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Pharmacol Ther (2014) 141(2):11724

For CPA unloading, brain slices were immersed into 10 C and 10 C solutions of decreasing concentration of V3 in the carrier solution with additional 300 mM mannitol, followed by a final step of carrier solution without added mannitol: 45% w/v V3 for 2 min and 30% at 10 C for 5 min, followed by 16% w/v V3 for 10 min, 8% for 3 min, 4% for 3 min, 2% for 3 min, 0% for 10 min at 10 C each, followed by a final step of LM5 without added mannitol at 10 C for 10 min ( table S3 ) Metabolic analysis After cryopreservation using the vitrification protocol, the slices were incubated for 2 hours in carbogenated aCSF at RT to allow recovery