Mercury is a well-known neurotoxin implicated in a wide range of neurological or psychiatric disorders including autism spectrum disorders, Alzheimer's disease, Parkinson's disease, epilepsy, ...depression, mood disorders and tremor. Mercury-induced neuronal degeneration is thought to invoke glutamate-mediated excitotoxicity, however, the underlying mechanisms remain poorly understood. Here, we examine the effects of various mercury concentrations (including pathological levels present in human plasma or cerebrospinal fluid) on cultured, rat cortical neurons.
We found that inorganic mercuric chloride (HgCl₂--at 0.025 to 25 μM) not only caused neuronal degeneration but also perturbed neuronal excitability. Whole-cell patch-clamp recordings of pyramidal neurons revealed that HgCl₂ not only enhanced the amplitude and frequency of synaptic, inward currents, but also increased spontaneous synaptic potentials followed by sustained membrane depolarization. HgCl₂ also triggered sustained, 2-5 fold rises in intracellular calcium concentration (Ca²⁺i). The observed increases in neuronal activity and Ca²⁺i were substantially reduced by the application of MK 801, a non-competitive antagonist of N-Methyl-D-Aspartate (NMDA) receptors. Importantly, our study further shows that a pre incubation or co-application of MK 801 prevents HgCl₂-induced reduction of cell viability and a disruption of β-tubulin.
Collectively, our data show that HgCl₂-induced toxic effects on central neurons are triggered by an over-activation of NMDA receptors, leading to cytoskeleton instability.
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The mechanism of postsynaptic neurotransmitter receptor clustering has been best described at the neuromuscular junction (NMJ), where packets of nicotinic acetylcholine receptors (nAChRs) in muscle ...fibre are redistributed to the synaptic site upon motor neuron innervation. This process of receptor localisation is facilitated by agrin signalling, and the stability of the resulting nAChR clusters depends upon the activation of Src family kinases (SFKs). In contrast to the NMJ however, the cellular signalling mechanisms orchestrating the clustering of nAChRs in the central nervous system (CNS) remain poorly defined. Furthermore, our understanding of the role of SFKs in the CNS is also limited. Here we provide evidence that SFK activation is required for synapse formation between pairs of identified neurons isolated from the CNS of Lymnaea stagnalis. Furthermore, we suggest that SFKs are involved in the functional redistribution of nAChRs to the synaptic contact sites in isolated axons. To the best of our knowledge, this study is the first to demonstrate a role for SFKs in the clustering of nAChRs in central neurons, suggesting that the mechanisms of receptor clustering between the peripheral and central nervous system are likely conserved.
As the saying goes: history repeats itself but for those who fail to learn lessons from it, they are often the worst off when the boomerang returns to strike them at the back. The 1918 influenza ...pandemic, caused by H1N1 virus was perhaps the worst that our humanity had yet encountered. It’s origin debatable (avian or otherwise), it infected over 500 Million people across the globe within a year, and wiped out almost 50 Million humans (40% world population) from the face of the earth.
is a unique model system for studying complex behaviors in animals. It has a large and centralized nervous system made up of lobes that are involved in controlling various sophisticated behaviors. As ...such, it may be considered as a model organism for untangling the neuronal mechanisms underlying behaviors-including learning and memory. However, despite considerable efforts, Octopus lags behind its other counterparts vis-à-vis its utility in deciphering the cellular, molecular and synaptic mechanisms underlying various behaviors. This study represents a novel approach designed to establish a neuronal cell culture protocol that makes this species amenable to further exploitation as a model system. Here we developed a protocol that enables dissociation of neurons from two specific Octopus' brain regions, the vertical-superior frontal system and the optic lobes, which are involved in memory, learning, sensory integration and adult neurogenesis. In particular, cells dissociated with enzyme papain and cultured on Poly-D-Lysine-coated dishes with L15-medium and fetal bovine serum yielded high neuronal survival, axon growth, and re-growth after injury. This model was also explored to define optimal culture conditions and to demonstrate the regenerative capabilities of adult Octopus neurons after axotomy. This study thus further underscores the importance of Octopus neurons as a model system for deciphering fundamental molecular and cellular mechanism of complex brain function and underlying behaviors.
Cholinergic neuronal networks in the hippocampus play a key role in the regulation of learning and memory in mammals. Perturbations of these networks, in turn, underlie neurodegenerative diseases. ...However, the mechanisms remain largely undefined. We have recently demonstrated that an in vitro
gene deletion perturbs nicotinic cholinergic plasticity at the hippocampal glutamatergic synapses. Furthermore,
neuronal conditional knockout in freely behaving animals has also been shown to result in learning and memory deficits, though the evidence remains equivocal. In this study, using an AVV viral vector transcription approach, we provide direct evidence that
gene deletion in the CA1 region of the hippocampus indeed leads to contextual fear conditioning deficits in conditional knockout animals. This loss of function was, however, recovered when the same animals were re-injected to overexpress
. This study provides the first direct evidence for the sufficiency and necessity of
in fear conditioning, and further endorses the role of menin in the regulation of cholinergic synaptic machinery in the hippocampus. These data underscore the importance of further exploring and revisiting the cholinergic hypothesis that underlies neurodegenerative diseases that affect learning and memory.
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Neurotrophic factors (NTFs) support neuronal survival, differentiation, and even synaptic plasticity both during development and throughout the life of an organism. However, their precise roles in ...central synapse formation remain unknown. Previously, we demonstrated that excitatory synapse formation in Lymnaea stagnalis requires a source of extrinsic NTFs and receptor tyrosine kinase (RTK) activation. Here we show that NTFs such as Lymnaea epidermal growth factor (L-EGF) act through RTKs to trigger a specific subset of intracellular signalling events in the postsynaptic neuron, which lead to the activation of the tumor suppressor menin, encoded by Lymnaea MEN1 (L-MEN1) and the expression of excitatory nicotinic acetylcholine receptors (nAChRs). We provide direct evidence that the activation of the MAPK/ERK cascade is required for the expression of nAChRs, and subsequent synapse formation between pairs of neurons in vitro. Furthermore, we show that L-menin activation is sufficient for the expression of postsynaptic excitatory nAChRs and subsequent synapse formation in media devoid of NTFs. By extending our findings in situ, we reveal the necessity of EGFRs in mediating synapse formation between a single transplanted neuron and its intact presynaptic partner. Moreover, deficits in excitatory synapse formation following EGFR knock-down can be rescued by injecting synthetic L-MEN1 mRNA in the intact central nervous system. Taken together, this study provides the first direct evidence that NTFs functioning via RTKs activate the MEN1 gene, which appears sufficient to regulate synapse formation between central neurons. Our study also offers a novel developmental role for menin beyond tumour suppression in adult humans.
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Our inability to accurately monitor individual neurons and their synaptic activity precludes fundamental understanding of brain function under normal and various pathological conditions. However, ...recent breakthroughs in micro- and nano-scale fabrication processes have advanced the development of neuro-electronic hybrid technology. Among such devices are three-dimensional and planar electrodes, offering the advantages of either high fidelity or longer-term recordings respectively. Here, we present the next generation of planar microelectrode arrays with "nano-edges" that enable long-term (≥1 month) and high fidelity recordings at a resolution 15 times higher than traditional planar electrodes. This novel technology enables better understanding of brain function and offers a tremendous opportunity towards the development of future bionic hybrids and drug discovery devices.
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In the central nervous system (CNS), cholinergic transmission induces synaptic plasticity that is required for learning and memory. However, our understanding of the development and maintenance of ...cholinergic circuits is limited, as the factors regulating the expression and clustering of neuronal nicotinic acetylcholine receptors (nAChRs) remain poorly defined. Recent studies from our group have implicated calpain-dependent proteolytic fragments of menin, the product of the MEN1 tumor suppressor gene, in coordinating the transcription and synaptic clustering of nAChRs in invertebrate central neurons. Here, we sought to determine whether an analogous cholinergic mechanism underlies menin's synaptogenic function in the vertebrate CNS. Our data from mouse primary hippocampal cultures demonstrate that menin and its calpain-dependent C-terminal fragment (C-menin) regulate the subunit-specific transcription and synaptic clustering of neuronal nAChRs, respectively. MEN1 knockdown decreased nAChR α5 subunit expression, the clustering of α7 subunit-containing nAChRs at glutamatergic presynaptic terminals, and nicotine-induced presynaptic facilitation. Moreover, the number and function of glutamatergic synapses was unaffected by MEN1 knockdown, indicating that the synaptogenic actions of menin are specific to cholinergic regulation. Taken together, our results suggest that the influence of menin on synapse formation and synaptic plasticity occur via modulation of nAChR channel subunit composition and functional clustering.
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All functions of the nervous system are contingent upon the precise organization of neuronal connections that are initially patterned during development, and then continually modified throughout ...life. Determining the mechanisms that specify the formation and functional modulation of synaptic circuitry are critical to advancing both our fundamental understanding of the nervous system as well as the various neurodevelopmental, neurological, neuropsychiatric, and neurodegenerative disorders that are met in clinical practice when these processes go awry. Defining the cellular and molecular mechanisms underlying nervous system development, function, and pathology has proven challenging, due mainly to the complexity of the vertebrate brain. Simple model system approaches with invertebrate preparations, on the other hand, have played pivotal roles in elucidating the fundamental mechanisms underlying the formation and plasticity of individual synapses, and the contributions of individual neurons and their synaptic connections that underlie a variety of behaviors, and learning and memory. In this Review, we discuss the experimental utility of the invertebrate mollusc Lymnaea stagnalis, with a particular emphasis on in vitro cell culture, semi-intact and in vivo preparations, which enable molecular and electrophysiological identification of the cellular and molecular mechanisms governing the formation, plasticity, and specificity of individual synapses at a single-neuron or single-synapse resolution.
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The perturbation of nicotinic cholinergic receptors is thought to underlie many neurodegenerative and neuropsychiatric disorders, such as Alzheimer's and schizophrenia. We previously identified that ...the tumor suppressor gene,
, regulates both the expression and synaptic targeting of α7 nAChRs in the mouse hippocampal neurons in vitro. Here we sought to determine whether the α7 nAChRs gene expression reciprocally regulates the expression of menin, the protein encoded by the
gene, and if this interplay impacts learning and memory. We demonstrate here that α7 nAChRs knockdown (KD) both in in vitro and in vivo, initially upregulated and then subsequently downregulated menin expression. Exogenous expression of menin using an AAV transduction approach rescued α7 nAChRs KD mediated functional and behavioral deficits specifically in hippocampal (CA1) neurons. These effects involved the modulation of the α7 nAChR subunit expression and functional clustering at the synaptic sites. Our data thus demonstrates a novel and important interplay between the
gene and the α7 nAChRs in regulating hippocampal-dependent learning and memory.
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