Considerations when using cre-driver rodent lines for studying ventral tegmental area circuitry

Abstract

The use of Cre-driver rodent lines for targeting ventral tegmental area (VTA) cell types has generated important and novel insights into how precise neurocircuits regulate physiology and behavior. While this approach generally results in enhanced cellular specificity, an important issue has recently emerged related to the selectivity and penetrance of viral targeting of VTA neurons using several Cre-driver transgenic mouse lines. Here, we highlight several considerations when utilizing these tools to study the function of genetically defined neurocircuits. While VTA dopaminergic neurons have previously been targeted and defined by the expression of single genes important for aspects of dopamine neurotransmission, many VTA and neighboring cells display dynamic gene expression phenotypes that are partially consistent with both classically described dopaminergic and non-dopaminergic neurons. Thus, in addition to varying degrees of selectivity and penetrance, distinct Cre lines likely permit targeting of partially overlapping, but not identical VTA cell populations. This Matters Arising Response paper addresses the Lammel et al. (2015) Matters Arising paper, published concurrently in Neuron.

Figure 1. Differences in TH immunofluoresence intesity across the medial-lateral VTA axis

(A) Confocal image of medial TH+ neurons with optimal gain (B) Confocal image of lateral TH+ neurons with gain optomized for the medial portion. Note that most neurons are oversaturated. (C) Confocal image of medial VTA TH+ neurons with gain optimized for lateral VTA TH+ neurons. Note that neurons that were previously visible (A, white arrows) are now difficult to discern. (D) Confocal image of lateral TH+ neurons with gain optimized for lateral portion.

Figure 2. DAT-Cre, TH-Cre, and VGat-Cre targeted neurons project to the LHb

Confocal image of a coronal LHb section (~1.7 posterior) showing expression of ChR2-eYFP fibers in the LHb following injection of cre-inducible ChR2-eYFP into the VTA of a DAT-Cre mouse (A), TH-Cre mouse (B), and VGat-Cre mouse (C). Scale bar = 200 µm.

Figure 3. Dual fluorescent in situ hybridization for TH and GAD67 in the VTA

(A,B) Confocal image of the anterior medial VTA (~2.8 posterior) from a wild type mouse showing appreciable co-localization between TH mRNA (green) and GAD67 mRNA (red). Overlap = 32/191 neurons (16.7%). Scale bar (A) = 200 µm. Scale bar (B) = 50 µm. (C,D) Confocal image of the posterior lateral VTA (~3.2 posterior) from a wild type mouse showing minimal co-localization between TH mRNA and GAD67 mRNA. Overlap = 1/263 neurons (0.38%) Scale bar (C) = 200 µm. Scale bar (D) = 50 µm.