Insertion Of Receptors Research Articles

Insertion of Calcium-Permeable AMPA Receptors during Epileptiform Activity In Vitro Modulates Excitability of Principal Neurons in the Rat Entorhinal Cortex.

Epileptic activity leads to rapid insertion of calcium-permeable α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (CP-AMPARs) into the synapses of cortical and hippocampal glutamatergic neurons, which generally do not express them. The physiological significance of this process is not yet fully understood; however, it is usually assumed to be a pathological process that augments epileptic activity. Using whole-cell patch-clamp recordings in rat entorhinal cortex slices, we demonstrate that the timing of epileptiform discharges, induced by 4-aminopyridine and gabazine, is determined by the shunting effect of Ca2+-dependent slow conductance, mediated predominantly by K+-channels. The blockade of CP-AMPARs by IEM-1460 eliminates this extra conductance and consequently increases the rate of discharge generation. The blockade of NMDARs reduced the additional conductance to a lesser extent than the blockade of CP-AMPARs, indicating that CP-AMPARs are a more significant source of intracellular Ca2+. The study’s main findings were implemented in a mathematical model, which reproduces the shunting effect of activity-dependent conductance on the generation of discharges. The obtained results suggest that the expression of CP-AMPARs in principal neurons reduces the discharge generation rate and may be considered as a protective mechanism.

β-Amyloid disruption of LTP/LTD balance is mediated by AKAP150-anchored PKA and Calcineurin regulation of Ca2+-permeable AMPA receptors.

SUMMARYRegulated insertion and removal of postsynaptic AMPA glutamate receptors (AMPARs) mediates hippocampal long-term potentiation (LTP) and long-term depression (LTD) synaptic plasticity underlying learning and memory. In Alzheimer’s disease β-amyloid (Aβ) oligomers may impair learning and memory by altering AMPAR trafficking and LTP/LTD balance. Importantly, Ca2+-permeable AMPARs (CP-AMPARs) assembled from GluA1 subunits are excluded from hippocampal synapses basally but can be recruited rapidly during LTP and LTD to modify synaptic strength and signaling. By employing mouse knockin mutations that disrupt anchoring of the kinase PKA or phosphatase Calcineurin (CaN) to the postsynaptic scaffold protein AKAP150, we find that local AKAP-PKA signaling is required for CP-AMPAR recruitment, which can facilitate LTP but also, paradoxically, prime synapses for Aβ impairment of LTP mediated by local AKAP-CaN LTD signaling that promotes subsequent CP-AMPAR removal. These findings highlight the importance of PKA/CaN signaling balance and CP-AMPARs in normal plasticity and aberrant plasticity linked to disease.

The processes underlying chronic migraine pathophysiology and its treatment with botulinum toxin type A

AbstractObjectiveTo comprehensively review both the models of migraine pathophysiology and transformation to chronic migraine, and align these with the known effects of botulinum toxin type A (BTX‐A).BackgroundMigraine is a relatively common, disabling, and debilitating condition that is notoriously difficult to manage by pain specialists. A comprehensive review of the mechanisms of its pathophysiology, chronification, and treatment with BTX‐A is lacking.MethodsThis narrative literature review is formed following an intensive search of scientific literature on PubMed, CENTRAL, and EMBASE databases using relevant key search terms.ResultsMigraine can be seen as a threshold disorder with multiple mechanisms: hyperexcitability of trigeminal afferents, impaired descending inhibition, and central sensitization. The mechanism of transformation to chronic migraine, where headache symptoms occur on at least 15 days per month, remains unclear, but key risk factors include sex (female), headache frequency, psychiatric co‐morbidities, obesity, and low socioeconomic status. Botulinum toxin A is an exotoxin produced by the gram‐positive rod bacterium Clostridium botulinum. Following the agent's initial use in aesthetic medicine, it is now an FDA‐approved treatment for chronic migraine. The toxin has multiple modes of action in chronic migraine, including neurotransmitter exocytosis blockade, inhibition of membrane receptor insertion, and anti‐inflammatory processes.ConclusionMigraine is likely caused by intrinsic hyperexcitability of the trigeminovascular nociceptive system, disrupted descending inhibitory pain networks, and central sensitization. Pathophysiology of chronification is poorly understood. BTX‐A acts primarily by attenuating neuropeptidergic signaling, inhibiting membrane receptor insertion, and downregulating expression of receptors implicated in cortical spreading depression.

Calcium-Permeable AMPA Receptors Mediate Timing-Dependent LTP Elicited by Low Repeat Coincident Pre- and Postsynaptic Activity at Schaffer Collateral-CA1 Synapses.

High-frequency stimulation induced long-term potentiation (LTP) and low-frequency stimulation induced LTD are considered as cellular models of memory formation. Interestingly, spike timing-dependent plasticity (STDP) can induce equally robust timing-dependent LTP (t-LTP) and t-LTD in response to low frequency repeats of coincident action potential (AP) firing in presynaptic and postsynaptic cells. Commonly, STDP paradigms relying on 25-100 repeats of coincident AP firing are used to elicit t-LTP or t-LTD, but the minimum number of repeats required for successful STDP is barely explored. However, systematic investigation of physiologically relevant low repeat STDP paradigms is of utmost importance to explain learning mechanisms in vivo. Here, we examined low repeat STDP at Schaffer collateral-CA1 synapses by pairing one presynaptic AP with either one postsynaptic AP (1:1t-LTP), or a burst of 4 APs (1:4t-LTP) and found 3-6 repeats to be sufficient to elicit t-LTP. 6× 1:1t-LTP required postsynaptic Ca2+ influx via NMDARs and L-type VGCCs and was mediated by increased presynaptic glutamate release. In contrast, 1:4t-LTP depended on postsynaptic metabotropic GluRs and ryanodine receptor signaling and was mediated by postsynaptic insertion of AMPA receptors. Unexpectedly, both 6× t-LTP variants were strictly dependent on activation of postsynaptic Ca2+-permeable AMPARs but were differentially regulated by dopamine receptor signaling. Our data show that synaptic changes induced by only 3-6 repeats of mild STDP stimulation occurring in ≤10s can take place on time scales observed also during single trial learning.

Biomimetic Recognition-Based Bioorthogonal Host–Guest Pairs for Cell Targeting and Tissue Imaging in Living Animals

Biomimetic Recognition-Based Bioorthogonal Host–Guest Pairs for Cell Targeting and Tissue Imaging in Living Animals

Adaptive Balance in Posterior Cerebellum.

Vestibular and optokinetic space is represented in three-dimensions in vermal lobules IX-X (uvula, nodulus) and hemisphere lobule X (flocculus) of the cerebellum. Vermal lobules IX-X encodes gravity and head movement using the utricular otolith and the two vertical semicircular canals. Hemispheric lobule X encodes self-motion using optokinetic feedback about the three axes of the semicircular canals. Vestibular and visual adaptation of this circuitry is needed to maintain balance during perturbations of self-induced motion. Vestibular and optokinetic (self-motion detection) stimulation is encoded by cerebellar climbing and mossy fibers. These two afferent pathways excite the discharge of Purkinje cells directly. Climbing fibers preferentially decrease the discharge of Purkinje cells by exciting stellate cell inhibitory interneurons. We describe instances adaptive balance at a behavioral level in which prolonged vestibular or optokinetic stimulation evokes reflexive eye movements that persist when the stimulation that initially evoked them stops. Adaptation to prolonged optokinetic stimulation also can be detected at cellular and subcellular levels. The transcription and expression of a neuropeptide, corticotropin releasing factor (CRF), is influenced by optokinetically-evoked olivary discharge and may contribute to optokinetic adaptation. The transcription and expression of microRNAs in floccular Purkinje cells evoked by long-term optokinetic stimulation may provide one of the subcellular mechanisms by which the membrane insertion of the GABAA receptors is regulated. The neurosteroids, estradiol (E2) and dihydrotestosterone (DHT), influence adaptation of vestibular nuclear neurons to electrically-induced potentiation and depression. In each section of this review, we discuss how adaptive changes in the vestibular and optokinetic subsystems of lobule X, inferior olivary nuclei and vestibular nuclei may contribute to the control of balance.

The amino-terminal domain of GluA1 mediates LTP maintenance via interaction with neuroplastin-65

Long-term potentiation (LTP) has long been considered as an important cellular mechanism for learning and memory. LTP expression involves NMDA receptor-dependent synaptic insertion of AMPA receptors (AMPARs). However, how AMPARs are recruited and anchored at the postsynaptic membrane during LTP remains largely unknown. In this study, using CRISPR/Cas9 to delete the endogenous AMPARs and replace them with the mutant forms in single neurons, we have found that the amino-terminal domain (ATD) of GluA1 is required for LTP maintenance. Moreover, we show that GluA1 ATD directly interacts with the cell adhesion molecule neuroplastin-65 (Np65). Neurons lacking Np65 exhibit severely impaired LTP maintenance, and Np65 deletion prevents GluA1 from rescuing LTP in AMPARs-deleted neurons. Thus, our study reveals an essential role for GluA1/Np65 binding in anchoring AMPARs at the postsynaptic membrane during LTP.

The WD40-Repeat Protein WDR-20 and the Deubiquitinating Enzyme USP-46 Promote Cell Surface Levels of Glutamate Receptors.

Reversible modification of AMPA receptors (AMPARs) with ubiquitin regulates receptor levels at synapses and controls synaptic strength. The conserved deubiquitinating enzyme (DUB) ubiquitin-specific protease-46 (USP-46) removes ubiquitin from AMPARs and protects them from degradation in both Caenorhabditis elegans and mammals. Although DUBs are critical for diverse physiological processes, the mechanisms that regulate DUBs, especially in the nervous system, are not well understood. We and others previously showed that the WD40-repeat proteins WDR-48 and WDR-20 bind to and stimulate the catalytic activity of USP-46. Here, we identify an activity-dependent mechanism that regulates WDR-20 expression and show that WDR-20 works together with USP-46 and WDR-48 to promote surface levels of the C. elegans AMPAR GLR-1. usp-46, wdr-48, and wdr-20 loss-of-function mutants exhibit reduced levels of GLR-1 at the neuronal surface and corresponding defects in GLR-1-mediated behavior. Increased expression of WDR-20, but not WDR-48, is sufficient to increase GLR-1 surface levels in an usp-46-dependent manner. Loss of usp-46, wdr-48, and wdr-20 function reduces the rate of local GLR-1 insertion in neurites, whereas overexpression of wdr-20 is sufficient to increase the rate of GLR-1 insertion. Genetic manipulations that chronically reduce or increase glutamate signaling result in reciprocal alterations in wdr-20 transcription and homeostatic compensatory changes in surface GLR-1 levels that are dependent on wdr-20 This study identifies wdr-20 as a novel activity-regulated gene that couples chronic changes in synaptic activity with increased local insertion and surface levels of GLR-1 via the DUB USP-46.SIGNIFICANCE STATEMENT Deubiquitinating enzymes (DUBs) are critical regulators of synapse development and function; however, the regulatory mechanisms that control their various physiological functions are not well understood. This study identifies a novel role for the DUB ubiquitin-specific protease-46 (USP-46) and its associated regulatory protein WD40-repeat protein-20 (WDR-20) in regulating local insertion of glutamate receptors into the neuronal cell surface. This work also identifies WDR-20 as an activity-regulated gene that couples chronic changes in synaptic activity with homeostatic compensatory increases in surface levels of GLR-1 via USP-46. Given that 35% of USP family DUBs associate with WDR proteins, understanding the mechanisms by which WDR proteins regulate USP-46 could have implications for a large number of DUBs in other cell types.

Inhibitors of synaptic vesicle exocytosis reduce surface expression of postsynaptic glutamate receptors

ABSTRACT Bafilomycin A1, a vacuolar H+-ATPase inhibitor, and botulinum toxin B and tetanus toxin, both vesicle fusion inhibitors, are widely known exocytosis blockers that have been used to inhibit the presynaptic release of neurotransmitters. However, protein trafficking mechanisms, such as the insertion of postsynaptic receptors and astrocytic glutamate-releasing channels into the plasma membrane, also require exocytosis. In our previous study, exocytosis inhibitors reduced the surface expression of astrocytic glutamate-releasing channels. Here, we further investigated whether exocytosis inhibitors influence the surface expression of postsynaptic receptors. Using pH-sensitive superecliptic pHluorin (SEP)-tagged postsynaptic glutamate receptors, including GluA1, GluA2, GluN1, and GluN2A, we found that bafilomycin A1, botulinum toxin B, and/or tetanus toxin reduce the SEP fluorescence of SEP-GluA1, SEP-GluA2, SEP-GluN1, and SEP-GluN2A. These findings indicate that presynaptic vesicle exocytosis inhibitors also affect the postsynaptic trafficking machinery for surface expression. Finally, this study provides profound insights assembling presynaptic, postsynaptic and astrocytic viewpoints into the interpretation of the data obtained using these synaptic vesicle exocytosis inhibitors.

Enhancing site-specific DNA integration by a Cas9 nuclease fused with a DNA donor-binding domain.

The CRISPR/Cas system is widely used for genome editing. However, robust and targeted insertion of a DNA segment remains a challenge. Here, we present a fusion nuclease (Cas9-N57) to enhance site-specific DNA integration via a fused DNA binding domain of Sleeping Beauty transposase to tether the DNA segment to the Cas9/sgRNA complex. The insertion was unidirectional and specific, and DNA fragments up to 12 kb in length were successfully integrated. As a test of the system, Cas9-N57 mediated the insertion of a CD19-specific chimeric antigen receptor (CD19-CAR) cassette into the AAVS1 locus in human T cells, and induced intrahepatic cholangiocarcinoma in mice by simultaneously mediating the insertion of oncogenic KrasG12D into the Rosa26 locus and disrupting Trp53 and Pten. Moreover, the nuclease-N57 fusion proteins based on AsCpf1 (AsCas12a) and CjCas9 exhibited similar activity. These findings demonstrate that CRISPR-associated nuclease-N57 protein fusion is a powerful tool for targeted DNA insertion and holds great potential for gene therapy applications.

Mechanism of Action of OnabotulinumtoxinA in Chronic Migraine: A Narrative Review.

ObjectiveTo review the literature on the mechanism of action of onabotulinumtoxinA in chronic migraine.BackgroundOnabotulinumtoxinA is a chronic migraine preventive treatment that significantly reduces headache frequency. The traditional mechanism described for onabotulinumtoxinA – reducing muscle contractions – is insufficient to explain its efficacy in migraine, which is primarily a sensory neurological disease.MethodsA narrative literature review on the mechanism of action of onabotulinumtoxinA in chronic migraine.ResultsFollowing injection into tissues, onabotulinumtoxinA inhibits soluble N‐ethylmaleimide‐sensitive fusion attachment protein receptor (SNARE)‐mediated vesicle trafficking by cleaving one of its essential proteins, soluble N‐ethylmaleimide‐sensitive fusion attachment protein (SNAP‐25), which occurs in both motor and sensory nerves. OnabotulinumtoxinA inhibits regulated exocytosis of motor and sensory neurochemicals and proteins, as well as membrane insertion of peripheral receptors that convey pain from the periphery to the brain, because both processes are SNARE dependent. OnabotulinumtoxinA can decrease exocytosis of pro‐inflammatory and excitatory neurotransmitters and neuropeptides such as substance P, calcitonin gene‐related peptide, and glutamate from primary afferent fibers that transmit nociceptive pain and participate in the development of peripheral and central sensitization. OnabotulinumtoxinA also decreases the insertion of pain‐sensitive ion channels such as transient receptor potential cation channel subfamily V member 1 (TRPV1) into the membranes of nociceptive neurons; this is likely enhanced in the sensitized neuron. For chronic migraine prevention, onabotulinumtoxinA is injected into 31‐39 sites in 7 muscles of the head and neck. Sensory nerve endings of neurons whose cell bodies are located in trigeminal and cervical ganglia are distributed throughout the injected muscles, and are overactive in people with migraine. Through inhibition of these sensory nerve endings, onabotulinumtoxinA reduces the number of pain signals that reach the brain and consequently prevents activation and sensitization of central neurons postulated to be involved in migraine chronification.ConclusionOnabotulinumtoxinA likely acts via sensory mechanisms to treat chronic migraine.

Netrin-1 contributes to peripheral nerve injury induced neuropathic pain via regulating phosphatidylinositol 4-kinase IIa in the spinal cord dorsal horn in mice

BackgroundThe burden of neuropathic pain is growing worldwide. Recent studies recapitulate the requirement for AMPA receptor in excitatory synaptic plasticity underlying pain-related syndromes. Netrin-1 and its receptor deleted in colorectal cancer (DCC) are fundamental for AMPA receptor dependent synaptic transmission. Phosphatidylinositol 4-kinase IIa (Pi4KIIa) mediates post-synaptic insertion of AMPA receptor in neuropathic disorders. This study investigates whether netrin-1 and Pi4KIIa regulate peripheral nerve injury-induced neuropathic pain. MethodsA model of chronic constriction injury (CCI) of the sciatic nerve in mice was established to induce neuropathic pain. Paw withdrawal mechanical threshold, paw withdrawal thermal latency, spinal netrin-1 secretion, DCC level and Pi4KIIa expression were examined. Netrin-1 knockdown by shRNA, recombinant netrin-1 and Pi4KIIa inhibitor were employed to elucidate the substantial mechanisms. ResultsCCI surgery initiated and sustained the persistent reduction in paw withdrawal mechanical threshold and paw withdrawal thermal latency, along with the increase in spinal netrin-1 release, DCC level and Pi4KIIa expression. Netrin-1 deficiency impaired CCI-induced neuropathic pain behaviors and spinal over-expression of DCC and Pi4KIIa. Pharmacological inhibition of Pi4KIIa attenuated peripheral nerve injury induced mechanical allodynia and thermal hyperalgesia in a dose-dependent manner. Spinal application of recombinant netrin-1 caused pain hypersensitivity and up-regulated spinal expression of DCC and Pi4KIIa. Central inhibition of Pi4KIIa reversed exogenous netrin-1 evoked acute pain phenotype. ConclusionOur current results demonstrate the contribution of spinal netrin-1 and DCC in modulating the expression of Pi4KIIa in the pathogenesis of neuropathic pain in mice.

Stress induces insertion of calcium-permeable AMPA receptors in the OFC\u2013BLA synapse and modulates emotional behaviours in mice

Stress increases the risk of neuropsychiatric disorders, such as major depression. Exposure to stress has been reported to induce various neuronal changes, such as alterations in synaptic transmission and structure. However, a causal link between stress-induced neural circuit alterations and changes in emotional behaviours is not well understood. In the present study, we focused on a projection pathway from the orbitofrontal cortex (OFC) to the basolateral nucleus of the amygdala (BLA) as a crucial circuit for negative emotions and examined the effect of stress on OFC–BLA excitatory synaptic transmission using optogenetic and whole-cell patch-clamp methods in mice. As a stress-inducing procedure, we used repeated tail-shock, which increased stress-related behaviours. We found greater α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA)/N-methyl-d-aspartate current ratios and insertion of calcium-permeable AMPA receptors (AMPARs) in the OFC–BLA synapse after stress. These stress-induced synaptic and behavioural changes were reduced by a blockade of protein kinase A, which plays a principal role in stress-induced targeting of AMPARs into the synaptic membrane. To examine a possible causal relationship between alterations in synaptic transmission in the OFC–BLA pathway and stress-related behaviour, we performed optogenetic activation or chemogenetic inactivation of OFC–BLA transmission in mice. We found that optogenetic activation and chemogenetic inactivation of OFC–BLA transmission increased and decreased stress-related behaviour, respectively. In conclusion, we have demonstrated that stress altered the postsynaptic properties of the OFC–BLA pathway. These synaptic changes might be one of the underlying mechanisms of stress-induced behavioural alterations.

Lipid-Mediated Insertion of Toll-Like Receptor (TLR) Ligands for Facile Immune Cell Engineering.

Cell-based immunotherapies have tremendous potential to treat many diseases, such as activating immunity in cancer or suppressing it in autoimmune diseases. Most cell-based cancer immunotherapies in the clinic provide adjuvant signals through genetic engineering to enhance T cell functions. However, genetically encoded signals have minimal control over dosing and persist for the life of a cell lineage. These properties make it difficult to balance increasing therapeutic efficacy with reducing toxicities. Here, we demonstrated the potential of phospholipid-coupled ligands as a non-genetic system for immune cell engineering. This system provides simple, controlled, non-genetic adjuvant delivery to immune cells via lipid-mediated insertion into plasma membranes. Lipid-mediated insertion (termed depoting) successfully delivered Toll-like receptor (TLR) ligands intracellularly and onto cell surfaces of diverse immune cells. These ligands depoted into immune cells in a dose-controlled fashion and did not compete during multiplex pairwise loading. Immune cell activation could be enhanced by autocrine and paracrine mechanisms depending on the biology of the TLR ligand tested. Depoted ligands functionally persisted on plasma membranes for up to 4 days in naïve and activated T cells, enhancing their activation, proliferation, and skewing cytokine secretion. Our data showed that depoted ligands provided a persistent yet non-permanent adjuvant signal to immune cells that may minimize the intensity and duration of toxicities compared to permanent genetic delivery. Altogether, these findings demonstrate potential for lipid-mediated depoting as a universal cell engineering approach with unique, complementary advantages to other cell engineering methods.

Rapid exchange of synaptic and extrasynaptic NMDA receptors in hippocampal CA1 neurons.

N-methyl-d-aspartate receptors (NMDARs) are fundamental coincidence detectors of synaptic activity necessary for the induction of synaptic plasticity and synapse stability. Adjusting NMDAR synaptic content, whether by receptor insertion or lateral diffusion between extrasynaptic and synaptic compartments, could play a substantial role defining the characteristics of the NMDAR-mediated excitatory postsynaptic current (EPSC), which in turn would mediate the ability of the synapse to undergo plasticity. Lateral NMDAR movement has been observed in dissociated neurons; however, it is currently unclear whether NMDARs are capable of lateral surface diffusion in hippocampal slices, a more physiologically relevant environment. To test for lateral mobility in rat hippocampal slices, we rapidly blocked synaptic NMDARs using MK-801, a use-dependent and irreversible NMDAR blocker. Following a 5-min washout period, we observed a strong recovery of NMDAR-mediated responses. The degree of the observed recovery was proportional to the amount of induced blockade, independent of levels of intracellular calcium, and mediated primarily by GluN2B-containing NMDA receptors. These results indicate that lateral diffusion of NMDARs could be a mechanism by which synapses rapidly adjust parameters to fine-tune synaptic plasticity.NEW & NOTEWORTHYN-methyl-d-aspartate-type glutamate receptors (NMDARs) have always been considered stable components of synapses. We show that in rat hippocampal slices synaptic NMDARs are in constant exchange with extrasynaptic receptors. This exchange of receptors is mediated primarily by NMDA receptors containing GluN2B, a subunit necessary to undergo synaptic plasticity. Thus this lateral movement of synaptic receptors allows synapses to rapidly regulate the total number of synaptic NMDARs with potential consequences for synaptic plasticity.

AMPA receptor and metabotropic glutamate receptor 1 adaptations in the nucleus accumbens core during incubation of methamphetamine craving.

Cue-induced drug craving progressively intensifies after withdrawal from self-administration of cocaine, methamphetamine, and other drugs of abuse, a phenomenon termed incubation of craving. For cocaine and methamphetamine, expression of incubated craving ultimately depends on strengthening of nucleus accumbens (NAc) synapses through an accumulation of high conductance Ca2+-permeable AMPA receptors (CP-AMPARs) that is detectable with electrophysiological approaches. This study sought to further characterize glutamate receptor adaptations in NAc core during methamphetamine incubation. Previous biochemical studies revealed that the CP-AMPARs accumulating after cocaine incubation are mainly homomeric GluA1 receptors and that their accumulation is reflected by increased cell surface GluA1. Here, for methamphetamine, we observed no significant change in surface or total GluA1 (GluA2 and GluA3 were also unchanged). Nonetheless, GluA1 translation was elevated after incubation of methamphetamine craving, as recently found for cocaine. Additionally, for cocaine, we previously observed a withdrawal-dependent decrease in mGlu1 surface expression that precedes and enables CP-AMPAR accumulation and incubation of craving, reflecting weakening of mGlu1-dependent mechanisms that normally limit synaptic CP-AMPAR levels in the NAc core. Here, we observed no change in surface or total mGlu1 protein or its coupling to Homer scaffolding proteins after methamphetamine withdrawal, nor did elevation of mGlu1 tone through repeated injections of an mGlu1-positive allosteric modulator delay incubation of craving. These findings suggest a common role for increased GluA1 translation, but not decreased mGlu1 function, in the incubation of methamphetamine and cocaine craving. We speculate that increased GluA1 translation near synapses may drive formation and synaptic insertion of homomeric GluA1 receptors in the absence of detectable changes in GluA1 protein levels.

Impairment and Restoration of Homeostatic Plasticity in Cultured Cortical Neurons From a Mouse Model of Huntington Disease.

Huntington disease (HD) is an inherited neurodegenerative disorder caused by a mutation in the huntingtin gene. The onset of symptoms is preceded by synaptic dysfunction. Homeostatic synaptic plasticity (HSP) refers to processes that maintain the stability of networks of neurons, thought to be required to enable new learning and cognitive flexibility. One type of HSP is synaptic scaling, in which the strength of all of the synapses onto a cell increases or decreases following changes in the cell’s level of activity. Several pathways implicated in synaptic scaling are dysregulated in HD, including brain-derived neurotrophic factor (BDNF) and calcium signaling. Here, we investigated whether HSP is disrupted in cortical neurons from an HD mouse model. We treated cultured cortical neurons from wild-type (WT) FVB/N or YAC128 HD mice with tetrodotoxin (TTX) for 48 h to silence action potentials and then recorded miniature excitatory postsynaptic currents. In WT cultures, these increased in both amplitude and frequency after TTX treatment, and further experiments showed that this was a result of insertion of AMPA receptors and formation of new synapses, respectively. Manipulation of BDNF concentration in the culture medium revealed that BDNF signaling contributed to these changes. In contrast to WT cortical neurons, YAC128 cultures showed no response to action potential silencing. Strikingly, we were able to restore the TTX-induced changes in YAC128 cultures by treating them with pridopidine, a drug which enhances BDNF signaling through stimulation of the sigma-1 receptor (S1R), and with the S1R agonist 3-PPP. These data provide evidence for disruption of HSP in cortical neurons from an HD mouse model that is restored by stimulation of S1R. Our results suggest a potential new direction for developing therapy to mitigate cognitive deficits in HD.

Single and Repeated Administration of Methylphenidate Modulates Synaptic Plasticity in Opposite Directions via Insertion of AMPA Receptors in Rat Hippocampal Neurons.

Methylphenidate (MPH) is widely used in the treatment of Attention Deficit Hyperactivity Disorder. Several lines of evidence support that MPH can modulate learning and memory processes in different ways including improvement and impairment of test performances. A relevant factor in the efficacy of treatment is whether administration is performed once or several times. In this study we demonstrate opposite effects of MPH on performance of preadolescent rats in the Morris Water Maze test. Animals treated with a single dose (1 mg/kg) performed significantly better compared to controls, while in animals treated with repetitive administration at the same concentration performance was reduced. We found that hippocampal LTP in slices from rats treated with a single dose was increased, while LTP from rats treated with repetitive injections of MPH was lower than in controls. Using Western blot of CA1 areas from potentiated slices of rats treated with a single dose we found a significant increase of phosphorylation at Ser845 of GluA1 subunits, associated to an increased insertion of GluA1-containing AMPARs in the plasma membrane. These receptors were functional, because AMPA-dependent EPSCs recorded on CA1 were enhanced, associated to a significant increase in short-term plasticity. In contrast, CA1 samples from rats injected with MPH during six consecutive days, showed a significant decrease in the phosphorylation at Ser845 of GluA1 subunits associated to a lower insertion of GluA1-containing AMPARs. Accordingly, a reduction of the AMPA-mediated EPSCs and short-term plasticity was also observed. Taken together, our results demonstrate that single and repeated doses with MPH can induce opposite effects at behavioral, cellular, and molecular levels. The mechanisms demonstrated here in preadolescent rats are relevant to understand the effects of this psychostimulant in the treatment of ADHD.

The Antibody Response to Plasmodium falciparum: Cues for Vaccine Design and the Discovery of Receptor-Based Antibodies.

Plasmodium falciparum remains a serious public health problem and a continuous challenge for the immune system due to the complexity and diversity of the pathogen. Recent advances from several laboratories in the characterization of the antibody response to the parasite have led to the identification of critical targets for protection and revealed a new mechanism of diversification based on the insertion of host receptors into immunoglobulin genes, leading to the production of receptor-based antibodies. These advances have opened new possibilities for vaccine design and passive antibody therapies to provide sterilizing immunity and control blood-stage parasites.

Metformin produces anxiolytic-like effects in rats by facilitating GABAA receptor trafficking to membrane.

Altered function or expression of GABAA receptors contributes to anxiety disorders. Benzodiazepines are widely prescribed for the treatment of anxiety. However, the long-term use of benzodiazepines increases the risk of developing drug dependence and tolerance. Thus, it is urgent to explore new therapeutic approaches. Metformin is widely used to treat Type 2 diabetes and other metabolic syndromes. However, the role of metformin in psychiatric disorders, especially anxiety, remains largely unknown. We examined the effects of metformin on anxiety-like behaviour of rats in open field test and elevated plus maze test. We also observed the effect of metformin (10μM, in vitro; 100mg·kg-1 , in vivo) on the trafficking of GABAA receptors, as mechanisms underlying the anxiolytic effects of metformin. Metformin (100mg·kg-1 , i.p. 30min) displayed a robust and rapid anxiolytic effect, without tolerance. Metformin up-regulated the surface expression of GABAA receptors and increased miniature inhibitory postsynaptic currents (mIPSCs). AMP-activated protein kinase (AMPK) activated by metformin-induced stimulation of forkhead box O3a (FoxO3a) transcriptional activity, followed by increased expression of GABAA receptor-associated protein (GABARAP) and its binding to GABAA receptors finally resulted in the membrane insertion of GABAA receptors. Metformin increased mIPSCs by up-regulating the membrane insertion of GABAA receptors, via a pathway involving AMPK, FoxO3a, and the GABAA receptor-associated protein. Thus metformin has a potential new use in the treatment of anxiety disorders.