Mice Lacking the 65 kDa Isoform of Glutamic Acid Decarboxylase (GAD65) Maintain Normal Levels of GAD67 and GABA in Their Brains but Are Susceptible to Seizures

Cleft palate and decreased brain γ-aminobutyric acid in mice lacking the 67-kDa isoform of glutamic acid decarboxylase

“…A TrpS-LacZ͞pGKNeo cassette (16) was inserted into a unique BamHI site in exon 1. Diphtheria toxin-A gene (pMC1DT-ApA) was included in the targeting vector to allow selection against random integration (12). The targeting vector (Fig.…”

“…These ES cells were injected into eight-cell embryos from ICR mice. The mutant mice were obtained by mating chimeric mice with C57BL͞6 mice, as described previously (12,16). The day when the vaginal plug appeared in the dam was designated as embryonic day 0.5 (E0.5), and the morning following birth was designated as postnatal day 0.5 (P0.5).…”

“…Total RNA was prepared from the cerebral cortex of P0.5 mice using TRIzol Reagent (GIBCO͞BRL). RNA was reverse transcribed and PCR amplified by using GAD67 5Ј and 3Ј primers [corresponding to nucleotides 282-303 and 759-777 of GAD67 cDNA (15), respectively] or GAD65 5Ј and 3Ј primers [corresponding to nucleotides 921-940 and 1257-1276 of the GAD65 cDNA (12), respectively]. Glyceraldehyde-3-phosphate dehydrogenase (G3PDH) primers were used to assess the quality of the RNA (17).…”

“…The use of gene targeting for the selective elimination of each GAD isoform is expected to settle these issues. The recent production of GAD65 Ϫ͞Ϫ mice (12,13) yielded the unexpected finding that lack of GAD65 does not change brain GABA contents or animal behavior, except for a slight increase in susceptibility to seizures. Here we report the production of GAD67 Ϫ͞Ϫ mice.…”

GAD1 (2q31.1), which encodes glutamic acid decarboxylase (GAD67), is associated with childhood-onset schizophrenia and cortical gray matter volume loss

“…5 Studies of GAD 67 and GAD 65 knockout mice revealed very different phenotypes: GAD 65 À/À maintained normal levels of GABA, which was in sharp contrast to GAD 67 À/À mice that died the first morning after birth and GAD 67 þ /À mice that had about one-third reduction in GABA in cortex. 6,7 A number of postmortem brain studies of schizophrenics have studied GAD expression. Akbarian et al 8 reported reduced prefrontal expression of GAD 67 in the absence of significant cell loss; axon terminals of chandelier neurons (which are GABAergic interneurons) were reduced in the middle layers of the prefrontal cortex (PFC); 9 GAD 67 mRNA expression was relatively unaltered in most PFC GABA neurons, but reduced below a detectable level in a subset of GABA neurons; 10 Guidotti et al 11 studied GAD 67 protein expression in the PFC as well as cerebellum, and found a 50% reduction in schizophrenic and bipolar patients as compared to controls.…”