Genetically-engineered inducible cell ablation mice selective for forebrain GABAergic cells

J. E. BELFORTE, G. KUNOS, K. NAKAZAWA

San Diego, USA

Congreso; Annual Meeting Society for Neuroscience; 2007

Society of Neuroscience

Local GABAergic interneurons sculpt the activities in cortical circuits controlling excitability and timing of principal neurons. They have been implicated in the generation and control of theta/gamma frequency oscillations and in the normal development of cortical circuitry, including the regulation of neuronal proliferation and migration. Converging clinical evidence suggests that altered interneuron function may underlie the pathophysiological process of major psychiatric disorders. In spite of these recent achievements, no pharmacological manipulations exist to specifically control in vivo activity of interneurons. To overcome this limitation we have developed a novel genetically engineered mouse strain in which Cre expression is targeted specifically to forebrain GABAergic cells. We homologously integrated Cre recombinase cDNA into the promoter region of the protein phosphatase 1 regulatory (inhibitor) subunit 2 (Ppp1r2) in bacterial artificial chromosome (BAC), and several BAC transgenic mice were generated. One of the strains, Cre#4127, showed a selective Cre recombination pattern on a significant population of GAD67-positive cells in the cerebral cortex and hippocampus but not in other brain regions, such as thalamus and cerebellum. Immunohistochemical analyses revealed that more than 90% of the beta-Gal positive cells, after crossing with R26R-LacZ reporter line, are GAD67 positive while they do not co-localize with glutamatergic neuron markers, such as TBR1 and alpha-CaMKII, suggesting that Cre recombination occurs predominantly in the forebrain GABAergic cells. Furthermore, 40-75% of the beta-Gal positive cells were parvalbumin (PV) positive depending on the cortical areas, while none of them were calretinin positive. Cre-mediated recombination began from the first postnatal week, allowing early manipulation of the targeted cells. To induce ablation of the targeted cells, we crossed the Cre line with a floxed-diphtheria toxin (DT) receptor strain (provided by Drs. Ari Waisman and Thorsten Buch), in which the Cre expressing cells can be ablated upon i.p. administration of DT. Within 24 hrs of DT administration apoptotic neurons were identified by activated caspase-3 immunoreactivity along the cortices, confirming the cell ablation. After a single administration of DT at postnatal day 10, we observed a decreased number of GAD67 positive cells, and up to a 30 % reduction of the PV positive cells. Behavioral analysis of this strain is underway to advance our understanding of in vivo function of cortical GABAergic interneurons.