Abstract
The brain's remarkable properties arise from the collective activity of millions of neurons. Widespread application of dimensionality reduction to multi-neuron recordings implies that neural dynamics can be approximated by low-dimensional "latent" signals reflecting neural computations. However, can such low-dimensional representations truly explain the vast range of brain activity, and if not, what is the appropriate resolution and scale of recording to capture them? Imaging neural activity at cellular resolution and near-simultaneously across the mouse cortex, we demonstrate an unbounded scaling of dimensionality with neuron number in populations up to 1 million neurons. Although half of the neural variance is contained within sixteen dimensions correlated with behavior, our discovered scaling of dimensionality corresponds to an ever-increasing number of neuronal ensembles without immediate behavioral or sensory correlates. The activity patterns underlying these higher dimensions are fine grained and cortex wide, highlighting that large-scale, cellular-resolution recording is required to uncover the full substrates of neuronal computations.
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💻 Code & Software
Analysis of the dimensionality of neuronal population dynamics as a function of neuron number
Scripts for controlling treadmill and stimuli delivery.
Pipeline based on NoRMCorre and CaImAn for segmenting large FOV, volumetric Light Bead Microscopy data.
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📋 Methods
RESOURCE AVAILABILITY
Lead contact Further information and requests for resources should be directed to the lead contact, Alipasha Vaziri ( vaziri@rockefeller.edu ).
Materials availability
This study did not generate new unique reagents.
Data and code availability
The codebase used to perform these analyses is available at https://github.com/vazirilab/scaling_analysis . This codebase and example processed datasets are deposited at https://doi.org/10.5281/zenodo.10403684 . All other information required to reanalyze the data reported in this paper is available from the lead contact upon reasonable request.
EXPERIMENTAL MODEL AND SUBJECT DETAILS Animal subjects
Transgenic mice expressing either GCaMP6s or GCaMP6f in glutamatergic neurons (Ai162D or Ai148D x Vglut1-IRES2-Cre-D, Jackson Labs stock numbers 031562, 030328, and 037512 respectively 60 ) and wild-type (C57BL/6J) mice were bred in house. Male and female mice were studied, and any effect of sex was not investigated. All mice were 30–95 days of age at the time of first procedure and were 47–170 days old during imaging experiments. Mice were allowed food and water ad libitum .
METHODS DETAILS Surgical procedures
All surgical and experimental procedures were approved by the Institutional Animal Care and Use Committee of The Rockefeller University. Cranial window implantation was performed as previously described. 24 During implantation, mice were anesthetized with isoflurane (1–1.5% maintenance at a flow rate of 0.7–0.9 L/min) and placed in a stereotaxic frame (RWD Life Science). The scalp and underlying connective tissue were removed and cleared from the skull. A custom stainless-steel head bar was fixed behind the occipital bone with cyanoacrylate glue (Loctite) and covered with black dental cement (Ortho-Jet, Lang Dental). A circular 8 mm diameter dual-hemisphere craniotomy was performed, leaving the dura intact. An approximately 1 mm segment of skull at the furthest posterior bound of the diameter of the craniotomy was left intact to avoid the junction of the sagittal and transverse sinus vessels while drilling. The cranial window was formed by implanting a circular 8 mm glass coverslip with 1 mm of the bottom removed (#1 thickness, Warner instruments) before sealing with tissue adhesive (Vetbond). The remaining exposed skull around the cranial window was covered with cyanoacrylate glue and then dental cement. Post-operative care included 2 days of subcutaneous delivery of dexamethasone (2 mg/kg), 3 days of subcutaneous delivery of meloxicam (0.125 mg/kg), and antibiotic-containing feed (LabDiet no. 58T7). After surgery, animals were returned to their home cages and were given at least one week to recover before imaging experiments. Mice with damaged dura, significant regrowth or otherwise unclear windows were euthanized and not used for imaging experiments. In the case of static GFP imaging experiments in Figures S3D and S3F - G , brain-wide expression of genetically encoded green fluorescent protein (GFP) was achieved by retro-orbital injection of 100 μL of AAV-PHP.eB CAG-GFP (2.7 * 10^12 vg/mL) into wild-type (C57BL/6J) mice. Animals were injected one week after implantation of the cranial window.
Show full methods section
RESOURCE AVAILABILITY
Lead contact Further information and requests for resources should be directed to the lead contact, Alipasha Vaziri ( vaziri@rockefeller.edu ).
Materials availability
This study did not generate new unique reagents.
Data and code availability
The codebase used to perform these analyses is available at https://github.com/vazirilab/scaling_analysis . This codebase and example processed datasets are deposited at https://doi.org/10.5281/zenodo.10403684 . All other information required to reanalyze the data reported in this paper is available from the lead contact upon reasonable request.
EXPERIMENTAL MODEL AND SUBJECT DETAILS Animal subjects
Transgenic mice expressing either GCaMP6s or GCaMP6f in glutamatergic neurons (Ai162D or Ai148D x Vglut1-IRES2-Cre-D, Jackson Labs stock numbers 031562, 030328, and 037512 respectively 60 ) and wild-type (C57BL/6J) mice were bred in house. Male and female mice were studied, and any effect of sex was not investigated. All mice were 30–95 days of age at the time of first procedure and were 47–170 days old during imaging experiments. Mice were allowed food and water ad libitum .
METHODS DETAILS Surgical procedures
All surgical and experimental procedures were approved by the Institutional Animal Care and Use Committee of The Rockefeller University. Cranial window implantation was performed as previously described. 24 During implantation, mice were anesthetized with isoflurane (1–1.5% maintenance at a flow rate of 0.7–0.9 L/min) and placed in a stereotaxic frame (RWD Life Science). The scalp and underlying connective tissue were removed and cleared from the skull. A custom stainless-steel head bar was fixed behind the occipital bone with cyanoacrylate glue (Loctite) and covered with black dental cement (Ortho-Jet, Lang Dental). A circular 8 mm diameter dual-hemisphere craniotomy was performed, leaving the dura intact. An approximately 1 mm segment of skull at the furthest posterior bound of the diameter of the craniotomy was left intact to avoid the junction of the sagittal and transverse sinus vessels while drilling. The cranial window was formed by implanting a circular 8 mm glass coverslip with 1 mm of the bottom removed (#1 thickness, Warner instruments) before sealing with tissue adhesive (Vetbond). The remaining exposed skull around the cranial window was covered with cyanoacrylate glue and then dental cement. Post-operative care included 2 days of subcutaneous delivery of dexamethasone (2 mg/kg), 3 days of subcutaneous delivery of meloxicam (0.125 mg/kg), and antibiotic-containing feed (LabDiet no. 58T7). After surgery, animals were returned to their home cages and were given at least one week to recover before imaging experiments. Mice with damaged dura, significant regrowth or otherwise unclear windows were euthanized and not used for imaging experiments. In the case of static GFP imaging experiments in Figures S3D and S3F - G , brain-wide expression of genetically encoded green fluorescent protein (GFP) was achieved by retro-orbital injection of 100 μL of AAV-PHP.eB CAG-GFP (2.7 * 10^12 vg/mL) into wild-type (C57BL/6J) mice. Animals were injected one week after implantation of the cranial window.
Imaging parameters
Data acquisition was performed utilizing LBM as previously described, 24 utilizing a custom multiplexing module interfaced with a commercial mesoscope (Thorlabs, Multiphoton Mesoscope). n=11 total mice, male or female, were imaged across n=25 sessions for a duration of 30 minutes (n=2), 60 minutes (n=13), 90 minutes (n=6), or 120 minutes (n=4). Three FOV types were recorded: 1.2×1.2×0.5 mm 3 , 2 μm lateral pixel spacing, at 10 Hz volume rate (yielding 6,519 – 21,087 neurons); 3×5×0.5 mm 3 , 5 μm spacing, at 4.7 Hz (134,628 – 315,363 neurons); and 5.4×6×0.5 mm 3 , 5 μm spacing, at 2.2 Hz (523,139 – 1,136,223 neurons). Imaging power was restricted to
📊 Figures
Figure 1.
Light Beads Microscopy (LBM) enables large-scale, volumetric recording of neuronal activity across cortex at cellular resolution.
A. A schematic representation of two possible scenarios: bounded versus unbounded scaling of the measured neuronal dimensionality as a function of number of recorded neurons. B. A schematic of the LBM...
Figure 2.
Shared variance component analysis (SVCA) reveals unbounded scaling of reliable neural dimensionality.
A. Schematics of SVCA: (i): SVCA splits the neurons into two sets (green and purple). (ii): SVCs are the maximally covarying projections of each neural set. (iii): The reliability of each neural SVC i...
Figure 3.
Behavior-related activity is encoded in a low-dimensional subspace of reliable neuronal dynamics.
A. Example behavior videography image, with a box denoting the area in which facial motion energy was monitored. B. The top three behavior PCs for an example mouse. C. Schematics of the prediction of ...
Figure 4.
Lack of correlation of high-dimensional neural SVCs with comprehensive behavioral monitoring and sensory-related activity.
A. Example images from simultaneous, all-around behavior monitoring. (i) The left side of mouse. (ii) Left: the right side of the face and pupil. Right: an inset depicting the pupil. (iii) The body of...
Figure 5.
Latent neural SVC dynamics represent a continuum of timescales.
A. Example autocorrelation curves for four neural SVCs from the same recording as Figure 2B - C . B. Characteristic dominant timescales within each SVC. The dominant autocorrelation timescale u03c4, c...
Figure 6.
Lower and higher neural SVCs form distinct, spatially organized neuronal assemblies.
A. Lower and higher SVCs exhibit distinct cortex-wide neuronal distribution profiles. The lateral spatial distribution of the neurons participating in four example SVCs in a single hemisphere recordin...
Figure images are served from the NIH/NLM PubMed Central Open Access Subset or Europe PMC; copyright remains with the publishers and authors.
💬 Discussion
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