A new era for functional labeling of neurons: activity-dependent promoters have come of age

Abstract

Genetic labeling of neurons with a specific response feature is an emerging technology for precise dissection of brain circuits that are functionally heterogeneous at the single-cell level. While immediate early gene mapping has been widely used for decades to identify brain regions which are activated by external stimuli, recent characterization of the promoter and enhancer elements responsible for neuronal activity-dependent transcription have opened new avenues for live imaging of active neurons. Indeed, these advancements provided the basis for a growing repertoire of novel experiments to address the role of active neuronal networks in cognitive behaviors. In this review, we summarize the current literature on the usage and development of activity-dependent promoters and discuss the future directions of this expanding new field.

Keywords: Arc; E-SARE; activity-dependent gene expression; c-fos; live imaging; neuronal ensemble.

FIGURE 1

Engineering of a synthetic E-SARE promoter. (A) The structure of SARE enhancer element which regulates activity-dependent expression of Arc/Arg3.1. Three distinct transcription factors, CREB, MEF2, and SRF, cooperate to induce strong transcription. (B) Top, construction strategy of E-SARE promoter. The locations of SARE and TATA-containing ArcMin in the Arc promoter are represented by black boxes. E-SARE is constructed by connecting SARE elements in tandem. Bottom, comparison of E-SARE with SARE-ArcMin and c-fos promoters by luciferase reporter assay in cultured neurons under resting (blue) and stimulated (pink) conditions. Dynamic ranges (calculated based on the ratio between stimulated and resting conditions; white) are also shown. Inset, expanded values obtained under resting conditions. Adapted from Kawashima et al. (2013).

FIGURE 2

In vivo recording reveals a strong correlation between evoked neuronal activity and E-SARE expression. (A) Top, experimental setup. Single-cell responses of E-SARE-driven GFP-positive and – negative neurons were recorded under the observation by two-photon microscopy. Bottom, representative images of a recorded neuron that is GFP reporter-positive (filled arrow). The dashed lines and line arrows indicate the recording electrode and GFP-negative neurons, respectively. Scale bar, 20 μm. (B) GFP brightness of recorded neurons correlates with their whisker-evoked activity (left) but not with spontaneous activity rate (right). Adapted from Kawashima et al. (2013).