Chunk #3 — Methods and Materials — Animals and Tissue Processing — Generation of CB1R heterozygous and knockout mice

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Cortical glutamic acid decarboxylase 67 deficiency results in lower cannabinoid 1 receptor messenger RNA expression: implications for schizophrenia.

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CB1R+/− and CB1R−/− mice were generated as previously described (12). The Cnr1 gene was mutated in MPI2 embryonic stem cells by replacing the coding region between amino acids 32 and 448 with PGK-neo. Chimeric mice derived from these cells were bred with C57BL/6J animals. Backcrossing of chimeric and heterozygous animals to C57BL/6J mice and interbreeding of CB1R+/− animals produced CB1R−/− mutants and wild-type mice. Male animals were euthanized at either eight weeks of age (CB1R+/−, CB1R−/−, and WT; n=6 per group) or four weeks of age (CB1R−/− and WT; n=7 per group) and brains were removed, frozen, and stored at −80°C. Fresh frozen brains were provided by Bristol-Myers Squibb.