MCU-enriched dendritic mitochondria regulate plasticity in distinct hippocampal circuits.

Mitochondria are dynamic organelles that are morphologically and functionally diverse across cell types and subcellular compartments in order to meet unique energy demands. Mitochondrial dysfunction has been implicated in a wide variety of neurological disorders, including psychiatric disorders like schizophrenia and bipolar disorder. Despite it being well known that mitochondria are essential for synaptic transmission and synaptic plasticity, the mechanisms regulating mitochondria in support of normal synapse function are incompletely understood. The mitochondrial calcium uniporter (MCU) regulates calcium entry into the mitochondria, which in turn regulates the bioenergetics and distribution of mitochondria to active synapses. Evidence suggests that calcium influx via MCU couples neuronal activity to mitochondrial metabolism and ATP production, which would allow neurons to rapidly adapt to changing energy demands. Intriguingly, MCU is uniquely enriched in hippocampal CA2 distal dendrites relative to neighboring hippocampal CA1 or CA3 distal dendrites, however, the functional significance of this enrichment is not clear. Synapses from the entorhinal cortex layer II (ECII) onto CA2 distal dendrites readily express long term potentiation (LTP), unlike the LTP- resistant synapses from CA3 onto CA2 proximal dendrites, but the mechanisms underlying these different plasticity profiles are unknown. We hypothesized that enrichment of MCU near ECII-CA2 synapses promotes LTP in an otherwise plasticity-restricted cell type. Using a CA2-specific MCU knockout (cKO) mouse, we found that MCU is required for LTP at distal dendrite synapses but does not affect the lack of LTP at proximal dendrite synapses. Loss of LTP at ECII-CA2 synapses correlated with a trend for decreased spine density in CA2 distal dendrites of cKO mice compared to control (CTL) mice, which was predominantly seen in immature spines. Moreover, mitochondria were significantly smaller and more numerous across all dendritic layers of CA2 in cKO mice compared to CTL mice, suggesting an overall increase in mitochondrial fragmentation. Fragmented mitochondria might have functional changes, such as altered ATP production, that might explain a deficit in synaptic plasticity. Collectively, our data reveal that MCU regulates layer-specific forms of plasticity in CA2 dendrites, potentially by maintaining proper mitochondria morphology and distribution within dendrites. Differences in MCU expression across different cell types and circuits might be a general mechanism to tune the sensitivity of mitochondria to cytoplasmic calcium levels to power synaptic plasticity. MAIN TAKE HOME POINTS: The mitochondrial calcium uniporter (MCU) regulates plasticity selectively at synapses in CA2 distal dendrites.The MCU-cKO induced LTP deficit correlates with a trending reduction in spine density in CA2 distal dendrites.Loss of MCU in CA2 results in ultrastructural changes in dendritic mitochondria that suggest an increase in mitochondrial fragmentation. These ultrastructural changes could result in functional consequences, such as decreased ATP production, that could underlie the plasticity deficit.Dendritic mitochondrial fragmentation in MCU cKO occurred throughout the dendritic laminae, suggesting that MCU is dispensable for establishing layer-specific mitochondrial structural diversity.

Layer-specific mitochondrial diversity across hippocampal CA2 dendrites.

CA2 is an understudied subregion of the hippocampus that is critical for social memory. Previous studies identified multiple components of the mitochondrial calcium uniporter (MCU) complex as selectively enriched in CA2. The MCU complex regulates calcium entry into mitochondria, which in turn regulates mitochondrial transport and localization to active synapses. We found that MCU is strikingly enriched in CA2 distal apical dendrites, precisely where CA2 neurons receive entorhinal cortical input carrying social information. Furthermore, MCU-enriched mitochondria in CA2 distal dendrites are larger compared to mitochondria in CA2 proximal apical dendrites and neighboring CA1 apical dendrites, which was confirmed in CA2 with genetically labeled mitochondria and electron microscopy. MCU overexpression in neighboring CA1 led to a preferential localization of MCU in the proximal dendrites of CA1 compared to the distal dendrites, an effect not seen in CA2. Our findings demonstrate that mitochondria are molecularly and structurally diverse across hippocampal cell types and circuits, and suggest that MCU can be differentially localized within dendrites, possibly to meet local energy demands.

LEC to CA2: A circuit to remember.

In this issue of Neuron, Lopez-Rojas et al. (2022) uncover a cortical circuit conveying social information to CA2, a region essential for social memory. Their findings suggest CA2 neurons integrate information from other extrahippocampal circuits to locally compute social novelty.

Protein-retention expansion microscopy for visualizing subcellular organelles in fixed brain tissue.

BACKGROUND: Protein expansion microscopy (proExM) is a powerful technique that crosslinks proteins to a swellable hydrogel to physically expand and optically clear biological samples. The resulting increased resolution (~70 nm) and physical separation of labeled proteins make it an attractive tool for studying the localization of subcellular organelles in densely packed tissues, such as the brain. However, the digestion and expansion process greatly reduce fluorescence signals making it necessary to optimize ExM conditions per sample for specific end goals. NEW METHOD: Here we compare the staining and digestion conditions of existing proExM workflows to identify the optimal protocol for visualizing subcellular organelles (mitochondria and the Golgi apparatus) within reporter-labeled neurons in fixed mouse brain tissue. RESULTS: We found that immunostaining before proExM and using a proteinase K based digestion for 8 h consistently resulted in robust fluorescence retention for immunolabeled subcellular organelles and genetically-encoded reporters. COMPARISON WITH EXISTING METHODS: With these methods, we more accurately quantified mitochondria size and number and better visualized Golgi ultrastructure in individual CA2 neurons in the mouse hippocampus. CONCLUSIONS: This organelle optimized proExM protocol will be broadly useful for investigators interested in visualizing the spatial distribution of immunolabeled subcellular organelles in various reporter mouse lines, reducing effort, time and resources on the optimization process.

Input clustering and the microscale structure of local circuits.

The recent development of powerful tools for high-throughput mapping of synaptic networks promises major advances in understanding brain function. One open question is how circuits integrate and store information. Competing models based on random vs. structured connectivity make distinct predictions regarding the dendritic addressing of synaptic inputs. In this article we review recent experimental tests of one of these models, the input clustering hypothesis. Across circuits, brain regions and species, there is growing evidence of a link between synaptic co-activation and dendritic location, although this finding is not universal. The functional implications of input clustering and future challenges are discussed.