All of the studies performed were approved by the University of Kentucky Institutional Animal Care and Use Committee, in compliance with the guidelines of the Association for the Assessment and Accreditation for Laboratory Animal Care, International and the National Institutes of Health Guide for the Care and Use of Laboratory Animals.12 Animal experiments complied with Animal Research: Reporting of In Vivo Experiments guidelines. All experiments were conducted using male C57BL/6J mice (2–3 months old; Jackson Laboratories, Bar Harbor, ME, USA).
The animals were housed five per cage, maintained in a 14 h light/10 h dark cycle, fed a balanced diet ad libitum. Animals were randomly assigned to groups, based on injury designation (sham or CCI) and treatment designation (vehicle or pioglitazone). Treatments were given in random order. All experimentation was performed blinded to treatment groups. For all outcomes, experiments were conducted with biological replicates of N = 4–8/group. Additionally, technical triplets were used in each assay.
Two separate cohorts were used for this study. The acute study was conducted to examine the effect of pioglitazone treatment after mild brain contusion on total, glia-enriched, and synaptic mitochondrial bioenergetic measures in the brain. Experimental groups for the acute study were euthanized at 48 h after injury. The subacute efficacy study was conducted to examine how modulation of acute mitochondrial bioenergetics with pioglitazone translated into cortical neuroprotection after mild brain contusion. Experimental groups for the subacute efficacy study were euthanized at 15 days after injury. Mice received a bolus (i.p.) administration of pioglitazone (100 µL volume; 1:1 DMSO and PEG400) at either 0.25, 3, 12, or 24 h after mild CCI, based on the treatment strategy outlined in Hubbard et al.4 (link) The 15-day efficacy study utilized 7-day duration Alzet Mini Pump 1007D (release of 0.5 µL/h; Alzet, Cupertino, CA, USA).
The animals were housed five per cage, maintained in a 14 h light/10 h dark cycle, fed a balanced diet ad libitum. Animals were randomly assigned to groups, based on injury designation (sham or CCI) and treatment designation (vehicle or pioglitazone). Treatments were given in random order. All experimentation was performed blinded to treatment groups. For all outcomes, experiments were conducted with biological replicates of N = 4–8/group. Additionally, technical triplets were used in each assay.
Two separate cohorts were used for this study. The acute study was conducted to examine the effect of pioglitazone treatment after mild brain contusion on total, glia-enriched, and synaptic mitochondrial bioenergetic measures in the brain. Experimental groups for the acute study were euthanized at 48 h after injury. The subacute efficacy study was conducted to examine how modulation of acute mitochondrial bioenergetics with pioglitazone translated into cortical neuroprotection after mild brain contusion. Experimental groups for the subacute efficacy study were euthanized at 15 days after injury. Mice received a bolus (i.p.) administration of pioglitazone (100 µL volume; 1:1 DMSO and PEG400) at either 0.25, 3, 12, or 24 h after mild CCI, based on the treatment strategy outlined in Hubbard et al.4 (link) The 15-day efficacy study utilized 7-day duration Alzet Mini Pump 1007D (release of 0.5 µL/h; Alzet, Cupertino, CA, USA).
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