Biology subjects

Rangaraju, V.

Publications and source records attributed to Rangaraju, V..

7 recordsLinked to original sources

bioRxiv2025

Mitochondria structurally remodel near synapses to fuel the sustained energy demands of plasticity

Brain synapses are hotspots of energy consumption, and the synaptic plasticity underlying learning sharply increases this demand. Because most synapses lie far from their cell body, their energy demands must be met locally. A dendritic mitochondrion can extend ~30 m and span many spines, yet ramps up ATP synthesis only near spines undergoing plasticity, providing immediate and sustained energy. Whether structural changes in these juxtaspinal mitochondrial regions (2 m segments at spine bases) support this response is unknown; resolving their energy-producing architecture poses substantial challenges. Compared with the lamellar cristae of axonal mitochondria, dendritic mitochondrial cristae form complex, tubular, and interconnected networks that require multiple high-resolution tomograms to characterize. We therefore developed a correlative light and electron microscopy (CLEM) pipeline with a custom deep-learning model that simultaneously segments five distinct 3D suborganellar structures from limited training data, enabling quantitative analysis across 42 tomograms at 2 nm pixel resolution. We find that juxtaspinal mitochondrial regions are larger than non-juxtaspinal regions within the same mitochondrion and enlarge further during synaptic plasticity. Juxtaspinal mitochondrial ATP synthesis and spine ATP levels rise in parallel; during single-spine plasticity, ATP rises within minutes, before detectable enlargement, indicating that juxtaspinal mitochondrial enlargement sustains rather than initiates the local energy response. The enlarged regions exhibit increased cristae membrane surface area, crista junction number, and membrane area with high curvature, together with increased MIC60 and ATP synthase copies, endoplasmic reticulum contacts, and more proximal ribosomes. Matrix-targeted PKA inhibition reduces MIC60 phosphorylation and impairs juxtaspinal mitochondrial enlargement, plasticity-associated ATP increase, and single-spine plasticity. Together, our findings support a model in which spine-specific remodeling within an individual mitochondrion boosts local metabolic capacity to sustain ATP synthesis and fuel synaptic plasticity.

neuroscience↗

bioRxiv2025

Synapses drive local mitochondrial ATP synthesis to fuel plasticity

Our brain constantly forms new memories and stabilizes existing memories. To achieve such cognitive flexibility, the brain is wired by plastic synapses that are hotspots of energy consumption. Supplying energy to distant synapses is challenging as they are distributed throughout dendrites and axons, spanning hundreds of microns from their cell body. Synapses, therefore, require an instant and local energy supply provided by mitochondria stabilized near dendritic spines. However, the mechanisms by which synapses communicate their energy demands to locally stable mitochondria to drive local energy production and sustain synaptic plasticity is unknown. Using highly sensitive spine- and mitochondrial ATP reporters and two-photon glutamate uncaging to stimulate individual spines, we find that synaptic plasticity input drives instant and sustained increase in spine ATP levels, provided by local ATP synthesis in [~]10-20 m spatially confined compartments within mitochondria. This spatially localized mitochondrial ATP generation is driven by a spatially localized mitochondrial calcium influx independent of the endoplasmic reticulum. Notably, the initial spine ATP increase, supported by local mitochondrial ATP synthesis, is independent of CaMKII and the energy demands of spine structural plasticity. Without local calcium signaling and mitochondrial stabilization, synapses do not meet their instant and sustained energy needs, resulting in synaptic plasticity defects, as observed in neurological disorders.

neuroscience↗

bioRxiv2025

An expanded palette of bright and photostable organellar Ca2+ sensors

The use of fluorescent sensors for functional imaging has revolutionized the study of organellar Ca2+ signaling. However, understanding the dynamic interplay between intracellular Ca2+ sinks and sources has been hindered by the lack of bright, photostable, and multiplexed measurements in different organelles, limiting our ability to define how Ca2+ shapes cell physiology across fields of biology. Here we introduce a new toolkit of chemigenetic organellar Ca2+ indicators whose color is tunable by reconstituting their fluorescence with different exogenous rhodamine dye-ligands, which significantly expand the capacity for multiplexing organellar Ca2+ measurements. These sensors, which we named ER-HaloCaMP and Mito-HaloCaMP, are optimized to report Ca2+ dynamics in the endoplasmic reticulum (ER) and mitochondria of mammalian cells and neurons, and show significantly improved brightness, photostability and responsiveness when compared to current best-in-class alternatives. Using either red or far-red dye-ligands, both ER-HaloCaMP and Mito-HaloCaMP enable visualizing ER and mitochondrial Ca2+ dynamics in neuronal axons, a subcellular location that only contains a few ER tubules and small mitochondria, structural limitations that have impaired measurements with previous red sensors. To show the expanded multiplexing capacities of our toolkit, we measured interorganellar Ca2+ fluxes simultaneously in three different subcellular compartments in live cells, revealing that the amplitude of ER Ca2+ release controls the efficacy of ER-mitochondria Ca2+ coupling in a cooperative manner. Organellar HaloCaMPs enable also measuring Ca2+ dynamics in intact brain tissue from flies and rodents, demonstrating their versatility across biological models. Our new toolkit provides an expanded palette of bright, photostable and responsive organellar Ca2+ sensors, which will facilitate future studies of intracellular Ca2+ signaling across fields of biology in health and disease.

cell biology↗

bioRxiv2024

Periodic ER-plasma membrane junctions support long-range Ca2+ signal integration in dendrites

Neuronal dendrites must relay synaptic inputs over long distances, but the mechanisms by which activity-evoked intracellular signals propagate over macroscopic distances remain unclear. Here, we discovered a system of periodically arranged endoplasmic reticulum-plasma membrane (ER-PM) junctions tiling the plasma membrane of dendrites at [~]1 m intervals, interlinked by a meshwork of ER tubules patterned in a ladder-like array. Populated with Junctophilin-linked plasma membrane voltage-gated Ca2+ channels and ER Ca2+-release channels (ryanodine receptors), ER-PM junctions are hubs for ER-PM crosstalk, fine-tuning of Ca2+ homeostasis, and local activation of the Ca2+/calmodulin-dependent protein kinase II. Local spine stimulation activates the Ca2+ modulatory machinery facilitating voltage-independent signal transmission and ryanodine receptor-dependent Ca2+ release at ER-PM junctions over 20 m away. Thus, interconnected ER-PM junctions support signal propagation and Ca2+ release from the spine-adjacent ER. The capacity of this subcellular architecture to modify both local and distant membrane-proximal biochemistry potentially contributes to dendritic computations. HighlightsO_LIPeriodic ER-PM junctions tile neuronal dendritic plasma membrane in rodent and fly. C_LIO_LIER-PM junctions are populated by ER tethering and Ca2+ release and influx machinery. C_LIO_LIER-PM junctions act as sites for local activation of CaMKII. C_LIO_LILocal spine activation drives Ca2+ release from RyRs at ER-PM junctions over 20 m. C_LI

neuroscience↗

bioRxiv2024

Mitochondrial Ca2+ efflux controls neuronal metabolism and long-term memory across species

While impairing neuronal metabolism limits brain performance, it remains poorly understood whether enhancing metabolism in neurons, in contrast, could boost brain function. We find that reducing the expression of the mitochondrial H+/Ca2+ exchanger Letm1 results in increased Ca2+ retention in the mitochondrial matrix of firing neurons, which overactivates neuronal metabolism in flies and rodents. We find that upscaled metabolic states in active neurons of central memory circuits of flies and mice enable storing long-term memories in training paradigms in which wild-type counterparts of both species fail to remember. Our findings unveil an evolutionarily conserved mechanism that controls mitochondrial metabolism in active neurons and prove its crucial role in governing higher brain functions, such as long-term memory formation. Highlights- Letm1 controls activity-driven mitochondrial Ca2+ efflux in neurons - Increased mitochondrial Ca2+ retention during activity overactivates neuronal metabolism - Activity-driven upscaling of neuronal metabolism facilitates long-term olfactory memory in flies and mice

neuroscience↗

bioRxiv2023

VAP spatially stabilizes dendritic mitochondria to locally fuel synaptic plasticity

Synapses are pivotal sites of memory formation and undergo plasticity in response to external inputs. Consequently, synapses are hotspots of energy consumption and are susceptible to dysfunction when their energy supplies are perturbed. Mitochondria are stabilized near synapses via cytoskeletal tethering and serve as local energy supplies to fuel synaptic plasticity. However, the mechanisms that tether and stabilize neuronal mitochondria for long durations and determine the spatial dendritic segment supported during synaptic plasticity are unknown. We identified a list of novel mitochondrial-cytoskeletal interactors in neurons using APEX-based proximity labeling. We narrowed down the protein candidates that exclusively tether mitochondria to actin near postsynaptic spines using high-resolution Airyscan confocal imaging. We find that VAP, the vesicle-associated membrane protein-associated protein implicated in Amyotrophic Lateral Sclerosis and interacts with the endoplasmic reticulum, stabilizes mitochondria via actin near the spines. To test if the VAP-dependent stable mitochondrial compartments can locally support synaptic plasticity, we investigated individual spines stimulated by two-photon glutamate uncaging for spine plasticity induction and their adjacent spines. We find that, along with actin, VAP functions as a spatial stabilizer of mitochondrial compartments to sustain synaptic plasticity for up to ~60 min and as a spatial ruler that determines the ~30 m length of the dendritic segment supporting synaptic plasticity.

neuroscience↗

bioRxiv2022

NMNAT2 is the major NAD+ provider for vesicular glycolysis generating on-board energy for fast axonal transport cargos

BackgroundBioenergetic maladaptations and axonopathy are often found in the early stages of neurodegeneration. Nicotinamide adenine dinucleotide (NAD), an essential cofactor for energy metabolism, is mainly synthesized by Nicotinamide mononucleotide adenylyl transferase 2 (NMNAT2) in CNS neurons. NMNAT2 mRNA levels are reduced in the brains of Alzheimers, Parkinsons and Huntingtons disease. Here we addressed whether NMNAT2 is required for axonal health of cortical glutamatergic neurons, whose far-projecting axons are vulnerable to neurodegenerative conditions. We also tested if NMNAT2 maintains axonal health by ensuring proper axonal ATP levels for axonal transport, a critical function of axons. MethodsWe generated mouse and cultured neuron models to determine the impact of NMNAT2 loss from cortical glutamatergic neurons on axonal transport, energetic metabolism, and morphological integrity. In addition, we determined if exogenous NAD supplementation or inhibiting NAD hydrolase sterile alpha and TIR motif-containing protein 1 (SARM1) prevented axonal deficits caused by NMNAT2 loss. Our study used a combination of genetic, molecular biology, immunohistochemistry, biochemistry, fluorescent time-lapse imaging, live imaging with optical sensors, and anti-sense oligos application. ResultsWe provide in vivo evidence that NMNAT2 in cortical glutamatergic neurons is required for axonal survival. Using in vivo and in vitro studies we demonstrate that NMNAT2 protects axons by ensuring the proper NAD-redox potential in distal axons of cortical neurons to support glycolysis on vesicular cargos, thus ensuring "onboard" ATP production fueling axonal transport. Exogenous NAD+ supplementation to NMNAT2 KO cortical neurons restores glycolysis and resumes fast axonal transport. Finally, we demonstrate both in vitro and in vivo that reducing the activity of SARM1, an NAD degradation enzyme, can reduce axonal transport deficits and suppress axon degeneration in NMNAT2 KO neurons. ConclusionNMNAT2 ensures axonal health by maintaining NAD redox potential in distal axons to ensure efficient vesicular glycolysis required for fast axonal transport.

neuroscience↗