We used three strategies to study the effects of deleting CNIH-2. Using Cnih2fl/fl mice we: 1) injected AAV-CRE-GFP into the hippocampus of P0–2 mouse pups and then made acute slices three weeks later; 2) made hippocampal slice cultures at P6–9, biolistically transfected neurons with CRE-GFP at DIV4 and recorded 2–3 weeks later (see Experimental Procedures for details); and 3) crossed Cnih2fl/fl mice with the NEX-CRE mouse line. In the first two sets of experiments, simultaneous recordings of AMPAR- and NMDAR-evoked excitatory postsynaptic currents (AMPAR- and NMDAR-eEPSCs, respectively) were made from a green infected/transfected CA1 pyramidal neuron expressing CRE and a neighboring control non-green pyramidal neuron during stimulation of excitatory axons in stratum radiatum. This approach permitted a pair-wise, internally controlled comparison of the consequence of our genetic manipulation. In the third approach using acute slices prepared from NexCnih2−/− mice, the ratio of the AMPAR- and NMDAR-eEPSCs was calculated and compared to wild-type neurons.
