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. 2023 Mar;44(3):538-545.
doi: 10.1038/s41401-022-00987-3. Epub 2022 Sep 20.

Upregulation of dynorphin/kappa opioid receptor system in the dorsal hippocampus contributes to morphine withdrawal-induced place aversion

Yan Chen #  1 , Chen-Yao Wang #  2   3 , Gui-Ying Zan  2 , Song-Yu Yao  4 , Ying-Zhi Deng  2 , Xue-Lian Shu  2   3 , Wei-Wei Wu  2 , Yan Ma  2 , Yu-Jun Wang  5 , Chang-Xi Yu  6 , Jing-Gen Liu  2
Affiliations

Upregulation of dynorphin/kappa opioid receptor system in the dorsal hippocampus contributes to morphine withdrawal-induced place aversion

Yan Chen et al. Acta Pharmacol Sin. 2023 Mar.

Abstract

Aversive emotion of opioid withdrawal generates motivational state leading to compulsive drug seeking and taking. Kappa opioid receptor (KOR) and its endogenous ligand dynorphin have been shown to participate in the regulation of aversive emotion. In the present study, we investigated the role of dynorphin/KOR system in the aversive emotion following opioid withdrawal in acute morphine-dependent mice. We found that blockade of KORs before pairing by intracerebroventricular injection of KOR antagonist norBNI (20, 40 μg) attenuated the development of morphine withdrawal-induced conditioned place aversion (CPA) behavior. We further found that morphine withdrawal increased dynorphin A expression in the dorsal hippocampus, but not in the amygdala, prefrontal cortex, nucleus accumbens, and thalamus. Microinjection of norBNI (20 μg) into the dorsal hippocampus significantly decreased morphine withdrawal-induced CPA behavior. We further found that p38 MAPK was significantly activated in the dorsal hippocampus after morphine withdrawal, and the activation of p38 MAPK was blocked by pretreatment with norBNI. Accordingly, microinjection of p38 MAPK inhibitor SB203580 (5 μg) into the dorsal hippocampus significantly decreased morphine withdrawal-produced CPA behavior. This study demonstrates that upregulation of dynorphin/KOR system in the dorsal hippocampus plays a critical role in the formation of aversive emotion associated with morphine withdrawal, suggesting that KOR antagonists may have therapeutic value for the treatment of opioid withdrawal-induced mood-related disorders.

Keywords: conditioned place aversion; dorsal hippocampus; kappa opioid receptor; morphine withdrawal; norBNI; p38 MAPK.

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Conflict of interest statement

The authors declare no competing interests.

Figures

Fig. 1
Fig. 1. Conditioned place aversion was induced by naloxone-precipitated morphine withdrawal in mice.
a Timeline for the experimental procedure. b Conditioned place aversion was elicited by two pairing with 0.2 mg/kg naloxone in mice exposed to 20 mg/kg morphine. Data are expressed as mean ± SEM, **P < 0.01 compared with the saline-treated control group, one-way ANOVA with Bonferroni’s post hoc test. Sal Saline, Mor Morphine, Nal Naloxone.
Fig. 2
Fig. 2. Effects of norBNI by intracerebroventricular (icv) administration on conditioned morphine withdrawal-induced CPA behavior.
Left column. Timeline for norBNI icv injection before a and after c naloxone injection and pairing and behavioral testing. Right column. b Injection of norBNI before pairing attenuated CPA induced by morphine withdrawal. NorBNI (10 μg/μL, 20 μg/μL, 1 μL/side) or vehicle (1 μL/side) was bilaterally microinjected into icv 2 h before naloxone injection. d Injection of norBNI after pairing did not affect CPA induced by morphine withdrawal. NorBNI (20 μg/μL, 1 μL/side) or vehicle (1 μL/side) was bilaterally microinjected into icv 1 h after naloxone injection. Values are expressed as mean ± SEM, *P < 0.05 compared with the corresponding vehicle-treated CMW group, ##P < 0.01 compared with saline-treated control group, one-way ANOVA with Bonferroni’s post hoc test. CMW conditioned morphine withdrawal.
Fig. 3
Fig. 3. The expression levels of dynorphin A in different brain regions after morphine withdrawal.
The dorsal hippocampus (a), amygdala (b), prefrontal cortex (c), nucleus accumbens (d) and thalamus (e) were extracted 1 h after naloxone-precipitated morphine withdrawal. The protein level of dynorphin A was measured by Western blot. Values are expressed as mean ± SEM, *P < 0.05 compared with control group in unpaired two-tailed t-test. CMW conditioned morphine withdrawal.
Fig. 4
Fig. 4. Effects of norBNI by intra-dorsal hippocampal administration on morphine withdrawal-induced CPA behavior.
a Timeline for the norBNI injection and behavioral testing. b Intra-dorsal hippocampal injection of norBNI before pairing attenuated CPA induced by morphine withdrawal. NorBNI (20 µg/µL, 0.5 µL/side) or vehicle (0.5 µL/side) was bilaterally microinjected into the dorsal hippocampus 2 h before naloxone injection. c Mice locomotor activity was measured. Nissl staining (d) and schematic representation of injection sites (e ○, control; ●, vehicle+CMW; ▲, norBNI+CMW) in the dorsal hippocampus for mice used in the experiments. Values are expressed as mean ± SEM, ***P < 0.001 compared with the saline-treated control group, #P < 0.05 compared with vehicle-treated CMW group, one-way ANOVA with Bonferroni’s post hoc test. CMW conditioned morphine withdrawal.
Fig. 5
Fig. 5. KOR dependent p38 MAPK activation was involved in the CPA produced by morphine withdrawal.
a p38 MAPK was activated in the dorsal hippocampus after morphine withdrawal. b Intra-dorsal hippocampal injection with norBNI attenuated p38 MAPK activation. Effects of p38 MAPK inhibitor SB203580 on morphine withdrawal-induced p38 MAPK activation and CPA behavior. c Timeline for SB203580 injection and behavioral testing. d, e Intra-dorsal hippocampal injection of SB203580 before pairing attenuated p38 MAPK activation and CPA induced by morphine withdrawal. SB203580 (5 µg/µL, 0.5 µL/side) or vehicle (0.5 µL/side) was bilaterally microinjected into the dorsal hippocampus 1 h before naloxone injection. f Mice locomotor activity was measured. Nissl staining (g) and schematic representation of injection sites (h ○, control; ●, vehicle + CMW; ▲, SB203580 + CMW) in the dorsal hippocampus for mice used in the experiments. Values are expressed as mean ± SEM, *P < 0.05, ***P < 0.001 compared with saline-treated control group, #P < 0.05, ##P < 0.01 compared with vehicle-treated CMW group, one-way ANOVA with Bonferroni’s post hoc test. CMW conditioned morphine withdrawal.
Fig. 6
Fig. 6. Effects of norBNI and SB203580 in the dorsal hippocampus on the novel object recognition test task.
a Experimental scheme. b, e Exploration time during the training phase. c, f Exploration time during the test phase. Microinjection of norBNI (20 µg/µL, 0.5 µL/side) or vehicle (0.5 µL/side) into the dorsal hippocampus 2 h before test. Microinjection of SB203580 (5 µg/µL, 0.5 µL/side) or vehicle (0.5 µL/side) into the dorsal hippocampus 1 h before test. All groups show more significant preference for the new object than the familiar object. d, g Discrimination indexes are plotted as a function of the drugs. There are no differences in discrimination index between vehicle-treated and norBNI-treated or SB203580-treated group. Values are expressed as mean ± SEM, *P < 0.05 compared with vehicle-F, norBNI-F, or SB203580-F group, two-way ANOVA with Bonferroni post hoc tests. F Familiar, N Novel.
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