Proceedings of The Physiological Society

Physiology 2015 (Cardiff, UK) (2015) Proc Physiol Soc 34, PC112

Poster Communications

Dose dependent aversive effects of chemogenetic Locus coeruleus activation

S. Hirschberg1, Y. Li1, A. Randall2,1, A. E. Pickering1

1. Physiology and Pharmacology, University of Bristol, Bristol, Please Select, United Kingdom. 2. Medical School, University of Exeter, Exeter, United Kingdom.

The Locus coeruleus (LC) is the principle noradrenergic nucleus in the CNS. Dysfunction of the noradrenergic system has been associated with pathologies such as major depression, Alzheimer's disease and neuropathic pain. We have therefore been interested in developing means to produce long term activation of the noradrenergic system. We describe here the use of an engineered excitatory receptor-ionophore (PSAM) to manipulate LC activity (Magnus et al. 2011). We developed a lenti-viral vector (lenti-PRS-EGFP2aPSAM-HA) with catecholaminergic promoter (PRS) to allow expression of a traceable (HAtag) excitatory ion channel (PSAM-HA) in NAergic LC neurons. Expression was evaluated using immunofluorescence for EGFP and dopamine beta hydroxylase (DBH) after vector delivery into the LC of Wistar rats (P19-21). Cell properties and effects of agonist application were investigated using patch clamp recordings from acute pontine slices. To assay for behavioural effects Wistar rats (250-300g) were transduced with either lenti-PRS-EGFP2aPSAM-HA (N=11) or a control vector lenti-PRS-EGFP (N=7) and tested in a conditioned place preference (CPP) protocol with intraperitoneal (i.p.) dosing of 5mg/Kg and 10mg/Kg PSEM308 (selective agonist). After behavioural testing, transduced animals were dosed with PSEM308 (5mg/Kg i.p.), culled 2.5 hours later and pontine tissue processed for c-Fos expression. After LC injection of lenti-PRS-EGFP2aPSAM-HA, fluorescence (285+/-75 cells per LC, N=3) was restricted to DBH-positive neurons and PSAM-HA was only detected in membranes of EGFP-positive cells. Patch recordings showed PSEM308 increased spontaneous action potential firing frequency or reversibly depolarized LC neurons to firing threshold. Conditioning with 10mg/Kg caused aversion to the drug paired chamber in PSAM-HA transduced rats (N=8; P<0.05, Bonferoni post-test saline vs drug compartment, 20% difference; Interaction: P=0.052 F(1.14)=4.47 Two way repeated ANOVA) whilst 5mg/Kg (N=11) had no effect (and no effect was seen in control animals at either dose). The percentage of transduced neurons that were double labelled with c-Fos was 18.4+/-5.9% in EGFP and 29.8+/-9.4% in PSAM-HA transduced animals. Our findings show that chemogenetic LC activation causes a dose-dependent aversive effect. It remains to be determined whether targeting of subsets of LC neurons (such as the pontospinal group) will produce the same aversive behaviour.

Where applicable, experiments conform with Society ethical requirements