Two-photon targeted patching and electroporation in vivo.

Abstract

By combining patch-clamp methods with two-photon microscopy, it is possible to target recordings to specific classes of neurons in vivo. Here we describe methods for imaging and recording from the soma and dendrites of neurons identified using genetically encoded probes such as green fluorescent protein (GFP) or functional indicators such as Oregon Green BAPTA-1. Two-photon targeted patching can also be adapted for use with wild-type brains by perfusing the extracellular space with a membrane-impermeable dye to visualize the cells by their negative image and target them for electrical recordings, a technique termed "shadowpatching." We discuss how these approaches can be adapted for single-cell electroporation to manipulate specific cells genetically. These approaches thus permit the recording and manipulation of rare genetically, morphologically, and functionally distinct subsets of neurons in the intact nervous system.