impair synaptic vesicle cycling. Although Drp1KO increased the distance between axonal mitochondria, mitochondrial-derived ATP still decreased similarly in Drp1KO boutons with and without mitochondria. This indicates that mitochondrial-derived ATP is rapidly dispersed in Drp1KO axons, and that the deficits in axonal bioenergetics and function are not caused by regional energy gradients. Instead, loss of Drp1 compromises the intrinsic bioenergetic function of axonal mitochondria, thus revealing a mechanism by which disrupting mitochondrial dynamics can cause dysfunction of axons.Mitochondrial dynamics – the balance between mitochondrial fission and fusion – regulates mitochondrial quality control by segregating poorly functioning mitochondria for degradation while mixing the contents of healthy mitochondria.^{1, 2} In neurons, fission uniquely facilitates movement of mitochondria down narrow distal axons.^{3, 4} Disruptions of this movement, and of other neuron-specific functions, may explain why systemic mutations in mitochondrial fusion and fission proteins specifically cause death of neurons. However, the roles and requirements of these proteins also differ between neuronal types.¹ For example, mutations in the fusion protein optic atrophy 1 cause degeneration of retinal ganglion neurons,⁵ and mutations in the fusion protein mitofusin-2 or the fission protein ganglioside-induced differentiation-associated protein 1 cause peripheral neuropathy (Charcot-Marie-Tooth types 2A and 4A^{6, 7}).There are several potential reasons why specific neurons have unique requirements for fission–fusion proteins. First, the functions of these proteins may be more critical in vulnerable neuronal populations. Recently, we showed that most midbrain DA neurons are uniquely vulnerable to loss of the central mitochondrial fission protein dynamin-related protein 1 (Drp1),⁴ a GTPase recruited to fission sites on the outer mitochondrial membrane.¹ Loss of Drp1 depletes axonal mitochondria, which is followed by axonal degeneration and neuronal death. However, a subpopulation of midbrain DA neurons survive, despite losing their axonal mitochondria, suggesting that they have lower needs for energy or other mitochondrial functions in their axons.⁴ Do unique requirements for mitochondrial dynamics underlie differential neuronal vulnerability? Do resistant neurons compensate with other fission or fusion mechanisms? Do the functions of fission differ between neurons? Notably, Drp1 may also have mitochondria-independent functions in synaptic vesicle release.⁸ Addressing these issues could help elucidate the physiological functions of mitochondrial dynamics in the nervous system and reveal how shifts in the fission–fusion balance contribute to selective neuronal death in neurodegenerative diseases, including Huntington''s disease, Parkinson''s disease and Alzheimer''s disease (AD),^{1, 4} and in other neurologic disorders, including stroke and epilepsy.^{9, 10, 11}To understand mitochondrial dynamics, it would be useful to know why mitochondrial fission is needed in the nervous system in the first place, and how loss of fission affects mitochondrial functions in specific cell types. Notably, Drp1 knockout did not change respiration or ATP levels in resuspended mouse embryonic fibroblasts (MEFs),^{12, 13} indicating that mitochondrial fission is not required for respiration in these cells. However, neuronal respiration may be more sensitive to Drp1 loss. Indeed, Drp1 loss markedly decreased the number of mitochondria in axons and the cell body in midbrain DA neurons in vivo,⁴ and reduced staining of complex I and IV activity in cerebellar neurons in vivo.¹⁴ However, it is unclear whether these changes translate into decreased ATP levels in neurons and, if so, whether this decrease compromises neuronal function. Furthermore, Drp1 loss caused cell death in cerebellar and most midbrain DA neurons,^{4, 14} which challenges our ability to dissociate the specific effects of Drp1 loss on mitochondrial function from other non-specific changes that accompany cell death.To learn how disrupting mitochondrial fission contributes to selective neurodegeneration, we studied the function of Drp1 in CA1 hippocampal neurons and its role in mitochondrial bioenergetics. Surprisingly, despite losing Drp1, most CA1 neurons survived for more than 1 year in vivo, although their function was compromised, leading to deficits in synaptic transmission and memory. To begin to understand how loss of Drp1 causes neuronal dysfunction, we examined the role of Drp1 in mitochondrial bioenergetics. We found that Drp1 is required to maintain normal mitochondrial-derived ATP levels specifically in axons (but not the cell body), and that the loss of this function is unrelated to the distribution of mitochondria within axons.     

Hypothesis: Tim-3/Galectin-9, A New Pathway for Leukemia Stem Cells Survival by Promoting Expansion of Myeloid-Derived Suppressor Cells and Differentiating into Tumor-Associated Macrophages

Despite the improvements in chemotherapy, about 60 % of acute myeloid leukemia (AML) remission patients still relapse. Leukemic stem cells (LSCs) are the main causes for the relapse and refractory. T cell immunoglobulin mucin-3 (TIM-3), a specific surface molecule expressed on LSCs in most types of AML, is a candidate for AML LSC-targeted therapies. It is important to know how this molecule functions in the maintenance of LSCs and suppression of anti-tumor immunity. Recent data have shown that Tim-3 which expresses on T cells can suppress immune responses indirectly by inducing expansion of myeloid-derived suppressor cells (MDSCs). MDSCs at the leukemia site can also differentiate into tumor-associated macrophages (TAMs). TAMs can promote proliferation and survival of LSCs by the diversion of adaptive immunity and the facilitation of extracellular matrix remodeling, angiogenesis, and lymphangiogenesis. Our previous study in AML patient bone marrow samples showed CD68⁺ macrophages around AML clone. Based on the known evidence and our experimental findings, we hypothesize that Tim-3, which specifically expresses on LSCs, is beneficial for LSCs survival and AML progression by promoting expansion of MDSCs and differentiating into TAMs at the leukemia site.     

Interaction of metal ions with neurotransmitter receptors and potential role in neurodiseases

There is increasing evidence that toxic metals play a role in diseases of unknown etiology. Their action is often mediated by membrane proteins, and in particular neurotransmitter receptors. This brief review will describe recent findings on the direct interaction of metal ions with ionotropic γ-aminobutyric acid (GABA_A) and glutamate receptors, the main inhibitory and excitatory neurotransmitter receptors in the mammalian central nervous system, respectively. Both hyper and hypo function of these receptors are involved in neurological and psychotic syndromes and modulation by metal ions is an important pharmacological issue. The focus will be on three xenobiotic metals, lead (Pb), cadmium (Cd) and nickel (Ni) that have no biological function and whose presence in living organisms is only detrimental, and two trace metals, zinc (Zn) and copper (Cu), which are essential for several enzymatic functions, but can mediate toxic actions if deregulated. Despite limited access to the brain and tight control by metalloproteins, exogenous metals interfere with receptor performances by mimicking physiological ions and occupying one or more modulatory sites on the protein. These interactions will be discussed as a potential cause of neuronal dysfunction.     

The Potential Role of Rho GTPases in Alzheimer's Disease Pathogenesis

Alzheimer's disease (AD) is characterized by a wide loss of synapses and dendritic spines. Despite extensive efforts, the molecular mechanisms driving this detrimental alteration have not yet been determined. Among the factors potentially mediating this loss of neuronal connectivity, the contribution of Rho GTPases is of particular interest. This family of proteins is classically considered a key regulator of actin cytoskeleton remodeling and dendritic spine maintenance, but new insights into the complex dynamics of its regulation have recently determined how its signaling cascade is still largely unknown, both in physiological and pathological conditions. Here, we review the growing evidence supporting the potential involvement of Rho GTPases in spine loss, which is a unanimously recognized hallmark of early AD pathogenesis. We also discuss some new insights into Rho GTPase signaling framework that might explain several controversial results that have been published. The study of the connection between AD and Rho GTPases represents a quite unchartered avenue that holds therapeutic potential.     

Granulocyte Colony Stimulating Factor Reduces Brain Injury in a Cardiopulmonary Bypass-Circulatory Arrest Model of Ischemia in a Newborn Piglet

Ischemic brain injury continues to be of major concern in patients undergoing cardiopulmonary bypass (CPB) surgery for congenital heart disease. Striatum and hippocampus are particularly vulnerable to injury during these processes. Our hypothesis is that the neuronal injury resulting from CPB and the associated circulatory arrest can be at least partly ameliorated by pre-treatment with granulocyte colony stimulating factor (G-CSF). Fourteen male newborn piglets were assigned to three groups: deep hypothermic circulatory arrest (DHCA), DHCA with G-CSF, and sham-operated. The first two groups were placed on CPB, cooled to 18 °C, subjected to 60 min of DHCA, re-warmed and recovered for 8–9 h. At the end of experiment, the brains were perfused, fixed and cut into 10 µm transverse sections. Apoptotic cells were visualized by in situ DNA fragmentation assay (TUNEL), with the density of injured cells expressed as a mean number ± SD per mm². The number of injured cells in the striatum and CA1 and CA3 regions of the hippocampus increased significantly following DHCA. In the striatum, the increase was from 0.46 ± 0.37 to 3.67 ± 1.57 (p = 0.002); in the CA1, from 0.11 ± 0.19 to 5.16 ± 1.57 (p = 0.001), and in the CA3, from 0.28 ± 0.25 to 2.98 ± 1.82 (p = 0.040). Injection of G-CSF prior to bypass significantly reduced the number of injured cells in the striatum and CA1 region, by 51 and 37 %, respectively. In the CA3 region, injured cell density did not differ between the G-CSF and control group. In a model of hypoxic brain insult associated with CPB, G-CSF significantly reduces neuronal injury in brain regions important for cognitive functions, suggesting it can significantly improve neurological outcomes from procedures requiring DHCA.