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1.
Nat Commun ; 15(1): 2142, 2024 Mar 08.
Article in English | MEDLINE | ID: mdl-38459070

ABSTRACT

Neuronal mitochondria play important roles beyond ATP generation, including Ca2+ uptake, and therefore have instructive roles in synaptic function and neuronal response properties. Mitochondrial morphology differs significantly between the axon and dendrites of a given neuronal subtype, but in CA1 pyramidal neurons (PNs) of the hippocampus, mitochondria within the dendritic arbor also display a remarkable degree of subcellular, layer-specific compartmentalization. In the dendrites of these neurons, mitochondria morphology ranges from highly fused and elongated in the apical tuft, to more fragmented in the apical oblique and basal dendritic compartments, and thus occupy a smaller fraction of dendritic volume than in the apical tuft. However, the molecular mechanisms underlying this striking degree of subcellular compartmentalization of mitochondria morphology are unknown, precluding the assessment of its impact on neuronal function. Here, we demonstrate that this compartment-specific morphology of dendritic mitochondria requires activity-dependent, Ca2+ and Camkk2-dependent activation of AMPK and its ability to phosphorylate two direct effectors: the pro-fission Drp1 receptor Mff and the recently identified anti-fusion, Opa1-inhibiting protein, Mtfr1l. Our study uncovers a signaling pathway underlying the subcellular compartmentalization of mitochondrial morphology in dendrites of neurons in vivo through spatially precise and activity-dependent regulation of mitochondria fission/fusion balance.


Subject(s)
Neurons , Pyramidal Cells , Neurons/metabolism , Pyramidal Cells/physiology , Hippocampus , Axons/metabolism , Mitochondria/metabolism , Dendrites/physiology
2.
bioRxiv ; 2023 Mar 26.
Article in English | MEDLINE | ID: mdl-36993655

ABSTRACT

Neuronal mitochondria play important roles beyond ATP generation, including Ca2+ uptake, and therefore have instructive roles in synaptic function and neuronal response properties. Mitochondrial morphology differs significantly in the axon and dendrites of a given neuronal subtype, but in CA1 pyramidal neurons (PNs) of the hippocampus, mitochondria within the dendritic arbor also display a remarkable degree of subcellular, layer-specific compartmentalization. In the dendrites of these neurons, mitochondria morphology ranges from highly fused and elongated in the apical tuft, to more fragmented in the apical oblique and basal dendritic compartments, and thus occupy a smaller fraction of dendritic volume than in the apical tuft. However, the molecular mechanisms underlying this striking degree of subcellular compartmentalization of mitochondria morphology are unknown, precluding the assessment of its impact on neuronal function. Here, we demonstrate that this compartment-specific morphology of dendritic mitochondria requires activity-dependent, Camkk2-dependent activation of AMPK and its ability to phosphorylate two direct effectors: the pro-fission Drp1 receptor Mff and the recently identified anti-fusion, Opa1-inhibiting protein, Mtfr1l. Our study uncovers a new activity-dependent molecular mechanism underlying the extreme subcellular compartmentalization of mitochondrial morphology in dendrites of neurons in vivo through spatially precise regulation of mitochondria fission/fusion balance.

3.
Sci Rep ; 12(1): 17074, 2022 10 12.
Article in English | MEDLINE | ID: mdl-36224243

ABSTRACT

Evidence from models and experiments suggests that the networked structure observed in mitochondria emerges at the critical point of a phase transition controlled by fission and fusion rates. If mitochondria are poised at criticality, the relevant network quantities should scale with the system's size. However, whether or not the expected finite-size effects take place has not been demonstrated yet. Here, we first provide a theoretical framework to interpret the scaling behavior of mitochondrial network quantities by analyzing two conceptually different models of mitochondrial dynamics. Then, we perform a finite-size scaling analysis of real mitochondrial networks extracted from microscopy images and obtain scaling exponents comparable with critical exponents from models and theory. Overall, we provide a universal description of the structural phase transition in mammalian mitochondria.


Subject(s)
Finite Element Analysis , Mitochondrial Dynamics , Animals , Mammals , Models, Theoretical , Phase Transition
4.
Dev Cell ; 57(11): 1369-1382.e6, 2022 06 06.
Article in English | MEDLINE | ID: mdl-35609616

ABSTRACT

The endoplasmic reticulum (ER) confronts a challenge to accommodate long, smooth ER tubules into the structural complexity of the axonal compartment. Here, we describe a morphological feature for the axonal ER network in developing neurons we termed the ER ladder. Axonal ER ladders are composed of rungs that wrap tightly around the microtubule bundle and dynamic rails, which slide across microtubules. We found that the ER-shaping protein Reticulon 2 determines the architecture and dynamics of the axonal ER ladder by modulating its interaction with microtubules. Moreover, we show that ER ladder depletion impairs the trafficking of associated vesicular axonal cargoes. Finally, we demonstrate that stromal interaction molecule 1 (Stim1) localizes to ER rungs and translocates to ER-plasma membrane contact sites upon depletion of luminal Ca2+. Our findings uncover fundamental insights into the structural and functional organization of the axonal ER network in developing mammalian neurons.


Subject(s)
Axons , Endoplasmic Reticulum , Animals , Axons/metabolism , Calcium/metabolism , Cytoskeleton/metabolism , Endoplasmic Reticulum/metabolism , Mammals/metabolism , Microtubules/metabolism , Neurons/metabolism
5.
Front Cell Neurosci ; 13: 350, 2019.
Article in English | MEDLINE | ID: mdl-31417367

ABSTRACT

Histopathological studies revealed that progressive neuropathies including Alzheimer, and Prion diseases among others, include accumulations of misfolded proteins intracellularly, extracellularly, or both. Experimental evidence suggests that among the accumulated misfolded proteins, small soluble oligomeric conformers represent the most neurotoxic species. Concomitant phenomena shared by different protein misfolding diseases includes alterations in phosphorylation-based signaling pathways synaptic dysfunction, and axonal pathology, but mechanisms linking these pathogenic features to aggregated neuropathogenic proteins remain unknown. Relevant to this issue, results from recent work revealed inhibition of fast axonal transport (AT) as a novel toxic effect elicited by oligomeric forms of amyloid beta and cellular prion protein PrPC, signature pathological proteins associated with Alzheimer and Prion diseases, respectively. Interestingly, the toxic effect of these oligomers was fully prevented by pharmacological inhibitors of casein kinase 2 (CK2), a remarkable discovery with major implications for the development of pharmacological target-driven therapeutic intervention for Alzheimer and Prion diseases.

6.
Aging Cell ; 17(5): e12812, 2018 Oct.
Article in English | MEDLINE | ID: mdl-30028071

ABSTRACT

Mounting evidence implicates chronic oxidative stress as a critical driver of the aging process. Down syndrome (DS) is characterized by a complex phenotype, including early senescence. DS cells display increased levels of reactive oxygen species (ROS) and mitochondrial structural and metabolic dysfunction, which are counterbalanced by sustained Nrf2-mediated transcription of cellular antioxidant response elements (ARE). Here, we show that caspase 3/PKCδdependent activation of the Nrf2 pathway in DS and Dp16 (a mouse model of DS) cells is necessary to protect against chronic oxidative damage and to preserve cellular functionality. Mitochondria-targeted catalase (mCAT) significantly reduced oxidative stress, restored mitochondrial structure and function, normalized replicative and wound healing capacity, and rendered the Nrf2-mediated antioxidant response dispensable. These results highlight the critical role of Nrf2/ARE in the maintenance of DS cell homeostasis and validate mitochondrial-specific interventions as a key aspect of antioxidant and antiaging therapies.


Subject(s)
Down Syndrome/metabolism , Down Syndrome/pathology , NF-E2-Related Factor 2/metabolism , Oxidative Stress , Animals , Antioxidants/metabolism , Caspase 3/metabolism , Catalase/metabolism , Cell Proliferation , Cell Survival , Cytoprotection , Fibroblasts/metabolism , Fibroblasts/pathology , HEK293 Cells , Humans , Mice, Inbred C57BL , Mitochondria/metabolism , Mitochondria/pathology , Models, Biological , Protein Kinase C-delta/metabolism , Protein Stability , Signal Transduction , Wound Healing
7.
Sci Rep ; 8(1): 363, 2018 01 10.
Article in English | MEDLINE | ID: mdl-29321534

ABSTRACT

Mitochondrial networks exhibit a variety of complex behaviors, including coordinated cell-wide oscillations of energy states as well as a phase transition (depolarization) in response to oxidative stress. Since functional and structural properties are often interwinded, here we characterized the structure of mitochondrial networks in mouse embryonic fibroblasts using network tools and percolation theory. Subsequently we perturbed the system either by promoting the fusion of mitochondrial segments or by inducing mitochondrial fission. Quantitative analysis of mitochondrial clusters revealed that structural parameters of healthy mitochondria laid in between the extremes of highly fragmented and completely fusioned networks. We confirmed our results by contrasting our empirical findings with the predictions of a recently described computational model of mitochondrial network emergence based on fission-fusion kinetics. Altogether these results offer not only an objective methodology to parametrize the complexity of this organelle but also support the idea that mitochondrial networks behave as critical systems and undergo structural phase transitions.


Subject(s)
Mitochondria/metabolism , Mitochondrial Dynamics , Models, Biological , Algorithms , Animals , Fibroblasts , Gene Expression , Genes, Reporter , Mice , Microscopy, Fluorescence
8.
PLoS One ; 12(12): e0188340, 2017.
Article in English | MEDLINE | ID: mdl-29261664

ABSTRACT

Prion diseases include a number of progressive neuropathies involving conformational changes in cellular prion protein (PrPc) that may be fatal sporadic, familial or infectious. Pathological evidence indicated that neurons affected in prion diseases follow a dying-back pattern of degeneration. However, specific cellular processes affected by PrPc that explain such a pattern have not yet been identified. Results from cell biological and pharmacological experiments in isolated squid axoplasm and primary cultured neurons reveal inhibition of fast axonal transport (FAT) as a novel toxic effect elicited by PrPc. Pharmacological, biochemical and cell biological experiments further indicate this toxic effect involves casein kinase 2 (CK2) activation, providing a molecular basis for the toxic effect of PrPc on FAT. CK2 was found to phosphorylate and inhibit light chain subunits of the major motor protein conventional kinesin. Collectively, these findings suggest CK2 as a novel therapeutic target to prevent the gradual loss of neuronal connectivity that characterizes prion diseases.


Subject(s)
Axonal Transport/physiology , Axons/metabolism , Casein Kinase II/metabolism , Prion Proteins/physiology , Animals , Cells, Cultured , Hippocampus/cytology , Hippocampus/metabolism , Kinesins/metabolism , Mice , Mitochondria/metabolism , Phosphorylation
9.
Traffic ; 18(9): 604-621, 2017 09.
Article in English | MEDLINE | ID: mdl-28696565

ABSTRACT

Our understanding of protein and lipid trafficking in eukaryotic cells has been challenged by the finding of different forms of compartmentalization and cargo processing in protozoan parasites. Here, we show that, in the absence of a Golgi compartment in Giardia, proteins destined for secretion are directly sorted and packaged at specialized ER regions enriched in COPII coatomer complexes and ceramide. We also demonstrated that ER-resident proteins are retained at the ER by the action of a KDEL receptor, which, in contrast to other eukaryotic KDEL receptors, showed no interorganellar dynamic but instead acts specifically at the limit of the ER membrane. Our study suggests that the ER-exit sites and the perinuclear ER-membranes are capable of performing protein-sorting functions. In our view, the description presented here suggests that Giardia adaptation represents an extreme example of reductive evolution without loss of function.


Subject(s)
Endoplasmic Reticulum/metabolism , Giardia lamblia/metabolism , Golgi Apparatus/metabolism , COP-Coated Vesicles/metabolism , Protein Transport/physiology , Protozoan Proteins/metabolism , Receptors, Peptide/metabolism
10.
Addict Biol ; 21(2): 326-38, 2016 Mar.
Article in English | MEDLINE | ID: mdl-25431310

ABSTRACT

Behavioral sensitization to cocaine is associated to neuroadaptations that contribute to addiction. Enkephalin is highly expressed in mesocorticolimbic areas associated with cocaine-induced sensitization; however, their influence on cocaine-dependent behavioral and neuronal plasticity has not been explained. In this study, we employed a knockout (KO) model to investigate the contribution of enkephalin in cocaine-induced behavioral sensitization. Wild-type (WT) and proenkephalin KO mice were treated with cocaine once daily for 9 days to induce sensitization. Additionally, to clarify the observations in KO mice, the same procedure was applied in C57BL/6 mice, except that naloxone was administered before each cocaine injection. All animals received a cocaine challenge on days 15 and 21 of the treatment to evaluate the expression of locomotor sensitization. On day 21, microdialysis measures of accumbal extracellular dopamine, Western blotting for GluR1 AMPA receptor (AMPAR), phosphorylated ERK2 (pERK2), CREB (pCREB), TrKB (pTrkB) were performed in brain areas relevant for sensitization from KO and WT and/or naloxone- and vehicle pre-treated animals. We found that KO mice do not develop sensitization to the stimulating properties of cocaine on locomotor activity and on dopamine release in the nucleus accumbens (NAc). Furthermore, pivotal neuroadaptations such as the increase in pTrkB receptor, pERK/CREB and AMPAR related to sensitized responses were absent in the NAc from KO mice. Consistently, full abrogation of cocaine-induced behavioral and neuronal plasticity after naloxone pre-treatment was observed. We show for first time that the proenkephalin system is essential in regulating long-lasting pivotal neuroadaptations in the NAc underlying behavioral sensitization to cocaine.


Subject(s)
Cocaine/pharmacology , Dopamine Uptake Inhibitors/pharmacology , Enkephalins/pharmacology , Neurotransmitter Agents/pharmacology , Analysis of Variance , Animals , Behavior, Animal/drug effects , Dopamine/metabolism , MAP Kinase Signaling System/drug effects , Male , Mice, Inbred C57BL , Mice, Knockout , Naloxone/pharmacology , Narcotic Antagonists/pharmacology , Nucleus Accumbens/drug effects , Phosphorylation/drug effects
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