Comparison of three DREADD agonists acting on Gq-DREADDs in the ventral tegmental area to alter locomotor activity in tyrosine hydroxylase:Cre male and female rats

Abstract

Rationale: Despite the increasingly pervasive use of chemogenetic tools in preclinical neuroscience research, the in vivo pharmacology of DREADD agonists remains poorly understood. The pharmacological effects of any ligand acting at receptors, engineered or endogenous, are influenced by numerous factors including potency, time course, and receptor selectivity. Thus, rigorous comparison of the potency and time course of available DREADD ligands may provide an empirical foundation for ligand selection.

Objectives: Compare the behavioral pharmacology of three different DREADD ligands clozapine-N-oxide (CNO), compound 21 (C21), and deschloroclozapine (DCZ) in a locomotor activity assay in tyrosine hydroxylase:cre recombinase (TH:Cre) male and female rats.

Methods: Locomotor activity in nine adult TH:Cre Sprague-Dawley rats (5 female, 4 male) was monitored for two hours following administration of d-amphetamine (vehicle, 0.1-3.2 mg/kg, IP), DCZ (vehicle, 0.32-320 µg/kg, IP), CNO (vehicle, 0.32-10 mg/kg), and C21 (vehicle, 0.1-3.2 mg/kg, IP). Behavioral sessions were conducted twice per week prior to and starting three weeks after bilateral intra-VTA hM3Dq DREADD virus injection.

Results: d-Amphetamine significantly increased locomotor activity pre- and post-DREADD virus injection. DCZ, CNO, and C21 did not alter locomotor activity pre-DREADD virus injection. There was no significant effect of DCZ, CNO, and C21 on locomotor activity post-DREADD virus injection; however, large individual differences in both behavioral response and receptor expression were observed.

Conclusions: Large individual variability was observed in both DREADD agonist behavioral effects and receptor expression. These results suggest further basic research would facilitate the utility of these chemogenetic tools for behavioral neuroscience research.

Keywords: Clozapine N-oxide; Compound 21; DREADD; Deschloroclozapine; Locomotor activity; TH:Cre rats.

Figure 1.

Effects of amphetamine, DCZ, CNO, and C21 alone on locomotor activity both pre-DREADD and post-DREADD virus injection. All points in panels A represent the mean±SEM of 6 rats. Panels B and C compare amphetamine potency (B) and time course (C) pre-DREADD and post-DREADD virus injection. Pre-DREADD points represent mean±SEM of 6 rats and post-DREADD points represent mean±SEM of 9 rats in panels B and C. Panels D-F show individual subject behavioral data for deschloroclozapine (D), clozapine-N-oxide (E), and compound 21 (F) in the six rats that showed mCherry expression and the three rats that failed to show mCherry expression “no expression.” * denotes p<0.05 compared to vehicle (0 drug dose).

Figure 2.

Individual locomotor activity effects of DCZ, CNO, and C21 alone (A, C, E) and corresponding intra-VTA Gq DREADD expression (B, D, F) in three TH:Cre rats that showed mCherry expression and a behavioral response to the chemogenetic ligands post-DREADD virus injections. DAPI staining is represented in blue, TH in red, and mCherry in green. Yellow represents colocalization of TH and mCherry.

Figure 3.

Individual locomotor activity effects of DCZ, CNO, and C21 alone (A, C, E) and corresponding intra-VTA hM3Dq DREADD expression (B, D, F) in three TH:Cre rats that showed mCherry expression, but little-to-no behavioral response to the chemogenetic ligands post-DREADD virus injections. DAPI staining is represented in blue, TH in red, and mCherry in green. Yellow represents colocalization of TH and mCherry.

Figure 4.

Individual locomotor activity effects of DCZ, CNO, and C21 alone (A, C, E) and corresponding intra-VTA hM3Dq DREADD expression (B, D, F) in three TH:Cre rats that showed no mCherry expression and little-to-no behavioral response to the chemogenetic ligands post-DREADD virus injections. DAPI staining is represented in blue, TH in red, and mCherry in green. Yellow represents colocalization of TH and mCherry.