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1.
Cell Rep ; 37(6): 109972, 2021 11 09.
Article in English | MEDLINE | ID: mdl-34758304

ABSTRACT

Cortical function relies on the balanced activation of excitatory and inhibitory neurons. However, little is known about the organization and dynamics of shaft excitatory synapses onto cortical inhibitory interneurons. Here, we use the excitatory postsynaptic marker PSD-95, fluorescently labeled at endogenous levels, as a proxy for excitatory synapses onto layer 2/3 pyramidal neurons and parvalbumin-positive (PV+) interneurons in the barrel cortex of adult mice. Longitudinal in vivo imaging under baseline conditions reveals that, although synaptic weights in both neuronal types are log-normally distributed, synapses onto PV+ neurons are less heterogeneous and more stable. Markov model analyses suggest that the synaptic weight distribution is set intrinsically by ongoing cell-type-specific dynamics, and substantial changes are due to accumulated gradual changes. Synaptic weight dynamics are multiplicative, i.e., changes scale with weights, although PV+ synapses also exhibit an additive component. These results reveal that cell-type-specific processes govern cortical synaptic strengths and dynamics.


Subject(s)
Disks Large Homolog 4 Protein/physiology , Excitatory Postsynaptic Potentials/physiology , Interneurons/physiology , Neural Inhibition , Parvalbumins/metabolism , Pyramidal Cells/physiology , Synapses/physiology , Animals , Female , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Neuronal Plasticity
2.
Mol Brain ; 14(1): 129, 2021 08 21.
Article in English | MEDLINE | ID: mdl-34419133

ABSTRACT

Hypobaric hypoxia (HH) is a typical characteristic of high altitude environment and causes a spectrum of pathophysiological effects, including headaches, gliovascular dysfunction and cognitive retardation. Here, we sought to understand the mechanisms underlying cognitive deficits under HH exposure. Our results showed that hypobaric hypoxia exposure impaired cognitive function and suppressed dendritic spine density accompanied with increased neck length in both basal and apical hippocampal CA1 region neurons in mice. The expression of PSD95, a vital synaptic scaffolding molecule, is down-regulated by hypobaric hypoxia exposure and post-transcriptionally regulated by cold-inducible RNA-binding protein (Cirbp) through 3'-UTR region binding. PSD95 expressing alleviates hypoxia-induced dendritic spine morphology changes of hippocampal neurons and memory deterioration. Moreover, overexpressed Cirbp in hippocampus rescues HH-induced abnormal expression of PSD95 and attenuates hypoxia-induced dendritic spine injury and cognitive retardation. Thus, our findings reveal a novel mechanism that Cirbp-PSD-95 axis appears to play an essential role in HH-induced cognitive dysfunction in mice.


Subject(s)
Altitude Sickness/physiopathology , CA1 Region, Hippocampal/pathology , Cognition Disorders/prevention & control , Dendritic Spines/ultrastructure , Disks Large Homolog 4 Protein/physiology , RNA-Binding Proteins/physiology , 3' Untranslated Regions , Animals , Avoidance Learning , Base Sequence , Cells, Cultured , Cognition Disorders/etiology , Disks Large Homolog 4 Protein/biosynthesis , Disks Large Homolog 4 Protein/genetics , Gene Expression Regulation , Genes, Reporter , Genetic Vectors/administration & dosage , Memory Disorders/etiology , Memory Disorders/prevention & control , Mice , Mice, Inbred C57BL , Morris Water Maze Test , Neurons/physiology , Neurons/ultrastructure , Open Field Test , RNA-Binding Proteins/biosynthesis , RNA-Binding Proteins/genetics , Random Allocation , Recombinant Fusion Proteins/metabolism
3.
Nat Commun ; 12(1): 2849, 2021 05 14.
Article in English | MEDLINE | ID: mdl-33990590

ABSTRACT

Long-term depression (LTD) of synaptic strength can take multiple forms and contribute to circuit remodeling, memory encoding or erasure. The generic term LTD encompasses various induction pathways, including activation of NMDA, mGlu or P2X receptors. However, the associated specific molecular mechanisms and effects on synaptic physiology are still unclear. We here compare how NMDAR- or P2XR-dependent LTD affect synaptic nanoscale organization and function in rodents. While both LTDs are associated with a loss and reorganization of synaptic AMPARs, only NMDAR-dependent LTD induction triggers a profound reorganization of PSD-95. This modification, which requires the autophagy machinery to remove the T19-phosphorylated form of PSD-95 from synapses, leads to an increase in AMPAR surface mobility. We demonstrate that these post-synaptic changes that occur specifically during NMDAR-dependent LTD result in an increased short-term plasticity improving neuronal responsiveness of depressed synapses. Our results establish that P2XR- and NMDAR-mediated LTD are associated to functionally distinct forms of LTD.


Subject(s)
Disks Large Homolog 4 Protein/physiology , Long-Term Synaptic Depression/physiology , Receptors, N-Methyl-D-Aspartate/physiology , Adenosine Triphosphate/administration & dosage , Animals , Autophagy/physiology , Cells, Cultured , Disks Large Homolog 4 Protein/deficiency , Female , Hippocampus/cytology , Hippocampus/physiology , In Vitro Techniques , Male , Mice , Mice, Inbred C57BL , Miniature Postsynaptic Potentials/physiology , Models, Neurological , N-Methylaspartate/administration & dosage , Neuronal Plasticity/physiology , Neurons/cytology , Neurons/drug effects , Neurons/physiology , Rats , Rats, Sprague-Dawley , Receptors, AMPA/physiology , Receptors, Purinergic P2X/physiology
4.
J Neurosci ; 41(11): 2329-2343, 2021 03 17.
Article in English | MEDLINE | ID: mdl-33472821

ABSTRACT

Cognitive processes that require spatial information rely on synaptic plasticity in the dorsal CA1 area (dCA1) of the hippocampus. Since the function of the hippocampus is impaired in aged individuals, it remains unknown how aged animals make spatial choices. Here, we used IntelliCage to study behavioral processes that support spatial choices of aged female mice living in a group. As a proxy of training-induced synaptic plasticity, we analyzed the morphology of dendritic spines and the expression of a synaptic scaffold protein, PSD-95. We observed that spatial choice training in young adult mice induced correlated shrinkage of dendritic spines and downregulation of PSD-95 in dCA1. Moreover, long-term depletion of PSD-95 by shRNA in dCA1 limited correct choices to a reward corner, while reward preference was intact. In contrast, old mice used behavioral strategies characterized by an increased tendency for perseverative visits and social interactions. This strategy resulted in a robust preference for the reward corner during the spatial choice task. Moreover, training decreased the correlation between PSD-95 expression and the size of dendritic spines. Furthermore, PSD-95 depletion did not impair place choice or reward preference in old mice. Thus, our data indicate that while young mice require PSD-95-dependent synaptic plasticity in dCA1 to make correct spatial choices, old animals observe cage mates and stick to a preferred corner to seek the reward. This strategy is resistant to the depletion of PSD-95 in the CA1 area. Overall, our study demonstrates that aged mice combine alternative behavioral and molecular strategies to approach and consume rewards in a complex environment.SIGNIFICANCE STATEMENT It remains poorly understood how aging affects behavioral and molecular processes that support cognitive functions. It is, however, essential to understand these processes to develop therapeutic interventions that support successful cognitive aging. Our data indicate that while young mice require PSD-95-dependent synaptic plasticity in dCA1 to make correct spatial choices (i.e., choices that require spatial information), old animals observe cage mates and stick to a preferred corner to seek the reward. This strategy is resistant to the depletion of PSD-95 in the CA1 area. Overall, our study demonstrates that aged mice combine alternative behavioral and molecular strategies to approach and consume rewards in a complex environment. Second, the contribution of PSD-95-dependent synaptic functions in spatial choice changes with age.


Subject(s)
CA1 Region, Hippocampal/physiology , Choice Behavior/physiology , Disks Large Homolog 4 Protein/physiology , Space Perception/physiology , Aging/physiology , Aging/psychology , Animals , Dendritic Spines/physiology , Disks Large Homolog 4 Protein/genetics , Environment , Female , Gene Expression Regulation/genetics , Mice , Mice, Inbred C57BL , Neuronal Plasticity/physiology , Reward , Social Interaction
5.
FASEB J ; 34(9): 12239-12254, 2020 09.
Article in English | MEDLINE | ID: mdl-33000527

ABSTRACT

α-Synuclein (α-syn)-induced neurotoxicity has been generally accepted as a key step in the pathogenesis of Parkinson's disease (PD). Microtubule-associated protein tau, which is considered second only to α-syn, has been repeatedly linked with PD in association studies. However, the underlying interaction between these two PD-related proteins in vivo remains unclear. To investigate how the expression of tau affects α-syn-induced neurodegeneration in vivo, we generated triple transgenic mice that overexpressed α-syn A53T mutation in the midbrain dopaminergic neurons (mDANs) with different expression levels of tau. Here, we found that tau had no significant effect on the A53T α-syn-mediated mDANs degeneration. However, tau knockout could modestly promote the formation of α-syn aggregates, accelerate the severe and progressive degeneration of parvalbumin-positive (PV+) neurons in substantia nigra pars reticulata (SNR), accompanied with anxiety-like behavior in aged PD-related α-syn A53T mice. The mechanisms may be associated with A53T α-syn-mediated specifically successive impairment of N-methyl-d-aspartate receptor subunit 2B (NR2B), postsynaptic density-95 (PSD-95) and microtubule-associated protein 1A (MAP1A) in PV+ neurons. Our study indicates that MAP1A may play a beneficial role in preserving the survival of PV+ neurons, and that inhibition of the impairment of NR2B/PSD-95/MAP1A pathway, may be a novel and preferential option to ameliorate α-syn-induced neurodegeneration.


Subject(s)
Mutation , Nerve Degeneration , Parkinson Disease/etiology , Parvalbumins/analysis , Substantia Nigra/pathology , alpha-Synuclein/genetics , tau Proteins/physiology , Animals , Disks Large Homolog 4 Protein/physiology , Homeodomain Proteins/physiology , Mice , Mice, Inbred C57BL , Microtubule-Associated Proteins/physiology , Parkinson Disease/pathology , Peptide Fragments/physiology , Protein Aggregates , Receptors, N-Methyl-D-Aspartate/physiology , Transcription Factors/physiology , alpha-Synuclein/physiology , tau Proteins/chemistry , tau Proteins/genetics
6.
Sci Rep ; 10(1): 14014, 2020 08 19.
Article in English | MEDLINE | ID: mdl-32814795

ABSTRACT

Determining the number of synapses that are present in different brain regions is crucial to understand brain connectivity as a whole. Membrane-associated guanylate kinases (MAGUKs) are a family of scaffolding proteins that are expressed in excitatory glutamatergic synapses. We used genetic labeling of two of these proteins (PSD95 and SAP102), and Spinning Disc confocal Microscopy (SDM), to estimate the number of fluorescent puncta in the CA1 area of the hippocampus. We also used FIB-SEM, a three-dimensional electron microscopy technique, to calculate the actual numbers of synapses in the same area. We then estimated the ratio between the three-dimensional densities obtained with FIB-SEM (synapses/µm3) and the bi-dimensional densities obtained with SDM (puncta/100 µm2). Given that it is impractical to use FIB-SEM brain-wide, we used previously available SDM data from other brain regions and we applied this ratio as a conversion factor to estimate the minimum density of synapses in those regions. We found the highest densities of synapses in the isocortex, olfactory areas, hippocampal formation and cortical subplate. Low densities were found in the pallidum, hypothalamus, brainstem and cerebellum. Finally, the striatum and thalamus showed a wide range of synapse densities.


Subject(s)
Brain/physiology , Disks Large Homolog 4 Protein/physiology , Guanylate Kinases/physiology , Hippocampus/physiology , Membrane Proteins/physiology , Synapses/physiology , Animals , Brain/ultrastructure , Hippocampus/ultrastructure , Male , Mice , Mice, Knockout , Microscopy, Electron , Synapses/ultrastructure
7.
Anesthesiology ; 133(4): 812-823, 2020 10 01.
Article in English | MEDLINE | ID: mdl-32773681

ABSTRACT

BACKGROUND: Experimental evidence shows postnatal exposure to anesthesia negatively affects brain development. The PDZ2 domain, mediating protein-protein interactions of the postsynaptic density-95 protein, serves as a molecular target for several inhaled anesthetics. The authors hypothesized that early postnatal disruption of postsynaptic density-95 PDZ2 domain interactions has persistent effects on dendritic spines and cognitive function. METHODS: One-week-old mice were exposed to 1.5% isoflurane for 4 h or injected with 8 mg/kg active postsynaptic density-95 wild-type PDZ2 peptide along with their respective controls. A subset of these mice also received 4 mg/kg of the nitric oxide donor molsidomine. Hippocampal spine density, long-term potentiation, novel object recognition memory, and fear learning and memory were evaluated in mice. RESULTS: Exposure of 7-day-old mice to isoflurane or postsynaptic density-95 wild-type PDZ2 peptide relative to controls causes: (1) a long-term decrease in mushroom spines at 7 weeks (mean ± SD [spines per micrometer]): control (0.8 ± 0.2) versus isoflurane (0.4 ± 0.2), P < 0.0001, and PDZ2MUT (0.7 ± 0.2) versus PDZ2WT (0.4 ± 0.2), P < 0.001; (2) deficits in object recognition at 6 weeks (mean ± SD [recognition index]): naïve (70 ± 8) versus isoflurane (55 ± 14), P = 0.010, and control (65 ± 13) versus isoflurane (55 ± 14), P = 0.045, and PDZ2MUT (64 ±11) versus PDZ2WT (53 ± 18), P = 0.045; and (3) deficits in fear learning at 7 weeks and memory at 8 weeks (mean ± SD [% freezing duration]): Learning, control (69 ± 12) versus isoflurane (52 ± 13), P < 0.0001, and PDZ2MUT (65 ± 14) versus PDZ2WT (55 ± 14) P = 0.011, and Memory, control (80 ± 17) versus isoflurane (56 ± 23), P < 0.0001 and PDZ2MUT (73 ± 18) versus PDZ2WT (44 ± 19) P < 0.0001. Impairment in long-term potentiation has fully recovered here at 7 weeks (mean ± SD [% baseline]): control (140 ± 3) versus isoflurane (137 ± 8), P = 0.560, and PDZ2MUT (136 ± 17) versus PDZ2WT (128 ± 11), P = 0.512. The isoflurane induced decrease in mushroom spines was preventable by introduction of a nitric oxide donor. CONCLUSIONS: Early disruption of PDZ2 domain-mediated protein-protein interactions mimics isoflurane in decreasing mushroom spine density and causing learning and memory deficits in mice. Prevention of the decrease in mushroom spine density with a nitric oxide donor supports a role for neuronal nitric oxide synthase pathway in mediating this cellular change associated with cognitive impairment.


Subject(s)
Anesthetics, Inhalation/toxicity , Cognition/drug effects , Dendritic Spines/drug effects , Disks Large Homolog 4 Protein/antagonists & inhibitors , Isoflurane/toxicity , Animals , Animals, Newborn , Cognition/physiology , Dendritic Spines/pathology , Dendritic Spines/physiology , Disks Large Homolog 4 Protein/physiology , Female , Male , Mice , Mice, Inbred C57BL , Organ Culture Techniques , Peptides/pharmacology , Post-Synaptic Density/drug effects , Post-Synaptic Density/pathology , Post-Synaptic Density/physiology
8.
Mol Neurobiol ; 57(5): 2479-2493, 2020 May.
Article in English | MEDLINE | ID: mdl-32157575

ABSTRACT

CRIPT, the cysteine-rich PDZ-binding protein, binds to the third PDZ domain of PSD-95 (postsynaptic density protein 95) family proteins and directly binds microtubules, linking PSD-95 family proteins to the neuronal cytoskeleton. Here, we show that overexpression of a full-length CRIPT leads to a modest decrease, and knockdown of CRIPT leads to an increase in dendritic branching in cultured rat hippocampal neurons. Overexpression of truncated CRIPT lacking the PDZ domain-binding motif, which does not bind to PSD-95, significantly decreases dendritic arborization. Conversely, overexpression of a full-length CRIPT significantly increases the number of immature and mature dendritic spines, and this effect is not observed when CRIPT∆PDZ is overexpressed. Competitive inhibition of CRIPT binding to the third PDZ domain of PSD-95 with PDZ3-binding peptides resulted in differential effects on dendritic arborization based on the origin of respective peptide sequence. These results highlight multifunctional roles of CRIPT during development and underscore the significance of the interaction between CRIPT and the third PDZ domain of PSD-95.


Subject(s)
Adaptor Proteins, Signal Transducing/physiology , Disks Large Homolog 4 Protein/physiology , Hippocampus/cytology , Neuronal Plasticity/physiology , Adaptor Proteins, Signal Transducing/antagonists & inhibitors , Adaptor Proteins, Signal Transducing/genetics , Amino Acid Motifs , Animals , Binding, Competitive , Cells, Cultured , Dendritic Spines/physiology , Dendritic Spines/ultrastructure , Gene Knockdown Techniques , Microtubules/metabolism , Microtubules/ultrastructure , Protein Binding , Protein Interaction Mapping , RNA Interference , RNA, Small Interfering/genetics , Rats
9.
Behav Neurol ; 2019: 1068260, 2019.
Article in English | MEDLINE | ID: mdl-31772680

ABSTRACT

OBJECTIVE: To explore the effects of sevoflurane on the latency and error times of the passive avoidance and levels of PSD-95 and AMPA receptors in the hippocampus. We evaluated the effects of sevoflurane on short-term memory in adult mice and explored the possible mechanism. METHODS: 144 Kunming mice (2-3 months, 30-35 g) were randomly divided into two groups A (n = 64) and B (n = 80) and received the dark-avoidance (DA) and step-down avoidance (SA) tests, respectively. The groups DA and SA were further divided into control (inhaled 40% O2 2 h) and sevoflurane (3.3% sevoflurane and 40% O2 2 h) subgroups. Before inhalation intervention, all mice were trained to be familiar with the Morris water maze (MWM). According to the test points of behavioral indicators, 8 mice were randomly selected from each subgroup at point 12 h (T1), 24 h (T2), 48 h (T3), and 72 h (T4) after inhalation intervention. The step-through latency and error times were measured in 5 min. After the behavioral test, the mice were killed and the tissues of the hippocampus were taken for hematoxylin and eosin (H&E) staining. The expression level of PSD-95 and AMPA receptors in the hippocampus was detected by immunohistochemistry and Western Blot. The changes of synaptic transmission were measured via electrophysiology analysis of hippocampal slices. RESULTS: The mice in the control subgroups found the platform in a shorter pathway than those in the sevoflurane subgroups during an MWM test. The step-through latency of T1 and T2 in the sevoflurane subgroup was shorter than baseline time, and the error times were increased in 5 min and higher than baseline time when compared with the control subgroup (P < 0.05) in the A and B groups. Compared with the control subgroup, the expression level of PSD-95 and AMPA receptors in the hippocampus was decreased at T1 and T2 in the sevoflurane subgroup (P < 0.05). The nerve cells were partially swelling. Electrophysiology analysis showed that the levels of PSD-95 and AMPA receptor expression were associated with synaptic transmission. CONCLUSION: Sevoflurane impaired short-term memory in adult mice by inhibiting the expression of PSD-95 and AMPA receptors in the hippocampus, which led to the decrease in synaptic transmission.


Subject(s)
Memory, Short-Term/drug effects , Sevoflurane/adverse effects , Animals , Brain/metabolism , China , Disks Large Homolog 4 Protein/metabolism , Disks Large Homolog 4 Protein/physiology , Female , Hippocampus/metabolism , Male , Maze Learning/drug effects , Memory, Short-Term/physiology , Mice , Neurons/metabolism , Receptors, AMPA/metabolism , Receptors, AMPA/physiology , Sevoflurane/pharmacology
10.
Sci Rep ; 9(1): 14060, 2019 Oct 01.
Article in English | MEDLINE | ID: mdl-31575955

ABSTRACT

AMPA receptors and interacting proteins are importantly involved in mediating stress-dependent plasticity. Previously we reported that GluA1-containing AMPA receptors and their interaction with PDZ-proteins are required for the experience-dependent expression of behavioral despair in the forced swim test. However, it is unclear if the expression of GluA1-containing AMPA receptors is affected by this type of behavior. Here we investigated in wild type mice, whether hippocampal gene or protein levels of GluA1 or associated PDZ proteins is altered following forced swim stress. We show that expression of Dlg4 (the gene coding for PSD-95) was strongly reduced after two days of forced swimming. In contrast, levels of Dlg1, Gria1, and Gria2 (coding for SAP97, GluA1, and GluA2 respectively) were not affected after one or two days of forced swimming. The changes in gene expression largely did not translate to the protein level. These findings indicate a limited acute effect of forced swim stress on the expression of the investigated targets and suggest that the acute involvement of GluA1-containing AMPA receptors tor forced swim behavior is a result of non-genomic mechanisms.


Subject(s)
Discs Large Homolog 1 Protein/metabolism , Disks Large Homolog 4 Protein/metabolism , Hippocampus/metabolism , Receptors, AMPA/metabolism , Animals , Blotting, Western , Discs Large Homolog 1 Protein/analysis , Discs Large Homolog 1 Protein/physiology , Disks Large Homolog 4 Protein/analysis , Disks Large Homolog 4 Protein/physiology , Female , Gene Expression Regulation , Hippocampus/chemistry , Hippocampus/physiology , Mice , Mice, Inbred C57BL , Real-Time Polymerase Chain Reaction , Receptors, AMPA/analysis , Receptors, AMPA/physiology , Stress, Physiological/physiology , Swimming
11.
J Neurosci ; 39(5): 876-887, 2019 01 30.
Article in English | MEDLINE | ID: mdl-30530507

ABSTRACT

Nitric oxide (NO) is a neurotransmitter synthesized in the brain by neuronal nitric oxide synthase (nNOS). Using immunohistochemistry and confocal imaging in the inferior colliculus (IC, auditory midbrain) of the guinea pig (Cavia porcellus, male and female), we show that nNOS occurs in two distinct cellular distributions. We confirm that, in the cortices of the IC, a subset of neurons show cytoplasmic labeling for nNOS, whereas in the central nucleus (ICc), such neurons are not present. However, we demonstrate that all neurons in the ICc do in fact express nNOS in the form of discrete puncta found at the cell membrane. Our multi-labeling studies reveal that nNOS puncta form multiprotein complexes with NMDA receptors, soluble guanylyl cyclase (sGC), and PSD95. These complexes are found apposed to glutamatergic terminals, which is indicative of synaptic function. Interestingly, these glutamatergic terminals express both vesicular glutamate transporters 1 and 2 denoting a specific source of brainstem inputs. With in vivo electrophysiological recordings of multiunit activity in the ICc, we found that local application of NMDA enhances sound-driven activity in a concentration-dependent and reversible fashion. This response is abolished by blockade of nNOS or sGC, indicating that the NMDA effect is mediated solely via the NO and cGMP signaling pathway. This discovery of a ubiquitous, but highly localized, expression of nNOS throughout the ICc and demonstration of the dramatic influence of the NMDA activated NO pathway on sound-driven neuronal activity imply a key role for NO signaling in auditory processing.SIGNIFICANCE STATEMENT We show that neuronal nitric oxide synthase (nNOS), the enzyme that synthesizes nitric oxide (NO), occurs as puncta in apparently all neurons in the central nucleus of the inferior colliculus (ICc) in the auditory midbrain. Punctate nNOS appears at glutamatergic synapses in a complex with glutamate NMDA receptors (NMDA-Rs), soluble guanylyl cyclase (sGC, the NO receptor), and PSD95 (a protein that anchors receptors and enzymes at the postsynaptic density). We show that NMDA-R modulation of sound-driven activity in the ICc is solely mediated by activation of nNOS and sGC. The presence of nNOS throughout this sensory nucleus argues for a major role of NO in hearing. Furthermore, this punctate form of nNOS expression may exist and have gone unnoticed in other brain regions.


Subject(s)
Auditory Cortex/physiology , Mesencephalon/physiology , Nitric Oxide Synthase Type I/physiology , Receptors, N-Methyl-D-Aspartate/physiology , Signal Transduction/physiology , Animals , Auditory Perception/physiology , Cyclic GMP/physiology , Disks Large Homolog 4 Protein/physiology , Female , Guinea Pigs , Inferior Colliculi/cytology , Inferior Colliculi/physiology , Male , Nitric Oxide/physiology , Nitric Oxide Synthase Type I/metabolism , Soluble Guanylyl Cyclase/metabolism , Synapses/physiology , Vesicular Glutamate Transport Proteins/metabolism
12.
PLoS Biol ; 16(12): e2006838, 2018 12.
Article in English | MEDLINE | ID: mdl-30586380

ABSTRACT

The disc-large (DLG)-membrane-associated guanylate kinase (MAGUK) family of proteins forms a central signaling hub of the glutamate receptor complex. Among this family, some proteins regulate developmental maturation of glutamatergic synapses, a process vulnerable to aberrations, which may lead to neurodevelopmental disorders. As is typical for paralogs, the DLG-MAGUK proteins postsynaptic density (PSD)-95 and PSD-93 share similar functional domains and were previously thought to regulate glutamatergic synapses similarly. Here, we show that they play opposing roles in glutamatergic synapse maturation. Specifically, PSD-95 promoted, whereas PSD-93 inhibited maturation of immature α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid-type glutamate receptor (AMPAR)-silent synapses in mouse cortex during development. Furthermore, through experience-dependent regulation of its protein levels, PSD-93 directly inhibited PSD-95's promoting effect on silent synapse maturation in the visual cortex. The concerted function of these two paralogs governed the critical period of juvenile ocular dominance plasticity (jODP), and fine-tuned visual perception during development. In contrast to the silent synapse-based mechanism of adjusting visual perception, visual acuity improved by different mechanisms. Thus, by controlling the pace of silent synapse maturation, the opposing but properly balanced actions of PSD-93 and PSD-95 are essential for fine-tuning cortical networks for receptive field integration during developmental critical periods, and imply aberrations in either direction of this process as potential causes for neurodevelopmental disorders.


Subject(s)
Disks Large Homolog 4 Protein/physiology , Guanylate Kinases/physiology , Membrane Proteins/physiology , Synapses/metabolism , Animals , Disks Large Homolog 4 Protein/metabolism , Excitatory Amino Acid Agents , Female , Glutamic Acid/metabolism , Guanylate Kinases/metabolism , Intracellular Signaling Peptides and Proteins/metabolism , Male , Membrane Proteins/metabolism , Mice , Mice, Inbred C57BL , Mice, Knockout , Neurons/physiology , Receptors, AMPA/metabolism , Receptors, Glutamate/metabolism , Receptors, N-Methyl-D-Aspartate/metabolism , Signal Transduction , Synaptic Transmission/physiology , Visual Cortex/metabolism
13.
Cell ; 174(5): 1172-1187.e16, 2018 08 23.
Article in English | MEDLINE | ID: mdl-30078712

ABSTRACT

Synapses are semi-membraneless, protein-dense, sub-micron chemical reaction compartments responsible for signal processing in each and every neuron. Proper formation and dynamic responses to stimulations of synapses, both during development and in adult, are fundamental to functions of mammalian brains, although the molecular basis governing formation and modulation of compartmentalized synaptic assemblies is unclear. Here, we used a biochemical reconstitution approach to show that, both in solution and on supported membrane bilayers, multivalent interaction networks formed by major excitatory postsynaptic density (PSD) scaffold proteins led to formation of PSD-like assemblies via phase separation. The reconstituted PSD-like assemblies can cluster receptors, selectively concentrate enzymes, promote actin bundle formation, and expel inhibitory postsynaptic proteins. Additionally, the condensed phase PSD assemblies have features that are distinct from those in homogeneous solutions and fit for synaptic functions. Thus, we have built a molecular platform for understanding how neuronal synapses are formed and dynamically regulated.


Subject(s)
Neurogenesis , Neuronal Plasticity , Post-Synaptic Density , Synapses/physiology , Animals , Brain/physiology , Disks Large Homolog 4 Protein/physiology , Hippocampus/physiology , Light , Mice , Microscopy, Confocal , Neurons/physiology , Scattering, Radiation , Signal Transduction , Synaptic Transmission
14.
Microcirculation ; 25(1)2018 01.
Article in English | MEDLINE | ID: mdl-29072364

ABSTRACT

Voltage-gated K+ (Kv ) channels are major determinants of membrane potential in vascular smooth muscle cells (VSMCs) and regulate the diameter of small cerebral arteries and arterioles. However, the intracellular structures that govern the expression and function of vascular Kv channels are poorly understood. Scaffolding proteins including postsynaptic density 95 (PSD95) recently were identified in rat cerebral VSMCs. Primarily characterized in neurons, the PSD95 scaffold has more than 50 known binding partners, and it can mediate macromolecular signaling between cell-surface receptors and ion channels. In cerebral arteries, Shaker-type Kv 1 channels appear to associate with the PSD95 molecular scaffold, and PSD95 is required for the normal expression and vasodilator influence of members of this K+ channel gene family. Furthermore, recent findings suggest that the ß1-subtype adrenergic receptor is expressed in cerebral VSMCs and forms a functional vasodilator complex with Kv 1 channels on the PSD95 scaffold. Activation of ß1-subtype adrenergic receptors in VSMCs enables protein kinase A-dependent phosphorylation and opening of Kv 1 channels in the PSD95 complex; the subsequent K+ efflux mediates membrane hyperpolarization and vasodilation of small cerebral arteries. Early evidence from other studies suggests that other families of Kv channels and scaffolding proteins are expressed in VSMCs. Future investigations into these macromolecular complexes that modulate the expression and function of Kv channels may reveal unknown signaling cascades that regulate VSMC excitability and provide novel targets for ion channel-based medications to optimize vascular tone.


Subject(s)
Cerebrovascular Circulation , Disks Large Homolog 4 Protein/physiology , Potassium Channels, Voltage-Gated/metabolism , Receptors, Adrenergic, beta/physiology , Animals , Disks Large Homolog 4 Protein/metabolism , Humans , Muscle, Smooth, Vascular/chemistry , Muscle, Smooth, Vascular/cytology , Rats , Receptors, Adrenergic, beta/metabolism
15.
Proc Natl Acad Sci U S A ; 114(41): E8760-E8769, 2017 10 10.
Article in English | MEDLINE | ID: mdl-28973854

ABSTRACT

Ubiquitination-directed proteasomal degradation of synaptic proteins, presumably mediated by lysine 48 (K48) of ubiquitin, is a key mechanism in synapse and neural circuit remodeling. However, more than half of polyubiquitin (polyUb) species in the mammalian brain are estimated to be non-K48; among them, the most abundant is Lys 63 (K63)-linked polyUb chains that do not tag substrates for degradation but rather modify their properties and activity. Virtually nothing is known about the role of these nonproteolytic polyUb chains at the synapse. Here we report that K63-polyUb chains play a significant role in postsynaptic protein scaffolding and synaptic strength and plasticity. We found that the postsynaptic scaffold PSD-95 (postsynaptic density protein 95) undergoes K63 polyubiquitination, which markedly modifies PSD-95's scaffolding potentials, enables its synaptic targeting, and promotes synapse maturation and efficacy. TNF receptor-associated factor 6 (TRAF6) is identified as a direct E3 ligase for PSD-95, which, together with the E2 complex Ubc13/Uev1a, assembles K63-chains on PSD-95. In contrast, CYLD (cylindromatosis tumor-suppressor protein), a K63-specific deubiquitinase enriched in postsynaptic densities, cleaves K63-chains from PSD-95. We found that neuronal activity exerts potent control of global and synaptic K63-polyUb levels and, through NMDA receptors, drives rapid, CYLD-mediated PSD-95 deubiquitination, mobilizing and depleting PSD-95 from synapses. Silencing CYLD in hippocampal neurons abolishes NMDA-induced chemical long-term depression. Our results unveil a previously unsuspected role for nonproteolytic polyUb chains in the synapse and illustrate a mechanism by which a PSD-associated K63-linkage-specific ubiquitin machinery acts on a major postsynaptic scaffold to regulate synapse organization, function, and plasticity.


Subject(s)
Disks Large Homolog 4 Protein/physiology , Hippocampus/physiology , Neurons/physiology , Polyubiquitin/metabolism , Post-Synaptic Density , Proteasome Endopeptidase Complex/metabolism , Synapses/physiology , Animals , Hippocampus/cytology , Lysine , Mice , Mice, Knockout , Neurons/cytology , Ubiquitination
16.
Med Sci Monit ; 23: 4954-4960, 2017 Oct 17.
Article in English | MEDLINE | ID: mdl-29038420

ABSTRACT

BACKGROUND Emotional state can be affected by different training loads. The aim of this study was to explore the changes of rat emotional state, as well as the mRNA and protein expressions of N-methyl-D-aspartate receptors (NMDARs), postsynaptic density 95 (PSD-95), and kinesin family member 17 (KIF-17) in the hippocampus, by long-term moderate-intensity and high-intensity training models in rats. MATERIAL AND METHODS The exercise model of SD rats was set up by treadmill running of moderate and high intensities for 4 weeks. The rats in the moderate-intensity training group were given endurance training with increasing intensity, while rats in the high-intensity training group were given high-speed training, and those in the normal control group were also established. The body weights of rats were measured before and after exercise to determine weight reduction. Real-time PCR and Western blotting were used to detect the mRNA and protein expressions of NMDARs, PSD-95, and KIF-17 in hippocampus of rats under different training loads. RESULTS Compared with the control group, the rats in the moderate-intensity training group had better body condition and emotional state, while the rats in the high-intensity training group had poor body condition and emotional state. The mRNA and protein expression of PSD-95, KIF-17, and NMDARs in the moderate-intensity training group were significantly elevated (P<0.05) while those in the high-intensity training group were suppressed (P<0.05). CONCLUSIONS Different training loads have remarkable influences on the cognition, emotion, and mental status of rats, and can affect the mRNA and protein expressions of NMDARs, PSD-95, and KIF-17 in rats. Appropriate training loads alleviate hypoxia damage to the hippocampus, and also effectively improve hippocampus function.


Subject(s)
Emotions/physiology , Physical Conditioning, Animal/physiology , Animals , Blotting, Western , Body Weight , Disks Large Homolog 4 Protein/metabolism , Disks Large Homolog 4 Protein/physiology , Gene Expression Regulation/genetics , Hippocampus/metabolism , Hippocampus/physiology , Intracellular Signaling Peptides and Proteins/metabolism , Kinesins/metabolism , Kinesins/physiology , Male , Post-Synaptic Density , RNA, Messenger , Rats , Rats, Sprague-Dawley , Receptors, N-Methyl-D-Aspartate/metabolism , Receptors, N-Methyl-D-Aspartate/physiology
17.
Proc Natl Acad Sci U S A ; 113(32): E4736-44, 2016 08 09.
Article in English | MEDLINE | ID: mdl-27457929

ABSTRACT

Phosphorylation regulates surface and synaptic expression of NMDA receptors (NMDARs). Both the tyrosine kinase Fyn and the tyrosine phosphatase striatal-enriched protein tyrosine phosphatase (STEP) are known to target the NMDA receptor subunit GluN2B on tyrosine 1472, which is a critical residue that mediates NMDAR endocytosis. STEP reduces the surface expression of NMDARs by promoting dephosphorylation of GluN2B Y1472, whereas the synaptic scaffolding protein postsynaptic density protein 95 (PSD-95) stabilizes the surface expression of NMDARs. However, nothing is known about a potential functional interaction between STEP and PSD-95. We now report that STEP61 binds to PSD-95 but not to other PSD-95 family members. We find that PSD-95 expression destabilizes STEP61 via ubiquitination and degradation by the proteasome. Using subcellular fractionation, we detect low amounts of STEP61 in the PSD fraction. However, STEP61 expression in the PSD is increased upon knockdown of PSD-95 or in vivo as detected in PSD-95-KO mice, demonstrating that PSD-95 excludes STEP61 from the PSD. Importantly, only extrasynaptic NMDAR expression and currents were increased upon STEP knockdown, as is consistent with low STEP61 localization in the PSD. Our findings support a dual role for PSD-95 in stabilizing synaptic NMDARs by binding directly to GluN2B but also by promoting synaptic exclusion and degradation of the negative regulator STEP61.


Subject(s)
Disks Large Homolog 4 Protein/physiology , Protein Tyrosine Phosphatases, Non-Receptor/metabolism , Receptors, N-Methyl-D-Aspartate/physiology , Animals , Female , HEK293 Cells , Humans , Mice , Proteasome Endopeptidase Complex/physiology , Protein Tyrosine Phosphatases, Non-Receptor/analysis , Rats , Rats, Sprague-Dawley , Receptors, N-Methyl-D-Aspartate/metabolism , Ubiquitination
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