Talk:Iglesias-Gonzalez 2013 J Neurosci Methods
Ice-cold isolation buffer used in study
225 mM Mannitol
75 mM Sucrose
5 mM HEPES,
1 mg/ml fatty acid free BSA (pH 7.4, isotonized with KOH)
- EGTA or EDTA added at concentrations in the range of: 1 - 3 mM
- Final mitochondrial pellet was resusbended in IB without chelators.
- Respiratory assays were carried out as previously described in Iglesias-Gonzalez 2012 Neurochem Res:
Standard respiration medium contains:
125 mM KCl
5 mM HEPES
3 mM MgCl2
2 mM KH2PO4
0,5 mM EGTA (pH 7.4)
Chemicals for MiR05/MiR06
Calcium and mitochondria
Abstract excerpt from Gunter et al.:
The literature suggests that the physiological functions for which mitochondria sequester Ca(2+) are (1). to stimulate and control the rate of oxidative phosphorylation, (2). to induce the mitochondrial permeability transition (MPT) and perhaps apoptotic cell death, and (3). to modify the shape of cytosolic Ca(2+) pulses or transients. There is strong evidence that intramitochondrial Ca(2+) controls both the rate of ATP production by oxidative phosphorylation and induction of the MPT. Since the results of these processes are so divergent, the signals inducing them must not be ambiguous. Furthermore, as pointed out by Balaban [J. Mol. Cell. Cardiol. 34 (2002 ) 11259-11271], for any repetitive physiological process dependent on intramitochondrial free Ca(2+) concentration ([Ca(2+)](m)), a kind of intramitochondrial homeostasis must exist so that Ca(2+) influx during the pulse is matched by Ca(2+) efflux during the period between pulses to avoid either Ca(2+) buildup or depletion. In addition, mitochondrial Ca(2+) transport modifies both spatial and temporal aspects of cytosolic Ca(2+) signaling...
MiRK03
>> from intern wiki MiRK03
KCl ? mM, adjust osmolarity (official: osmotic concentration) to MiR05
HEPES free acid 20 mM
KH2PO4 10 mM
MgCl2 3 mM
EGTA 0.5 mM
BSA: 0.1%
pH = 7.0
KCl 130 mM
