Patric K. Stanton

Chronic Traumatic Encephalopathy in Blast-Exposed Military Veterans and a Blast Neurotrauma Mouse Model

Blast exposure is associated with chronic traumatic encephalopathy, impaired neuronal function, and persistent cognitive deficits in blast-exposed military veterans and experimental animals. Blast Brain: An Invisible Injury Revealed Traumatic brain injury (TBI) is the “signature” injury of the conflicts in Afghanistan and Iraq and is associated with psychiatric symptoms and long-term cognitive disability. Recent estimates indicate that TBI may affect 20% of the 2.3 million U.S. servicemen and women deployed since 2001. Chronic traumatic encephalopathy (CTE), a tau protein–linked neurodegenerative disorder reported in athletes with multiple concussions, shares clinical features with TBI in military personnel exposed to explosive blast. However, the connection between TBI and CTE has not been explored in depth. In a new study, Goldstein et al. investigate this connection in the first case series of postmortem brains from U.S. military veterans with blast exposure and/or concussive injury. They report evidence for CTE neuropathology in the military veteran brains that is similar to that observed in the brains of young amateur American football players and a professional wrestler. The investigators developed a mouse model of blast neurotrauma that mimics typical blast conditions associated with military blast injury and discovered that blast-exposed mice also demonstrate CTE neuropathology, including tau protein hyperphosphorylation, myelinated axonopathy, microvascular damage, chronic neuroinflammation, and neurodegeneration. Surprisingly, blast-exposed mice developed CTE neuropathology within 2 weeks after exposure to a single blast. In addition, the neuropathology was accompanied by functional deficits, including slowed axonal conduction, reduced activity-dependent long-term synaptic plasticity, and impaired spatial learning and memory that persisted for 1 month after exposure to a single blast. The investigators then showed that blast winds with velocities of more than 330 miles/hour—greater than the most intense wind gust ever recorded on earth—induced oscillating head acceleration of sufficient intensity to injure the brain. The researchers then demonstrated that blast-induced learning and memory deficits in the mice were reduced by immobilizing the head during blast exposure. These findings provide a direct connection between blast TBI and CTE and indicate a primary role for blast wind–induced head acceleration in blast-related neurotrauma and its aftermath. This study also validates a new blast neurotrauma mouse model that will be useful for developing new diagnostics, therapeutics, and rehabilitative strategies for treating blast-related TBI and CTE. Blast exposure is associated with traumatic brain injury (TBI), neuropsychiatric symptoms, and long-term cognitive disability. We examined a case series of postmortem brains from U.S. military veterans exposed to blast and/or concussive injury. We found evidence of chronic traumatic encephalopathy (CTE), a tau protein–linked neurodegenerative disease, that was similar to the CTE neuropathology observed in young amateur American football players and a professional wrestler with histories of concussive injuries. We developed a blast neurotrauma mouse model that recapitulated CTE-linked neuropathology in wild-type C57BL/6 mice 2 weeks after exposure to a single blast. Blast-exposed mice demonstrated phosphorylated tauopathy, myelinated axonopathy, microvasculopathy, chronic neuroinflammation, and neurodegeneration in the absence of macroscopic tissue damage or hemorrhage. Blast exposure induced persistent hippocampal-dependent learning and memory deficits that persisted for at least 1 month and correlated with impaired axonal conduction and defective activity-dependent long-term potentiation of synaptic transmission. Intracerebral pressure recordings demonstrated that shock waves traversed the mouse brain with minimal change and without thoracic contributions. Kinematic analysis revealed blast-induced head oscillation at accelerations sufficient to cause brain injury. Head immobilization during blast exposure prevented blast-induced learning and memory deficits. The contribution of blast wind to injurious head acceleration may be a primary injury mechanism leading to blast-related TBI and CTE. These results identify common pathogenic determinants leading to CTE in blast-exposed military veterans and head-injured athletes and additionally provide mechanistic evidence linking blast exposure to persistent impairments in neurophysiological function, learning, and memory.

GLYX-13, a NMDA Receptor Glycine-Site Functional Partial Agonist, Induces Antidepressant-Like Effects Without Ketamine-Like Side Effects

Recent human clinical studies with the NMDA receptor (NMDAR) antagonist ketamine have revealed profound and long-lasting antidepressant effects with rapid onset in several clinical trials, but antidepressant effects were preceded by dissociative side effects. Here we show that GLYX-13, a novel NMDAR glycine-site functional partial agonist, produces an antidepressant-like effect in the Porsolt, novelty induced hypophagia, and learned helplessness tests in rats without exhibiting substance abuse-related, gating, and sedative side effects of ketamine in the drug discrimination, conditioned place preference, pre-pulse inhibition and open-field tests. Like ketamine, the GLYX-13-induced antidepressant-like effects required AMPA/kainate receptor activation, as evidenced by the ability of NBQX to abolish the antidepressant-like effect. Both GLYX-13 and ketamine persistently (24 h) enhanced the induction of long-term potentiation of synaptic transmission and the magnitude of NMDAR-NR2B conductance at rat Schaffer collateral-CA1 synapses in vitro. Cell surface biotinylation studies showed that both GLYX-13 and ketamine led to increases in both NR2B and GluR1 protein levels, as measured by Western analysis, whereas no changes were seen in mRNA expression (microarray and qRT-PCR). GLYX-13, unlike ketamine, produced its antidepressant-like effect when injected directly into the medial prefrontal cortex (MPFC). These results suggest that GLYX-13 produces an antidepressant-like effect without the side effects seen with ketamine at least in part by directly modulating NR2B-containing NMDARs in the MPFC. Furthermore, the enhancement of ‘metaplasticity’ by both GLYX-13 and ketamine may help explain the long-lasting antidepressant effects of these NMDAR modulators. GLYX-13 is currently in a Phase II clinical development program for treatment-resistant depression.

A human intracranial study of long-range oscillatory coherence across a frontal-occipital-hippocampal brain network during visual object processing.

Visual object-recognition is thought to involve activation of a distributed network of cortical regions, nodes of which include the lateral prefrontal cortex, the so-called lateral occipital complex (LOC), and the hippocampal formation. It has been proposed that long-range oscillatory synchronization is a major mode of coordinating such a distributed network. Here, intracranial recordings were made from three humans as they performed a challenging visual object-recognition task that required them to identify barely recognizable fragmented line-drawings of common objects. Subdural electrodes were placed over the prefrontal cortex and LOC, and depth electrodes were placed within the hippocampal formation. Robust beta-band coherence was evident in all subjects during processing of recognizable fragmented images. Significantly lower coherence was evident during processing of unrecognizable scrambled versions of the same. The results indicate that transient beta-band oscillatory coupling between these three distributed cortical regions may reflect a mechanism for effective communication during visual object processing.

Blockade of Gap Junctions In Vivo Provides Neuroprotection After Perinatal Global Ischemia

Background and Purpose— We investigated the contribution of gap junctions to brain damage and delayed neuronal death produced by oxygen-glucose deprivation (OGD). Methods— Histopathology, molecular biology, and electrophysiological and fluorescence cell death assays in slice cultures after OGD and in developing rats after intrauterine hypoxia-ischemia (HI). Results— OGD persistently increased gap junction coupling and strongly activated the apoptosis marker caspase-3 in slice cultures. The gap junction blocker carbenoxolone applied to hippocampal slice cultures before, during, or 60 minutes after OGD markedly reduced delayed neuronal death. Administration of carbenoxolone to ischemic pups immediately after intrauterine HI prevented caspase-3 activation and dramatically reduced long-term neuronal damage. Conclusions— Gap junction blockade may be a useful therapeutic tool to minimize brain damage produced by perinatal and early postnatal HI.

BDNF increases release probability and the size of a rapidly recycling vesicle pool within rat hippocampal excitatory synapses

Exerting its actions pre‐, post‐ and peri‐synaptically, brain‐derived neurotrophic factor (BDNF) is one of the most potent modulators of hippocampal synaptic function. Here, we examined the effects of BDNF on a rapidly recycling pool (RRP) of vesicles within excitatory synapses. First, we estimated vesicular release in hippocampal cultures by performing FM4‐64 imaging in terminals impinging on enhanced green fluorescent protein (eGFP)‐labelled dendritic spines – a hallmark of excitatory synapses. Consistent with a modulation of the RRP, BDNF increased the evoked destaining rate of FM4‐64 only during the initial phase of field stimulation. Multiphoton microscopy in acute hippocampal slices confirmed these observations by selectively imaging the RRP, which was loaded with FM1‐43 by hyperosmotic shock. Slices exposed to BDNF showed an increase in the evoked and spontaneous rates of FM1‐43 destaining from terminals in CA1 stratum radiatum, mostly representing excitatory terminals of Schaffer collaterals. Variance‐mean analysis of evoked EPSCs in CA1 pyramidal neurons further confirmed that release probability is increased in BDNF‐treated slices, without changes in the number of independent release sites or average postsynaptic quantal amplitude. Because BDNF was absent during dye loading, imaging, destaining and whole‐cell recordings, these results demonstrate that BDNF induces a long‐lasting enhancement in the probability of transmitter release at hippocampal excitatory synapses by modulating the RRP. Since the endogenous BDNF scavenger TrkB‐IgG prevented the enhancement of FM1‐43 destaining rate caused by induction of long‐term potentiation in acute hippocampal slices, the modulation of a rapidly recycling vesicle pool may underlie the role of BDNF in hippocampal long‐term synaptic plasticity.

Alterations in metabolism and gap junction expression may determine the role of astrocytes as “good samaritans” or executioners

Our knowledge of astroglia and their physiological and pathophysiological role(s) in the central nervous system (CNS) has grown during the past decade, revealing a complex picture. It is becoming increasingly clear that glia play a significant role in the homeostasis and function of the CNS and that neurons should no longer be considered the only cell type that responds, both rapidly and slowly, to electrochemical activity. We discuss recent advances in the field with an emphasis on the impact of hypoxia and ischemia on astrocytic metabolism and the functional relationship between glucose metabolism and gap junctions in astrocytes. We also address the controversy over whether astrocytic gap junctions mediate protection or killing of neurons during or after hypoxic or ischemic insults.

The N-methyl-d-aspartate receptor modulator GLYX-13 enhances learning and memory, in young adult and learning impaired aging rats

NMDA receptor (NMDAR) activity has been strongly implicated in both in vitro and in vivo learning models and the decline in cognitive function associated with aging and is linked to a decrease in NMDAR functional expression. GLYX-13 is a tetrapeptide (Thr-Pro-Pro-Thr) which acts as a NMDAR receptor partial agonist at the glycine site. GLYX-13 was administered to young adult (3 months old) and aged (27-32 months old) Fischer 344 X Brown Norway F1 rats (FBNF1), and behavioral learning tested in trace eye blink conditioning (tEBC), a movable platform version of the Morris water maze (MWM), and alternating t-maze tasks. GLYX-13 (1mg/kg, i.v.) enhanced learning in both young adult and aging animals for MWM and alternating t-maze, and increased tEBC in aging rats. We previously showed optimal enhancement of tEBC in young adult rats given GLYX-13 at the same dose. Of these learning tasks, the MWM showed the most robust age related deficit in learning. In the MWM, GLYX-13 enhancement of learning was greater in the old compared to the young adult animals. Examination of the induction of long-term potentiation (LTP) and depression (LTD) at Schaffer collateral-CA1 synapses in hippocampal slices showed that aged rats showed marked, selective impairment in the magnitude of LTP evoked by a sub-maximal tetanus, and that GLYX-13 significantly enhanced the magnitude of LTP in slices from both young adult and aged rats without affecting LTD. These data, combined with the observation that the GLYX-13 enhancement of learning was greater in old than in young adult animals, suggest that GLYX-13 may be a promising treatment for deficits in cognitive function associated with aging.

A NMDA receptor glycine site partial agonist, GLYX-13, simultaneously enhances LTP and reduces LTD at Schaffer collateral–CA1 synapses in hippocampus

N-methyl-D-aspartate glutamate receptors (NMDARs) are a key route for Ca2+ influx into neurons important to both activity-dependent synaptic plasticity and, when uncontrolled, triggering events that cause neuronal degeneration and death. Among regulatory binding sites on the NMDAR complex is a glycine binding site, distinct from the glutamate binding site, which must be co-activated for NMDAR channel opening. We developed a novel glycine site partial agonist, GLYX-13, which is both nootropic and neuroprotective in vivo. Here, we assessed the effects of GLYX-13 on long-term synaptic plasticity and NMDAR transmission at Schaffer collateral-CA1 synapses in hippocampal slices in vitro. GLYX-13 simultaneously enhanced the magnitude of long-term potentiation (LTP) of synaptic transmission, while reducing long-term depression (LTD). GLYX-13 reduced NMDA receptor-mediated synaptic currents in CA1 pyramidal neurons evoked by low frequency Schaffer collateral stimulation, but enhanced NMDAR currents during high frequency bursts of activity, and these actions were occluded by a saturating concentration of the glycine site agonist d-serine. Direct two-photon imaging of Schaffer collateral burst-evoked increases in [Ca2+] in individual dendritic spines revealed that GLYX-13 selectively enhanced burst-induced NMDAR-dependent spine Ca2+ influx. Examining the rate of MK-801 block of synaptic versus extrasynaptic NMDAR-gated channels revealed that GLYX-13 selectively enhanced activation of burst-driven extrasynaptic NMDARs, with an action that was blocked by the NR2B-selective NMDAR antagonist ifenprodil. Our data suggest that GLYX-13 may have unique therapeutic potential as a learning and memory enhancer because of its ability to simultaneously enhance LTP and suppress LTD.

Imaging LTP of presynaptic release of FM1-43 from the rapidly recycling vesicle pool of Schaffer collateral-CA1 synapses in rat hippocampal slices.

Recent studies using the styryl dye FM1‐43 and two‐photon microscopy to directly visualize transmitter release at CA3–CA1 excitatory synapses in the hippocampus have demonstrated that activity‐dependent long‐term potentiation (LTP) and long‐term depression are associated with alterations in vesicular release. It is not known whether particular vesicle pools preferentially express these alterations or what second messenger cascades are involved. To address these questions, we selectively loaded FM1‐43 into the rapidly recycling pool (RRP) of vesicles by use of a brief hypertonic shock to release and load the RRP. We demonstrate here that the induction of LTP can lead to a selective long‐lasting enhancement in presynaptic release from the RRP, while reserve pool kinetics remain unchanged. LTP of RRP release was N‐methyl‐d‐aspartate receptor‐dependent and also required production of the intercellular messenger NO and activation of receptor tyrosine kinase. Measurement of FM1‐43 stimulus‐evoked uptake rates following induction of LTP confirmed that LTP produces more rapid recycling of vesicles released by electrical stimulation, consistent with an enhanced release probability from the RRP.

GLYX-13, an NMDA receptor glycine site functional partial agonist enhances cognition and produces antidepressant effects without the psychotomimetic side effects of NMDA receptor antagonists

Introduction: The N-methyl-d-aspartate receptor-ionophore complex plays a key role in learning and memory and has efficacy in animals and humans with affective disorders. GLYX-13 is an N-methyl-d-aspartate receptor (NMDAR) glycine-site functional partial agonist and cognitive enhancer that also shows rapid antidepressant activity without psychotomimetic side effects. Areas covered: The authors review the mechanism of action of GLYX-13 that was investigated in preclinical studies and evaluated in clinical studies. Specifically, the authors review its pharmacology, pharmacokinetics, and drug safety that were demonstrated in clinical studies. Expert opinion: NMDAR full antagonists can produce rapid antidepressant effects in treatment-resistant subjects; however, they are often accompanied by psychotomimetic effects that make chronic use outside of a clinical trial inpatient setting problematic. GLYX-13 appears to exert its antidepressant effects in the frontal cortex via NMDAR-triggered synaptic plasticity. Understanding the mechanistic underpinning of GLYX-13s antidepressant action should provide both novel insights into the role of the glutamatergic system in depression and identify new targets for therapeutic development.