A brain-sparing diphtheria toxin for chemical genetic ablation of peripheral cell lineages

Abstract

Conditional expression of diphtheria toxin receptor (DTR) is widely used for tissue-specific ablation of cells. However, diphtheria toxin (DT) crosses the blood–brain barrier, which limits its utility for ablating peripheral cells using Cre drivers that are also expressed in the central nervous system (CNS). Here we report the development of a brain-sparing DT, termed BRAINSPAReDT, for tissue-specific genetic ablation of cells outside the CNS. We prevent blood–brain barrier passage of DT through PEGylation, which polarizes the molecule and increases its size. We validate BRAINSPAReDT with regional genetic sympathectomy: BRAINSPAReDT ablates peripheral but not central catecholaminergic neurons, thus avoiding the Parkinson-like phenotype associated with full dopaminergic depletion. Regional sympathectomy compromises adipose tissue thermogenesis, and renders mice susceptible to obesity. We provide a proof of principle that BRAINSPAReDT can be used for Cre/DTR tissue-specific ablation outside the brain using CNS drivers, while consolidating the link between adiposity and the sympathetic nervous system.

Profiling of G protein-coupled receptors in vagal afferents reveals novel gut-to-brain sensing mechanisms

“…In summary, the unexpected identification of many GPCRs with unknown ligands and functions in vagal afferents highlights how little is known about vagal signaling. Many of the genes identified in this study may be used to generate novel transgenic mice to investigate the functional neuroanatomy of vagal afferents, for instance, using diphtheria toxin-assisted ablation tools [86] to determine their exact functions. Thus, we are hopeful that our data will guide future studies aimed at addressing the role of GPCRs signaling in visceral afferents in relation to metabolic and immune sensing, as well as metabolic diseases and their complications.…”

Leptin and brain–adipose crosstalks

“…2). Newly developed genetic tools that enable targeted manipulations of molecules involved in sympathetic signalling represent a promising way to delineate the tissue-specific regulation of leptin production 59 . Overall, these observations indicate that SNS activity decreases leptin expression independently of adiposity, and most likely through activation of β 3 -AR 25–29 , although other receptors may also regulate leptin.…”

DTR‐mediated conditional cell ablation—Progress and challenges

“…Likewise, DTx access to the brain, but not the peripheral lymphoid organs was successfully inhibited by neutralizing antibodies . In an interesting extension of this approach Domingos and colleagues recently introduced a brain‐sparing DTx form . Specifically, the authors prevented the blood–brain barrier passage of DTx through DTx PEGylation allowing them to specifically ablate peripheral sympathetic neurons without affecting the CNS .…”

“…In an interesting extension of this approach Domingos and colleagues recently introduced a brain‐sparing DTx form . Specifically, the authors prevented the blood–brain barrier passage of DTx through DTx PEGylation allowing them to specifically ablate peripheral sympathetic neurons without affecting the CNS . More detailed characterization of the pharmacokinetic availability and the half‐life of DTx and its modified variants could be an interesting future avenue to improve the use of the DTx/DTR system.…”