Title: Primary cortical neurons from SLC13A5 nTg mice have normal morphology but reduced synapse formation and increased spontaneous activity.
Legend: (A) Morphologic assessment of cultured neurons at 15 days in vitro (DIV). Phalloidin staining (left) and unbiased computer-driven reconstruction (right) are shown along with quantification from n=3 embryos per genotype. Data are mean+SEM (Sholl analysis) or SD (spine density and volume) with each data point representing one embryo. (B) Immunostaining of cultured neurons for pre-synaptic marker Syn1 and post-synaptic marker Psd-95 at 15 DIV. Puncta were fit with 2 μm spots and normalized to neuron volume, and spots co-localized if they were within 1 μm of each other. Data are shown with each data point representing one embryo, n=4 WT and n=3 SLC13A5 nTg. Statistical testing was conducted via unpaired t-test with the t statistics and P-values as follows: Syn-1 (3.817; 0.0124), Psd-95 (6.203; 0.0016), co-localized (5.931; 0.0019). (C) Multi-electrode array spontaneous activity. Left: histogram showing the number of active electrodes per network expressed as the relative frequency in per cent. The vertical line demarks eight active electrodes, which is the minimum value required to be considered a mature network. Statistical testing was conducted via the Mann–Whitney test comparing frequency distributions at each DIV with Mann–Whitney U-values and P-values as follows: DIV7 (108.5; 0.0001), DIV14 (155; 0.0061), DIV 21 (138.5; 0.0020), DIV 28 (110; 0.0002). Right: spontaneous activity measured by mean firing rate, burst frequency, network burst frequency and synchronicity index. Each data point is an independent network of cultured neurons and exhibits at least 8 of 16 active electrodes. Statistical testing was conducted via mixed effects analysis (DIV×genotype) with Sidak’s multiple comparison test. DIV7 data were excluded from analysis due to the lack of WT values. Test details are as follows, listing the F statistics for the genotype factor and adjusted P-values from multiple comparison testing: mean firing rate (8.956; DIV14, 0.0001; DIV 21, 0.0303), burst frequency (5.505; DIV14, 0.0013; DIV 21, 0.0178), network burst frequency (3.789; DIV14, 0.0013; DIV28, 0.0346) and network synchronicity (6.745; DIV14, 0.0160). Data are from three WT and seven SLC13A5 nTg embryos.
Citation: Michael J. Rigby, Nicola Salvatore Orefice, Alexis J. Lawton, Min Ma, Samantha L. Shapiro, Sue Y. Yi, Inca A. Dieterich, Alyssa Frelka, Hannah N. Miles, Robert A. Pearce, John Paul J. Yu, Lingjun Li, John M. Denu, Luigi Puglielli. SLC13A5/sodium-citrate co-transporter overexpression causes disrupted white matter integrity and an autistic-like phenotype. Brain Communications, Volume 4, Issue 1, 2022, fcac002, https://doi.org/10.1093/braincomms/fcac002
Abstract: Endoplasmic reticulum-based Nɛ-lysine acetylation serves as an important protein quality control system for the secretory pathway. Dysfunctional endoplasmic reticulum-based acetylation, as caused by overexpression of the acetyl coenzyme A transporter AT-1 in the mouse, results in altered glycoprotein flux through the secretory pathway and an autistic-like phenotype. AT-1 works in concert with SLC25A1, the citrate/malate antiporter in the mitochondria, SLC13A5, the plasma membrane sodium/citrate symporter, and ATP citrate lyase, the cytosolic enzyme that converts citrate into acetyl coenzyme A. Here, we report that mice with neuron-specific overexpression of SLC13A5 exhibit autistic-like behaviours with a jumping stereotypy. The mice displayed disrupted white matter integrity and altered synaptic structure and function. Analysis of both the proteome and acetyl-proteome revealed unique adaptations in the hippocampus and cortex, highlighting a metabolic response that likely plays an important role in the SLC13A5 neuron transgenic phenotype. Overall, our results support a mechanistic link between aberrant intracellular citrate/acetyl coenzyme A flux and the development of an autistic-like phenotype.
About the Lab: The Puglielli Lab’s research interests focus on molecular mechanisms of neurodevelopment and neurodegeneration. The laboratory employs a combination of biochemical, cellular, molecular, and genetic approaches in in vitro, ex vivo and in vivo models.
Investigator: Luigi Puglielli, MD, PhD