A locus coeruleus to dorsal hippocampus pathway mediates cue-induced reinstatement of opioid self-administration in male and female rats

Abstract

Opioid use disorder is a chronic relapsing disorder encompassing misuse, dependence, and addiction to opioid drugs. Long term maintenance of associations between the reinforcing effects of the drug and the cues associated with its intake are a leading cause of relapse. Indeed, exposure to the salient drug-associated cues can lead to drug cravings and drug seeking behavior. The dorsal hippocampus (dHPC) and locus coeruleus (LC) have emerged as important structures for linking the subjective rewarding effects of opioids with environmental cues. However, their role in cue-induced reinstatement of opioid use remains to be further elucidated. In this study, we showed that chemogenetic inhibition of excitatory dHPC neurons during re-exposure to drug-associated cues significantly attenuates cue-induced reinstatement of morphine-seeking behavior. In addition, the same manipulation reduced reinstatement of sucrose-seeking behavior but failed to alter memory recall in the object location task. Finally, intact activity of tyrosine hydroxylase (TH) LC-dHPC_Th afferents is necessary to drive cue induced reinstatement of morphine-seeking as inhibition of this pathway blunts cue-induced drug-seeking behavior. Altogether, these studies show an important role of the dHPC and LC-dHPC_Th pathway in mediating cue-induced reinstatement of opioid seeking.

Fig. 1. Inhibition of excitatory dHPC neurons attenuates cue-induced reinstatement of morphine-seeking.

A Schematic representation of viral injection and behavioral methodology. B Number of active and inactive lever presses during morphine SA acquisition (n = 56). C Number of active and inactive lever presses steadily decreased over the 10 days of extinction (n = 56). D Number of active lever presses during the last day of extinction and cue-induced reinstatement. Chemogenetic inhibition of dHPC excitatory neurons during reinstatement attenuates cue-driven drug-seeking. E Spread of overlay of individual animal viral expression across dHPC in hM4D(Gi) and control mCherry injected rats. F Representative traces and images from whole-cell current clamp recordings showing increased rheobase in hM4D(Gi) expressing neuron upon application of CNO in dHPC (left mCherry, right hM4D(Gi); top baseline, bottom CNO). G Coronal section of dHPC CaMKII neurons expressing hM4D(Gi). (Blue – DAPI; Red – mCherry (hM4D(Gi)). The data are presented as the mean ± s.e.m.

Fig. 2. Inhibition of excitatory dHPC neurons does not alter memory recall during object location test.

A Schematic representation of viral injection and behavioral methodology. Discrimination index is calculated as time spent with the moved object/time spent with both objects. B Discrimination index for the moved object during training and test session. Chemogenetic inhibition of dHPC excitatory neurons during test session does not affect the preference for moved object. C Spread of overlay of individual animal viral expression across dHPC in hM4D(Gi) and control mCherry injected rats. The data are presented as the mean ± s.e.m.

Fig. 3. Inhibition of excitatory dHPC neurons attenuates cue-induced reinstatement of sucrose-seeking.

A Schematic representation of viral injections and behavioral methodology. B Number of active and inactive lever presses during sucrose acquisition (n = 27). C Number of active and inactive lever presses steadily decreased over the 10 days of extinction (n = 27). D Number of active lever presses during the last day of extinction and cue-induced reinstatement. Chemogenetic inhibition of dHPC excitatory neurons during reinstatement prevents cue-driven sucrose-seeking. E Spread of overlay of individual animal viral expression across dHPC in hM4D(Gi) and control mCherry injected rats. F Representative coronal section of dHPC CaMKII neurons expressing hM4D(Gi). (Blue – DAPI; Red – mCherry (hM4D(Gi)). The data are presented as the mean ± s.e.m.

Fig. 4. Inhibition of LC-dHPC_Th pathway diminishes cue-induced reinstatement of drug-seeking.

A Schematic representation of viral injection and behavioral methodology. B Representative coronal section of LC TH neurons expressing hM4D(Gi). (Blue – DAPI; Red – mCherry (hM4D(Gi); Green – TH). Expression of mCherry is highly specific to TH positive neurons. C Number of active and inactive lever presses during morphine acquisition (n = 31). D Number of active and inactive lever presses steadily decreased over the 10 days of extinction (n = 31). E Number of active lever presses during the last day of extinction and cue-induced reinstatement. Chemogenetic inhibition of LC-dHPC_Th pathway during reinstatement attenuates cue-driven drug-seeking. F Spread of overlay of individual animal viral expression across LC in hM4D(Gi) and control mCherry injected Th cre rats. G Individual animal cannula placement in dHPC for microinjection of aCSF/CNO. H Representative traces and images from whole-cell current clamp recordings showing increased rheobase in hM4D(Gi) expressing neuron upon application of CNO in LC (left mCherry, right hM4D(Gi); top baseline, bottom CNO). The data are presented as the mean ± s.e.m.