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Robust activity-dependent mitochondrial calcium dynamics at the AIS is dispensable for action potential generation.

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The authors' code

MATLAB · 215 lines · 9.9 KB · no license

  1. function options = NoRMCorreSetParms(varargin)
  3. % Struct for setting the NoRMCorre algorithm parameters. Any parameter that is
  4. % not set gets a default value
  6. % Author: Eftychios A. Pnevmatikakis
  7. % Simons Foundation, 2016
  9. Names = [
  10. % dataset info
  11. 'd1 ' % number of rows
  12. 'd2 ' % number of cols
  13. 'd3 ' % number of planes (for 3d imaging, default: 1)
  14. % patches
  15. 'grid_size ' % size of non-overlapping regions (default: [d1,d2,d3])
  16. 'overlap_pre ' % size of overlapping region (default: [32,32,16])
  17. 'min_patch_size ' % minimum size of patch (default: [32,32,16])
  18. 'us_fac ' % upsampling factor for subpixel registration (default: 20)
  19. 'mot_uf ' % degree of patches upsampling (default: [4,4,1])
  20. 'max_dev ' % maximum deviation of patch shift from rigid shift (default: [3,3,1])
  21. 'overlap_post ' % size of overlapping region after upsampling (default: [32,32,16])
  22. 'max_shift ' % maximum rigid shift in each direction (default: [15,15,5])
  23. 'phase_flag ' % flag for using phase correlation (default: false)
  24. 'shifts_method ' % method to apply shifts ('FFT','cubic','linear')
  25. % template updating
  26. 'upd_template ' % flag for online template updating (default: true)
  27. 'init_batch ' % length of initial batch (default: 100)
  28. 'bin_width ' % width of each bin (default: 10)
  29. 'buffer_width ' % number of local means to keep in memory (default: 50)
  30. 'method ' % method for averaging the template (default: {'median';'mean})
  31. 'iter ' % number of data passes (default: 1)
  32. 'boundary ' % method of boundary treatment 'NaN','copy','zero','template' (default: 'copy')
  33. % misc
  34. 'add_value ' % add dc value to data (default: 0)
  35. 'use_parallel ' % for each frame, update patches in parallel (default: false)
  36. 'memmap ' % flag for saving memory mapped motion corrected file (default: false)
  37. 'mem_filename ' % name for memory mapped file (default: 'motion_corrected.mat')
  38. 'mem_batch_size ' % batch size during memory mapping for speed (default: 5000)
  39. % plotting
  40. 'plot_flag ' % flag for plotting results in real time (default: false)
  41. 'make_avi ' % flag for making movie (default: false)
  42. 'name ' % name for movie (default: 'motion_corrected.avi')
  43. 'fr ' % frame rate for movie (default: 30)
  44. % output type
  45. 'output_type ' % 'mat' (load in memory), 'memmap', 'tiff', 'hdf5', 'bin' (default:mat)
  46. 'h5_groupname ' % name for hdf5 dataset (default: 'mov')
  47. 'h5_filename ' % name for hdf5 saved file (default: 'motion_corrected.h5')
  48. 'tiff_filename ' % name for saved tiff stack (default: 'motion_corrected.tif')
  49. % use windowing
  50. 'use_windowing ' % flag for windowing data before fft (default: false)
  51. 'window_length ' % length of window on each side of the signal as a fraction of signal length
  52. % total length = length(signal)(1 + 2*window_length). (default: 0.5)
  53. % bitsize for reading .raw files
  54. 'bitsize ' % (default: 2 (uint16). other choices 1 (uint8), 4 (single), 8 (double))
  55. % offset from bidirectional sampling
  56. 'correct_bidir ' % check for offset due to bidirectional scanning (default: true)
  57. 'nFrames ' % number of frames to average (default: 50)
  58. 'bidir_us ' % upsampling factor for bidirectional sampling (default: 10)
  59. 'col_shift ' % known bi-directional offset provided by the user (default: [])
  60. ];
  62. [m,n] = size(Names);
  63. names = lower(Names);
  65. % Combine all leading options structures o1, o2, ... in l1Set(o1,o2,...).
  66. options = [];
  67. for j = 1:m
  68. eval(['options.' Names(j,:) '= [];']);
  69. end
  70. i = 1;
  71. while i <= nargin
  72. arg = varargin{i};
  73. if ischar(arg), break; end
  74. if ~isempty(arg) % [] is a valid options argument
  75. if ~isa(arg,'struct')
  76. error(sprintf(['Expected argument %d to be a string parameter name ' ...
  77. 'or an options structure\ncreated with OPTIMSET.'], i));
  78. end
  79. for j = 1:m
  80. if any(strcmp(fieldnames(arg),deblank(Names(j,:))))
  81. eval(['val = arg.' Names(j,:) ';']);
  82. else
  83. val = [];
  84. end
  85. if ~isempty(val)
  86. eval(['options.' Names(j,:) '= val;']);
  87. end
  88. end
  89. end
  90. i = i + 1;
  91. end
  93. % A finite state machine to parse name-value pairs.
  94. if rem(nargin-i+1,2) ~= 0
  95. error('Arguments must occur in name-value pairs.');
  96. end
  97. expectval = 0; % start expecting a name, not a value
  98. while i <= nargin
  99. arg = varargin{i};
  101. if ~expectval
  102. if ~ischar(arg)
  103. error(sprintf('Expected argument %d to be a string parameter name.', i));
  104. end
  106. lowArg = lower(arg);
  107. j = strmatch(lowArg,names);
  108. if isempty(j) % if no matches
  109. error(sprintf('Unrecognized parameter name ''%s''.', arg));
  110. elseif length(j) > 1 % if more than one match
  111. % Check for any exact matches (in case any names are subsets of others)
  112. k = strmatch(lowArg,names,'exact');
  113. if length(k) == 1
  114. j = k;
  115. else
  116. msg = sprintf('Ambiguous parameter name ''%s'' ', arg);
  117. msg = [msg '(' deblank(Names(j(1),:))];
  118. for k = j(2:length(j))'
  119. msg = [msg ', ' deblank(Names(k,:))];
  120. end
  121. msg = sprintf('%s).', msg);
  122. error(msg);
  123. end
  124. end
  125. expectval = 1; % we expect a value next
  127. else
  128. eval(['options.' Names(j,:) '= arg;']);
  129. expectval = 0;
  131. end
  132. i = i + 1;
  133. end
  135. if expectval
  136. error(sprintf('Expected value for parameter ''%s''.', arg));
  137. end
  139. Values = [
  140. % dataset info
  141. {[]}
  142. {[]}
  143. {1}
  144. % patches
  145. {[]} % size of non-overlapping regions (default: [d1,d2,d3])
  146. {[32,32,16]} % size of overlapping region (default: [32,32,16])
  147. {[32,32,16]} % minimum size of patch (default: [32,32,16])
  148. {50} % upsampling factor for subpixel registration (default: 50)
  149. {[4,4,1]} % degree of patches upsampling (default: [4,4,1])
  150. {[3,3,1]} % maximum deviation of patch shift from rigid shift (default: [3,3,1])
  151. {[32,32,16]} % size of overlapping region after upsampling (default: [32,32,16])
  152. {[15,15,5]} % maximum rigid shift in each direction
  153. {false} % use phase correlation (good for high SNR)
  154. {'FFT'} % method for applying shifts ('FFT', 'linear', 'cubic')
  155. % template updating
  156. {true} % flag for online template updating (default: true)
  157. {100} % length of initial batch (default: 100)
  158. {50} % width of each bin (default: 10)
  159. {50} % number of local means to keep in memory (default: 50)
  160. {{'median';'mean'}} % method for averaging the template (default: {'median';'mean'}
  161. {1} % number of data passes (default: 1)
  162. {'copy'} % method of boundary treatment (default: 'copy')
  163. % misc
  164. {0} % add dc value to data (default: 0)
  165. {false} % for each frame, update patches in parallel (default: false)
  166. {false} % flag for saving memory mapped motion corrected file (default: false)
  167. {'motion_corrected.mat'} % name for memory mapped file (default: 'motion_corrected.mat')
  168. {1000} % batch size used during memory mapping for faster mapping
  169. % plotting
  170. {false} % flag for plotting results in real time (default: false)
  171. {false} % flag for making movie (default: false)
  172. {'motion_corrected.avi'} % name for movie (default: 'motion_corrected.avi')
  173. {30} % frame rate for movie (default: 30)
  174. % output_type
  175. {'mat'}
  176. {'mov'}
  177. {'motion_corrected.h5'}
  178. {'motion_corrected.tif'}
  179. % use_windowing
  180. {false}
  181. {0.5}
  182. % bitsize for reading .raw files
  183. {2}
  184. % offset from bidirectional sampling
  185. {true}
  186. {50}
  187. {10}
  188. {[]}
  189. ];
  191. for j = 1:m
  192. if eval(['isempty(options.' Names(j,:) ')'])
  193. eval(['options.' Names(j,:) '= Values{j};']);
  194. end
  195. end
  197. if isempty(options.d1); options.d1 = input('What is the total number of rows? \n'); end
  198. if isempty(options.d2); options.d2 = input('What is the total number of columns? \n'); end
  199. %if options.d3 == 1; nd = 2; else nd = 3; end
  200. if isempty(options.grid_size); options.grid_size = [options.d1,options.d2,options.d3]; end
  201. if length(options.grid_size) == 1; options.grid_size = options.grid_size*ones(1,3); end
  202. if length(options.grid_size) == 2; options.grid_size(3) = 1; end
  203. if length(options.min_patch_size) == 1; options.min_patch_size = options.min_patch_size*ones(1,3); end
  204. if length(options.min_patch_size) == 2; options.min_patch_size(3) = 1; end
  205. if length(options.overlap_pre) == 1; options.overlap_pre = options.overlap_pre*ones(1,3); end
  206. if length(options.overlap_pre) == 2; options.overlap_pre(3) = 1; end
  207. if length(options.overlap_post) == 1; options.overlap_post = options.overlap_post*ones(1,3); end
  208. if length(options.overlap_post) == 2; options.overlap_post(3) = 1; end
  209. if length(options.max_shift) == 1; options.max_shift = options.max_shift*ones(1,3); end
  210. if length(options.max_shift) == 2; options.max_shift(3) = 1; end
  211. if length(options.max_dev) == 1; options.max_dev = options.max_dev*ones(1,3); end
  212. if length(options.max_dev) == 2; options.max_dev(3) = 1; end
  213. if length(options.mot_uf) == 1; options.mot_uf = options.mot_uf*ones(1,3); end
  214. if length(options.mot_uf) == 2; options.mot_uf(3) = 1; end
  215. options.mot_uf(options.grid_size >= [options.d1,options.d2,options.d3]) = 1;

NoRMCorreSetParms.m at commit c329c3b, no license · at the source

Overview

Authors: Koen Kole1,2, Maarten HP Kole1,3
ORCID iDs: Koen Kole, Maarten HP Kole
  1. Department of Axonal Signaling, Netherlands Institute for Neuroscience, Royal Netherlands Academy of Arts and Science, Amsterdam, The Netherlands
  2. Donders Center for Neuroscience, Donders Institute for Brain, Cognition and Behaviour, Nijmegen, The Netherlands
  3. Cell Biology, Neurobiology and Biophysics, Department of Biology, Faculty of Science, University of Utrecht, Utrecht, The Netherlands
Institutions: Netherlands Institute for Neuroscience (Netherlands); Royal Netherlands Academy of Arts and Sciences (Netherlands); Donders Institute for Brain, Cognition and Behaviour (Netherlands); Donders Centre for Neuroscience (Netherlands); Utrecht University (Netherlands)
Journal: The Journal of physiology, volume 604, issue 7, pages 3032-3052
Dates: received 24 May 2025; accepted 11 February 2026; published online 10 March 2026; in print 1 April 2026
Type: Research article · Language: English
License: CC BY
Identifiers: DOI 10.1113/jp289290 · PMID 41806299 · PMCID PMC13039255 · OpenAlex W7134897345
Open access: hybrid, a free copy (OpenAlex)
Status: code verified
Categories: optical imaging (calcium, voltage, 2-photon) (modality), mouse (organism), cellular / molecular (subfield)
Methods: Statistics, Spectral & time-frequency, Evoked potentials, Connectivity, Single-unit activity, calcium imaging
Keywords: action potential, axon initial segment, calcium buffer, mitochondria, pyramidal neuron
MeSH: Action Potentials*, Axon Initial Segment*, Calcium*, Mitochondria*, Pyramidal Cells*, Animals, Calcium Channels, Calcium Signaling, Mice (* major topic)
Topic: Mitochondrial Function and Pathology (Molecular Biology, Biochemistry, Genetics and Molecular Biology), according to OpenAlex
Funding: ZonMw (0 451 001 201 0066); Exacte en Natuurwetenschappen (865.17.003); Netherlands Organisation for Health Research and Development (0 451 001 201 0066)
Citations: not cited yet (Europe PMC); 67 references in the paper
Research resources: we used wild‐type mice RRID:IMSR_RJ:C57BL, RRID:MMRRC_03, RRID:SCR_0, RRID:SCR_01

Abstract

Abstract: Mitochondria are diverse and multifaceted intracellular organelles regulating oxidative energy supply, lipid metabolism and calcium (Ca2+) signalling. In neurons the spatial sequestration of cytoplasmic Ca2+ by mitochondria plays a critical role in determining activity‐dependent spine plasticity, shaping the presynaptic transmitter release characteristics and contributing to sustained action potential firing. Here, we tested the hypothesis that mitochondria at the axon initial segment (AIS) affect the microdomain cytoplasmic Ca2+ transients, thereby regulating Ca2+‐dependent voltage‐gated ion channels at the plasma membrane and initiation of action potentials. Using 3D electron microscopy reconstructions and virally injecting genetically encoded fluorescence indicators we visualized the ultrastructure and distribution of mitochondria selectively in thick‐tufted layer 5 pyramidal neurons. We found that most mitochondria were stably clustered to the proximal AIS, while few were observed at distal sites. Simultaneous two‐photon imaging of action potential‐dependent cytoplasmic and mitochondrial Ca2+, combined with electrophysiological recordings showed that AIS mitochondria exhibit powerful activity‐dependent cytosolic Ca2+ uptake. However, while intracellular application of the mitochondrial Ca2+ uniporter inhibitor Ru360 fully blocked mitochondrial Ca2+ import and increased the slow afterhyperpolarization duration, it did not affect action potential input–output function, action potential dynamics nor the ability to produce high‐frequency burst output. Together, the results indicate that AIS mitochondria are dispensable for temporal and rate encoding, suggesting that mt‐Ca2+ buffering at the AIS may be involved in non‐electrical roles.

Key points: Mitochondrial Ca2+ buffering controls multiple Ca2+‐dependent intracellular processes and their subcellular location of the organelles defines local physiological properties in neurons.

Recent studies implicate mitochondrial Ca2+ uptake in the slow afterhyperpolarization and maintenance of action potential firing.

Using electron microscopy and virally delivered genetically encoded tools we examined mitochondria in the layer 5 pyramidal neuron axon initial segment (AIS), the site where action potentials initiate, and found that cytoplasmic Ca2+ influx is powerfully buffered by proximally clustered mitochondria.

Electrophysiological recordings during the block of the mitochondrial calcium uniporter reveal a role in the slow afterhyperpolarization, while AIS action potential initiation and action potential waveforms are independent from mitochondria.

These findings indicate AIS mitochondria under physiological conditions exert non‐electrical roles.

Reproduced under the paper's license (CC BY), from the paper cited above.

Repositories

Its files are read in the Code ↔ Paper reader above.

Kolelab/mtGCaMP

License: none: the authors keep all their rights
State: the link answers, verified on 30 September 2026
Evidence: files inventoried
Commit: c329c3b7e46844ef53edc10c8fc2af124ad32aff, 14 April 2022
Languages: MATLAB (18)
Size: 54 files, 18 scripts
Software Heritage: not archived
Found in: “Data availability statement”
Holds: README
Not found: license file, CITATION.cff, environment file, tests, continuous integration, documentation
Tools: Image Processing Toolbox (3 files), Curve Fitting Toolbox (1 file), Parallel Computing Toolbox (1 file), Signal Processing Toolbox (1 file), Statistics and Machine Learning Toolbox (1 file)
Availability: 1 check, the latest on 30 September 2026: the link answers
  • 30 September 2026: the link answers
18 files
At the source: github.com/Kolelab/mtGCaMP

microns-explorer.org

License: none: the authors keep all their rights
State: the link answers, verified on 30 September 2026
Evidence: the link answers
Software Heritage: not checked
Found in: “Data availability statement”
Not found: README, license file, CITATION.cff, environment file, tests, continuous integration, documentation
Availability: 1 check, the latest on 30 September 2026: the link answers (HTTP 200)
  • 30 September 2026: the link answers (HTTP 200)
At the source: microns-explorer.org

The paper's code and data availability statement is in the Data section.

Tracing map

Proposed by the machine: these links were found in the paper and verified at the source, without human review. The map will receive a Zenodo DOI once one of the paper's authors has validated it with their ORCID.

What the map holds:

  • 2 repositories of the authors' code, each at its verified commit, with its license and how the link was found in the paper;
  • 18 scripts, each with its path and the digest of its content;
  • no match between paragraphs and code yet;
  • neither the text of the paper nor the code itself.

Its JSON (tracing-map.json) is deposited on Zenodo with its DOI once the map is validated.

Data

No dataset and no data link were found in the paper.

Data availability statement

All data are available upon request to the corresponding authors. The Microns dataset used for 3D EM analysis is available at www.microns‐explorer.org (http://www.microns-explorer.org). The script used to analyse Ca2+ imaging data is available via https://github.com/Kolelab/mtGCaMP.

Reproduced under the paper's license (CC BY), from the paper cited above.

Versions

The history of this record: each version stored by the harvester or made by a correction of its authors or of the maintainers of its code, and what changed in its facts. The texts of the paper (its abstract, its availability statements) are not part of it; versions that changed only those are not listed.

Version 1, 30 September 2026: the first record

Recorded: type, language, journal, volume, issue, pages, dates, 2 authors, 5 keywords, 9 MeSH terms, 3 funders, 67 references, 4 RRIDs.

Cite

This paper

Kole, K., & Kole, M. H. (2026). Robust activity-dependent mitochondrial calcium dynamics at the AIS is dispensable for action potential generation. The Journal of physiology, 604(7), 3032-3052. https://doi.org/10.1113/jp289290

BibTeX

@article{kole2026robust,
author = {Kole, Koen and Kole, Maarten HP},
title = {{Robust activity-dependent mitochondrial calcium dynamics at the AIS is dispensable for action potential generation}},
journal = {The Journal of physiology},
year = {2026},
month = mar,
volume = {604},
number = {7},
pages = {3032--3052},
publisher = {Wiley},
issn = {0022-3751},
doi = {10.1113/jp289290},
url = {https://doi.org/10.1113/jp289290},
pmid = {41806299},
pmcid = {PMC13039255}
}

RIS

TY - JOUR
AU - Kole, Koen
AU - Kole, Maarten HP
TI - Robust activity-dependent mitochondrial calcium dynamics at the AIS is dispensable for action potential generation
T2 - The Journal of physiology
J2 - J Physiol
PY - 2026
DA - 2026/03/10
VL - 604
IS - 7
SP - 3032
EP - 3052
SN - 0022-3751
PB - Wiley
DO - 10.1113/jp289290
UR - https://doi.org/10.1113/jp289290
LA - en
ER -

CSL-JSON

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"id": "10.1113/jp289290",
"type": "article-journal",
"title": "Robust activity-dependent mitochondrial calcium dynamics at the AIS is dispensable for action potential generation",
"container-title": "The Journal of physiology",
"author": [
{
"family": "Kole",
"given": "Koen"
},
{
"family": "Kole",
"given": "Maarten HP"
}
],
"container-title-short": "J Physiol",
"volume": "604",
"issue": "7",
"page": "3032-3052",
"DOI": "10.1113/jp289290",
"PMID": "41806299",
"PMCID": "PMC13039255",
"ISSN": "0022-3751",
"publisher": "Wiley",
"URL": "https://doi.org/10.1113/jp289290",
"language": "en",
"issued": {
"date-parts": [
[
2026,
3,
10
]
]
}
}

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