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New inducible genetic method reveals critical roles of GABA in the control of feeding and metabolism

Significance We established a new genetic method for rapidly activating Cre recombinase that is based on suppression of a nonsense codon within Cre by a new generation of aminoglycosides. We applied this strategy to successfully silence both enzymes responsible for GABA biosynthesis in hypothalamic agouti-related peptide (AgRP) neurons that control appetite and other behaviors. Inactivation of GABA signaling in AgRP neurons of young adult mice resulted in severe loss of body weight and abnormal glucose metabolism, whereas older mice only manifested transient appetite deficiency. This genetic system adds another robust tool that can be used to understand complex neurological processes. Currently available inducible Cre/loxP systems, despite their considerable utility in gene manipulation, have pitfalls in certain scenarios, such as unsatisfactory recombination rates and deleterious effects on physiology and behavior. To overcome these limitations, we designed a new, inducible gene-targeting system by introducing an in-frame nonsense mutation into the coding sequence of Cre recombinase (nsCre). Mutant mRNAs transcribed from nsCre transgene can be efficiently translated into full-length, functional Cre recombinase in the presence of nonsense suppressors such as aminoglycosides. In a proof-of-concept model, GABA signaling from hypothalamic neurons expressing agouti-related peptide (AgRP) was genetically inactivated within 4 d after treatment with a synthetic aminoglycoside. Disruption of GABA synthesis in AgRP neurons in young adult mice led to a dramatic loss of body weight due to reduced food intake and elevated energy expenditure; they also manifested glucose intolerance. In contrast, older mice with genetic inactivation of GABA signaling by AgRP neurons had only transient reduction of feeding and body weight; their energy expenditure and glucose tolerance were unaffected. These results indicate that GABAergic signaling from AgRP neurons plays a key role in the control of feeding and metabolism through an age-dependent mechanism. This new genetic technique will augment current tools used to elucidate mechanisms underlying many physiological and neurological processes.