Project description:Epilepsy, a common neurological disorder, is featured with recurrent seizures. Its underlying pathological mechanisms remain elusive. Here, we provide evidence for loss of neogenin (NEO1), a coreceptor for multiple ligands, including netrins and bone morphological proteins, in the development of epilepsy. NEO1 is reduced in hippocampi from patients with epilepsy based on transcriptome and proteomic analyses. Neo1 knocking out (KO) in mouse brains displays elevated epileptiform spikes and seizure susceptibility. These phenotypes were undetectable in mice, with selectively depleted NEO1 in excitatory (NeuroD6-Cre+) or inhibitory (parvalbumin+) neurons, but present in mice with specific hippocampal astrocytic Neo1 KO. Additionally, neurons in hippocampal dentate gyrus, a vulnerable region in epilepsy, in mice with astrocyte-specific Neo1 KO show reductions in inhibitory synaptic vesicles and the frequency of miniature inhibitory postsynaptic current(mIPSC), but increase of the duration of miniature excitatory postsynaptic current and tonic NMDA receptor currents, suggesting impairments in both GABAergic transmission and extracellular glutamate clearance. Further proteomic and cell biological analyses of cell-surface proteins identified GLAST, a glutamate-aspartate transporter that is marked reduced in Neo1 KO astrocytes and the hippocampus. NEO1 interacts with GLAST and promotes GLAST surface distribution in astrocytes. Expressing NEO1 or GLAST in Neo1 KO astrocytes in the hippocampus abolishes the epileptic phenotype. Taken together, these results uncover an unrecognized pathway of NEO1-GLAST in hippocampal GFAP+ astrocytes, which is critical for GLAST surface distribution and function, and GABAergic transmission, unveiling NEO1 as a valuable therapeutic target to protect the brain from epilepsy.
Project description:Memory consolidation is the process by which acquired information is converted to something concrete to be retrieved later. Here we examined a potential role for brain-derived neurotrophic factor (BDNF) in mediating the enhanced memory consolidation induced by the GABA(A) receptor antagonist, bicuculline methiodide. With the administration of an acquisition trial in naïve mice using a passive avoidance task, mature BDNF (mBDNF) levels were temporally changed in the hippocampal CA1 region, and the lowest levels were observed 9?h after the acquisition trial. In the passive avoidance task, bicuculline methiodide administration within 1?h of training but not after 3?h significantly increased latency time in the retention trial 24?h after the acquisition trial. Concomitantly, 1?h post-training administration of bicuculline methiodide, which enhanced memory consolidation, significantly increased mBDNF levels 9?h after training compared to those of the vehicle-treated control group. In addition, exogenous human recombinant BDNF (hrBDNF) administration 9?h after training into the hippocampal CA1 region facilitated memory consolidation confirming that the increase in mBDNF at around 9?h after training plays a key role in the enhancement of memory consolidation. Moreover, the increases in latency time and immediate early gene expressions by bicuculline methiodide or hrBDNF were significantly blocked by anisomycin, a protein synthesis inhibitor, K252a, a tyrosine receptor kinase (Trk) inhibitor, or anti-TrkB IgG. These findings suggest that the increase in the level of mBDNF and its function during a restricted time window after training are required for the enhancement of memory consolidation by GABA(A) receptor blockade.
Project description:Pharmacological suppression of γ-aminobutyric acid (GABA) transaminase (GABA-T), the sole GABA-degrading enzyme and a potential therapeutic target for treating brain disorders such as epilepsy, increases not only phasic inhibition but also tonic inhibition. However, the specific cellular source, neuromodulatory effects and potential therapeutic benefits of this enhanced tonic inhibition remain unexplored due to the lack of cell-type-specific gene manipulation studies. Here we report that the increase in tonic GABA currents observed after GABA-T suppression is predominantly due to increased tonic GABA release from astrocytes rather than action-potential-dependent synaptic GABA spillover. General GABA-T knockdown (KD) by a short hairpin RNA considerably increased tonic GABA currents in dentate granule cells, thereby enhancing tonic inhibition. An astrocyte-specific rescue of GABA-T following general GABA-T KD normalized the elevated tonic GABA currents to near control levels. Tetrodotoxin-insensitive tonic GABA currents were significantly increased after general GABA-T KD, whereas tetrodotoxin-sensitive tonic GABA currents showed no significant increase, suggesting that this enhanced tonic inhibition is primarily action-potential independent. General GABA-T KD reduced the spike probability of granule cells and impaired dorsal hippocampus-dependent spatial memory, which were fully reversed by astrocyte-specific GABA-T rescue. These findings suggest that suppressing astrocytic GABA-T may be sufficient to influence the excitatory/inhibitory balance in the brain and associated behaviors. Our study implies that the therapeutic benefits of pharmacological GABA-T suppression may be largely attributed to the modulation of astrocytic GABA-T and its impact on tonic GABA release from astrocytes. Here, we report distinct effects of GABA-T suppression depending on cell type; suppressing GABA-T in astrocytes enhances tonic inhibition, while its suppression in GABAergic neurons augments phasic inhibition. Our findings demonstrate that targeted suppression of astrocytic GABA-T not only enhances tonic GABA release from astrocytes but also significantly influences the excitation/inhibition balance in the brain, with consequential effects on behavior. This suggests that astrocytic GABA-T modulation holds promising potential for developing novel therapeutic strategies aimed at treating cognitive and neurological disorders through the regulation of astrocytic GABA metabolism. GAD glutamate decarboxylase, MAO-B monoamine oxidase B, BEST1 bestrophin 1, GABA-T GABA transaminase, GAT GABA transporter, DG dentate gyrus, SSA succinic semialdehyde.
Project description:The val(66)met polymorphism on the BDNF gene has been reported to explain individual differences in hippocampal volume and memory-related activity. These findings, however, have not been replicated consistently and no studies to date controlled for the potentially confounding impact of early life stress, such as childhood abuse, and psychiatric status. Using structural and functional MRI, we therefore investigated in 126 depressed and/or anxious patients and 31 healthy control subjects the effects of val(66)met on hippocampal volume and encoding activity of neutral, positive and negative words, while taking into account childhood abuse and psychiatric status. Our results show slightly lower hippocampal volumes in carriers of a met allele (n=54) relative to val/val homozygotes (n=103) (P=0.02, effect size (Cohen's d)=0.37), which appeared to be independent of childhood abuse and psychiatric status. For hippocampal encoding activity, we found a val(66)met-word valence interaction (P=0.02) such that carriers of a met allele showed increased levels of activation in response to negative words relative to activation in the neutral word condition and relative to val/val homozygotes. This, however, was only evident in the absence of childhood abuse, as abused val/val homozygotes showed hippocampal encoding activity for negative words that was comparable to that of carriers of a met allele. Neither psychiatric status nor memory accuracy did account for these associations. In conclusion, BDNF val(66)met has a significant impact on hippocampal volume independently of childhood abuse and psychiatric status. Furthermore, early adverse experiences such as childhood abuse account for individual differences in hippocampal encoding activity of negative stimuli but this effect manifests differently as a function of val(66)met.
Project description:It is well recognized that astrocytes can produce factors known to affect the myelination process. One such factor, brain-derived neurotrophic factor (BDNF), can enhance the differentiation of oligodendrocyte lineage cells following a demyelinating lesion. Our previous work indicated that enhancing astrocyte-derived BDNF via injection of a general agonist of Group I/II metabotropic glutamate receptors (mGluRs) into the lesion increased myelin proteins in the cuprizone model of demyelination after 4 hr. To determine if this observation has potential therapeutic significance, we now use a more specific mGluR agonist, 2-chloro-5-hydroxyphenylglycine (CHPG), which binds to mGluR5, to examine effects on myelination through the clinically relevant approach of a peripheral injection. In initial studies, intraperitoneal injection of CHPG resulted in an increase in myelin proteins within the lesioned corpus callosum. These effects were blocked when either BDNF or the CHPG receptor, mGluR5, was deleted from glial fibrillary acidic protein (GFAP)+ astrocytes or when the BDNF receptor, tropomyosin receptor kinase B (TrkB), was deleted from proteolipid protein (PLP)+ oligodendrocytes. Moreover, injection of CHPG over 2 weeks not only elevated BDNF and myelin proteins, but also enhanced myelination and reversed behavioral deficits. Interestingly, effects on myelin and myelin proteins were not seen in the control animals, indicating that a lesion is critical in eliciting effects. Taken together, the data suggest that the mGluR agonist CHPG may be a potential therapeutic strategy for treating demyelinating diseases and that it works by enhancing the release of BDNF from astrocytes.
Project description:The role of glutamate N-methyl-D-aspartate receptor (NMDAR) hypofunction has been extensively studied in schizophrenia; however, less is known about its role in anxiety disorders. Recently, it was demonstrated that astrocytic GLT-1 blockade leads to an anxiety-like phenotype. Although astrocytes are capable of modulating NMDAR activity through glutamate uptake transporters, the relationship between astrocytic glutamate uptake and the development of an anxiety phenotype remains poorly explored. Here, we aimed to investigative whether long-term antagonism of NMDAR impacts anxiety-related behaviors and astrocytic glutamate uptake. Memantine, an NMDAR antagonist, was administered daily for 24 days to healthy adult CF-1 mice by oral gavage at doses of 5, 10, or 20 mg/kg. The mice were submitted to a sequential battery of behavioral tests (open field, light-dark box and elevated plus-maze tests). We then evaluated glutamate uptake activity and the immunocontents of glutamate transporters in the frontoparietal cortex and hippocampus. Our results demonstrated that long-term administration of memantine induces anxiety-like behavior in mice in the light-dark box and elevated plus-maze paradigms. Additionally, the administration of memantine decreased glutamate uptake activity in both the frontoparietal cortex and hippocampus without altering the immunocontent of either GLT-1 or GLAST. Remarkably, the memantine-induced reduction in glutamate uptake was correlated with enhancement of an anxiety-like phenotype. In conclusion, long-term NMDAR antagonism with memantine induces anxiety-like behavior that is associated with reduced glutamate uptake activity but that is not dependent on GLT-1 or GLAST protein expression. Our study suggests that NMDAR and glutamate uptake hypofunction may contribute to the development of conditions that fall within the category of anxiety disorders.
Project description:Glutamatergic transmission prompts K+ efflux through postsynaptic NMDA receptors. The ensuing hotspot of extracellular K+ elevation depolarizes presynaptic terminal, boosting glutamate release, but whether this also affects glutamate uptake in local astroglia has remained an intriguing question. Here, we find that the pharmacological blockade, or conditional knockout, of postsynaptic NMDA receptors suppresses use-dependent increase in the amplitude and duration of the astrocytic glutamate transporter current (IGluT ), whereas blocking astrocytic K+ channels prevents the duration increase only. Glutamate spot-uncaging reveals that astrocyte depolarization, rather than extracellular K+ rises per se, is required to reduce the amplitude and duration of IGluT . Biophysical simulations confirm that local transient elevations of extracellular K+ can inhibit local glutamate uptake in fine astrocytic processes. Optical glutamate sensor imaging and a two-pathway test relate postsynaptic K+ efflux to enhanced extrasynaptic glutamate signaling. Thus, repetitive glutamatergic transmission triggers a feedback loop in which postsynaptic K+ efflux can transiently facilitate presynaptic release while reducing local glutamate uptake.
Project description:During early postnatal development, sensory regions of the brain undergo periods of heightened plasticity which sculpt neural networks and lay the foundation for adult sensory perception. Such critical periods were also postulated for learning and memory but remain elusive and poorly understood. Here, we present evidence that the activity-regulated and memory-linked gene Arc/Arg3.1 is transiently up-regulated in the hippocampus during the first postnatal month. Conditional removal of Arc/Arg3.1 during this period permanently alters hippocampal oscillations and diminishes spatial learning capacity throughout adulthood. In contrast, post developmental removal of Arc/Arg3.1 leaves learning and network activity patterns intact. Long-term memory storage continues to rely on Arc/Arg3.1 expression throughout life. These results demonstrate that Arc/Arg3.1 mediates a critical period for spatial learning, during which Arc/Arg3.1 fosters maturation of hippocampal network activity necessary for future learning and memory storage.
Project description:Adult neurogenesis in the hippocampus is a remarkable phenomenon involved in various aspects of learning and memory as well as disease pathophysiology. Brain-derived neurotrophic factor (BDNF) represents a major player in the regulation of this unique form of neuroplasticity, yet the mechanisms underlying its pro-neurogenic actions remain unclear. Here, we examined the effects associated with brief (25 min), unilateral infusion of BDNF in the rat dentate gyrus. Acute BDNF infusion induced long-term potentiation (LTP) of medial perforant path-evoked synaptic transmission and, concomitantly, enhanced hippocampal neurogenesis bilaterally, reflected by increased dentate gyrus BrdU + cell numbers. Importantly, inhibition of activity-regulated cytoskeleton-associated protein (Arc/Arg3.1) translation through local, unilateral infusion of anti-sense oligodeoxynucleotides (ArcAS) prior to BDNF infusion blocked both BDNF-LTP induction and the associated pro-neurogenic effects. Notably, basal rates of proliferation and newborn cell survival were unaltered in homozygous Arc/Arg3.1 knockout mice. Taken together these findings link the pro-neurogenic effects of acute BDNF infusion to induction of Arc/Arg3.1-dependent LTP in the adult rodent dentate gyrus.
Project description:The dentate gyrus of the hippocampus is one of the few regions of the mammalian brain where new neurons are generated throughout adulthood. This adult neurogenesis has been proposed as a novel mechanism that mediates spatial memory. However, data showing a causal relationship between neurogenesis and spatial memory are controversial. Here, we developed an inducible transgenic strategy allowing specific ablation of adult-born hippocampal neurons. This resulted in an impairment of spatial relational memory, which supports a capacity for flexible, inferential memory expression. In contrast, less complex forms of spatial knowledge were unaltered. These findings demonstrate that adult-born neurons are necessary for complex forms of hippocampus-mediated learning.