Wild type c57bl 6 mice

Wild-type C57BL/6 mice were purchased from Taconic. Mice carrying Fcgr2b^fl or Fcgr2b⁻ alleles were generated from B6 ES cells (Supplementary Figure 1). Mb1Cre/Fcgr2b^fl/fl, Cg1Cre/Fcgr2b^fl/fl, CD11cCre/Fcgr2b^fl/fl, LysMCre/Fcgr2b^fl/fl mice were generated by crossing mice carrying Fcgr2b^fl alleles to Mb1Cre mice (23 (link)), Cg1Cre mice (24 (link)), CD11cCre mice (25 (link)), and LysMCre mice (26 (link)), respectively, which have been backcrossed to the B6 background for at least 10 times. In some mice, Mb1Cre and CD11cCre also mediated germline deletion of one Fcgr2b^fl allele and gave rise to Mb1Cre/Fcgr2b^fl/− and CD11cCre/Fcgr2b^fl/− mice. B6.Fcgr2b₁₂₉^−/−(N12) were obtained from Taconic. Fcer1g^−/−Fcgr2b^−/− mice had been described previously (27 (link)). All mice were maintained in The Rockefeller University Comparative Bioscience Center. All experiments were performed in compliance with federal laws and institutional guidelines and had been approved by the Rockefeller University IACUC.

Wild-type C57BL/6 mice, were obtained from Taconic (Cambridge City, IN). Male mice age 5 weeks (18–23gr) were grouped and housed in single grommet ventilated plexiglass cages at ambient temperature (21°C) in a room maintained on a reversed 12L:12D cycle (lights off at 10:00, lights on at 22:00) in our animal facility that has Association for Assessment and Accreditation of Laboratory Animal Care approval. Food and water were provided ad libitum. The mice were given ~7 days to acclimatize to the housing conditions and reverse light cycle before the start of the experiments. Mice were then habituated to the containment boxes for the Von Frey assay. All animal procedures were pre-approved by our Institutional Animal Care and Use Committee and were in accordance with the National Institutes of Health Guide for the Care and Use of Laboratory Animals. Mice were not deprived of food or water at any time.

All mice were housed indoors with food and water provided ad libitum. For 3D motion tracking under the microscope, mice with endogenous mCherry fluorescence within the vasculature were used in all in vivo experiments except for uncoupling experiments using TMRM. To generate a knock-in mouse line with mCherry-tagged to the N-terminus of endogenous non-muscle myosin 2A, the mCherry sequence was inserted at the 5′ of the initiating ATG codon in exon 2. The 4-kb 5’ arm immediately upstream of the ATG is followed by the mCherry-tagged exon2 plus 160-bp DNA sequence immediately 3’ of exon2, a loxP-flanked PGK-Neor cassette and the 2-kb 3’ arm. Nucleotide sequences of the cloned DNA fragments were confirmed by sequencing. Male and Female Mice 2–3 months of age were used for flux experiments. For uncoupling experiments, wild-type C57BL/6 mice (Taconic Biosciences) were used to allow for multiplex imaging with TMRM. Mice were euthanized via anoxia (nitrogen exposure).

All animal work was done following a protocol approved by the Institutional Animal Care and Use Committee of New York Presbyterian Hospital/Weill Cornell Medical College (IACUC # 2010–0050, 2015–0028). Wild type C57BL/6 mice (strain: C57BL/6NTac) and athymic nude mice (strain: B6.Cg/NTac-Foxn1nu NE10) were purchased from Taconic Biosciences. All mice were maintained under pathogen-free conditions in the Weill Cornell Medicine animal facility. All animal studies were done in accordance with Animal Research: Reporting of In Vivo Experiments (ARRIVE) guidelines.