Behavioral and Physiological Effects of a Novel Kappa-Opioid Receptor-Based DREADD in Rats

Abstract

In the past decade, novel methods using engineered receptors have enabled researchers to manipulate neuronal activity with increased spatial and temporal specificity. One widely used chemogenetic method in mice and rats is the DREADD (designer receptors exclusively activated by designer drugs) system in which a mutated muscarinic G protein-coupled receptor is activated by an otherwise inert synthetic ligand, clozapine-N-oxide (CNO). Recently, the Roth laboratory developed a novel inhibitory DREADD in which a mutated kappa-opioid receptor (KORD) is activated by the pharmacologically inert drug salvinorin B (SalB; Vardy et al, 2015). They demonstrated the feasibility of using KORD to study brain circuits involved in motivated behavior in mice. Here, we used behavioral, electrophysiological, and neuroanatomical methods to demonstrate the feasibility of using the novel KORD to study brain circuits involved in motivated behavior in rats. In Exp. 1, we show that SalB dose-dependently decreased spontaneous and cocaine-induced locomotor activity in rats expressing KORD to midbrain (ventral tegmental area/substantia nigra). In Exp. 2, we show that SalB completely inhibited tonic firing in KORDexpressing putative dopamine neurons in midbrain. In Exp. 3, we used a 'retro-DREADD' dual-virus approach to restrict expression of KORD in ventral subiculum neurons that project to nucleus accumbens shell. We show that KORD activation selectively decreased novel context-induced Fos expression in this projection. Our results indicate that the novel KORD is a promising tool to selectively inactivate brain areas and neural circuits in rat studies of motivated behavior.

Role of projections from ventral subiculum to nucleus accumbens shell in context-induced reinstatement of heroin seeking in rats

“…In this regard, our present and previous data (Bossert et al 2012) suggest that context-induced reinstatement may be particularly susceptible to ipsilateral disconnections. The development of optogenetic (Ma et al 2014; Yizhar et al 2011) and DREADD (Boender et al 2014; Marchant et al 2015; Nair et al 2013) methods for projection-specific inhibition will allow us and other investigators to directly test this idea by comparing the effects of single-hemisphere projection inhibition versus dual-hemisphere projection inhibition on reinstatement of drug seeking.…”

DREADDs for Neuroscientists

“…In the neuronal cell bodies, recombination of AAV-FLEX-DREADD constructs can occur to allow expression of DREADDs in a projection-specific fashion (Boender et al, 2014) (Figure 4B). The use of CAV-Cre and FLEX-DREADD constructs has been dubbed the “Retro-DREADD” technique (Marchant et al, 2016) and in theory could be used for intersectional and multiplexed applications.…”

“…Salvinorin B does retain modest affinity for KOR (>100 nM) so that investigators using the KORD should use the lowest dose possible and verify no effects of salvinorin B in the absence of KORD. Several labs have reported successful inhibition of neural activity with KORD (Marchant et al, 2016; Vardy et al, 2015; Denis et al, 2015). …”

Assaying the Stability and Inactivation of AAV Serotype 1 Vectors

“…The prevalence of AAV as a vector stems from its low immunogenicity, 1,2 high titers (*1 · 10 14 viral genomes/mL), no associated human disease, 3 helper virus-free production, 4 and high stability in terms of both duration of transgene expression in transduced cell and stability as a viral particle. 5,6 AAV serotype 1 (AAV1) is used for neuroscience applications such as opotogenetics, 7,8 pharmacogenetics, [9][10][11] and gene silencing. 12 AAV1 is also being used in human clinical trials for heart disease, 13 alpha-1 antitrypsin deficiency, 14 limbgirdle muscular dystrophy type 2D, 15 CharcotMarie-Tooth neuropathy, 16 and Becker muscular dystrophy.…”