Mapping Neuronal Connectivity using Laser Photostimulation and Calcium Imaging

Abstract

Knowledge of the connectivity of a neuronal network is essential for understanding its activity and function. Current anatomical imaging techniques can be tedious and have a high degree of uncertainty. Electrophysiological techniques, although precise, are limited by network size. We present here a novel technique that allows relatively fast determination of connectivity in cultured neurons. This technique combines laser scanning photostimulation (via glutamate uncaging) to stimulate and calcium imaging to record activity of a large population of neurons. Mapping proceeds by selectively stimulating a single neuron and monitoring the response of nearby neurons to infer the postsynaptic connections. Within a few hours, we can generate a map of the functional connectivity of relatively large neuronal networks consisting of ∼150 neurons and ∼1500 connections. We describe the method in detail and discuss the feasibility of mapping neuronal microcircuits in acute brain slices.