Method for labeling and reconstruction of single neurons using Sindbis virus vectors

Neuronal dendrites and axons are key substrates for the input and output of information, respectively, so establishing the precise and complete morphological description of dendritic and axonal processes of a single neuron is essential for understanding the neuron's functional role in the neuronal circuits. The whole structure of single neurons was originally revealed using Golgi staining, and later the intracellular labeling method was developed, although this is technically too difficult to stain entire neurons in vivo. Since the late 1980s, molecular biology techniques have been applied to neuroscience research, leading to the development of various virus vectors, such as the Sindbis and adeno-associated virus vectors, which have facilitated the reconstruction of neurons at a single cell level. In the present review, we focus on a method for labeling and reconstruction of single neurons using Sindbis virus vectors that express membrane-targeted fluorescent proteins. We describe in detail a protocol for single-neuron labeling using Sindbis virus vectors, and we provide an example of a recent project at our laboratory in which we successfully applied these methods to study thalamocortical projection neurons. Further, we discuss the strengths and limitations of Sindbis virus vectors for single neuron reconstruction, comparing them with adeno-associated virus vectors.