Figure 2.

Setup and experimental strategies used to prevent spikes during functional mapping of synaptic inputs in vivo. (A) Schematic of the preparation showing simultaneous intracellular recording and dendritic calcium imaging in a layer 2/3 pyramidal neuron in vivo. (B) The electrode enters the cortex through a hole in the coverglass. (C) Spikes are suppressed either by injection of a constant negative current to hyperpolarize the neuron below spike threshold (left), or by including QX-314 in the pipette solution (middle), or by iontophoresis of GABA from an adjacent patch pipette glued to the sharp microelectrode (right). (D) Fabrication of the double barrel electrode used for simultaneous intracellular recording and extracellular iontophoresis of GABA: a bent patch pipette and a straight sharp microelectrode are bonded together by UV-sensitive glue. Bottom right, micrograph of the two tips of a completed double barrel electrode (inter-tip distance: 15 μm). Also see Figure A4 in Appendix for detailed steps on the assembly of double barrel electrodes.