Isoproterenol (Sigma-Aldrich) was subcutaneously delivered to WT, DN-Mst1 and Mst1-KO mice (average age of 6 months) by mini-osmotic pumps (ALZET model 2001, DURECT Corp, Cupertino, CA, USA) at a dose of 60 mg/kg/day as described [23 (link)] for 24 h. Immediately after pump implantation, WT, DN-Mst1 and Mst1-KO mice were injected with 1 % Evans blue dye (EBD) via tail-vein at a dose of 100 mg/kg [28 (link)]. After 24 h, animals were killed. LV was mounted in Tissue-Tek OCT compound (Sakura, Torrance, CA, USA) and frozen in isopentane pre-cooled in liquid nitrogen. Cryostat sections (10 µm thickness) were counterstained with WGA Alexa Fluor 488 conjugate (Invitrogen) and EBD positive myocytes were quantified. Necrotic myocytes were presented as percentage of EBD positive myocytes per myocyte nuclei.
Protocol detail
Isoproterenol-Induced Myocardial Necrosis
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Alexa fluor 488
Animals
Evans blue
Frozen
Implantation
Isopentane
Isoproterenol
Mice
Myocyte
Necrotic
Nitrogen
Nuclei
Osmotic
Tail
Tissue
Vein
Corresponding Organization :
Other organizations : Rutgers, The State University of New Jersey
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Variable analysis
independent variables
- Isoproterenol (Sigma-Aldrich) delivered subcutaneously to WT, DN-Mst1 and Mst1-KO mice at a dose of 60 mg/kg/day for 24 h
- Percentage of Evans blue dye (EBD) positive myocytes per myocyte nuclei (necrotic myocytes)
- Average age of mice: 6 months
- Mini-osmotic pumps (ALZET model 2001) used for Isoproterenol delivery
- Tail-vein injection of 1% Evans blue dye at 100 mg/kg
- Tissue processing: LV mounted in Tissue-Tek OCT compound, frozen in isopentane, cryostat sections (10 µm) counterstained with WGA Alexa Fluor 488 conjugate
- Not explicitly mentioned
- WT, DN-Mst1 and Mst1-KO mice
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