Impaired emotional-like behavior and serotonergic function during protracted abstinence from chronic morphine

Abstract

Background: Opiate abuse is a chronic relapsing disorder, and maintaining prolonged abstinence remains a major challenge. Protracted abstinence is characterized by lowered mood, and clinical studies show elevated comorbidity between addiction and depressive disorders. At present, their relationship remains unclear and has been little studied in animal models. Here we investigated emotional alterations during protracted abstinence, in mice with a history of chronic morphine exposure.

Methods: C57BL6J mice were exposed to a chronic intermittent escalating morphine regimen (20-100 mg/kg). Physical dependence (naloxone-precipitated withdrawal), despair-related behaviors (tail suspension test), and social behaviors were examined after 1 or 4 weeks of abstinence. Stress hormones and forebrain bioamine levels were analyzed at the end of morphine regimen and after 4 weeks of abstinence. Finally, we examined the effects of chronic fluoxetine during abstinence on morphine-induced behavioral deficits.

Results: Acute naloxone-induced withdrawal was clearly measurable after 1 week, and became undetectable after 4 weeks. In contrast, social and despair-related behaviors were unchanged after 1 week, but low sociability and despair-like behavior became significant after 4 weeks. Chronic morphine regimen increased both corticosterone levels and forebrain serotonin turnover, but only serotonergic activity in the dorsal raphe remained impaired after 4 weeks. Remarkably, chronic fluoxetine prevented depressive-like behavioral deficits in 4-week abstinent mice.

Conclusions: During protracted abstinence, the immediate consequences of morphine exposure attenuate, whereas fluoxetine-sensitive emotional alterations strengthen with time. Our study establishes a direct link between morphine abstinence and depressive-like symptoms and strongly suggests that serotonin dysfunction represents a main mechanism contributing to mood disorders in opiate abstinence.

Figure 1

Physical dependence and HPA axis hyperactivity decline during protracted abstinence. (A) Time line of the experiment. Dashed arrows indicate morphine doses injected twice daily. Black arrows indicate chronic, 1-week and 4-week abstinence time points under study. Morphine treatment reduced body weight over time (2 experiments pooled, n=14/group). (B) Withdrawal signs following naloxone injection (s.c. 1 mg/kg) 2 h after the last morphine administration (n=4 mice/group). (C) Withdrawal signs measured following morphine re-exposure (50 mg/kg) after either 1 or 4 weeks of abstinence (n=5/group). Physical symptoms decreased after 4 weeks. (D) Corticosterone and ACTH levels measured in chronic and 4-week abstinent groups (n=5-6/group). HPA axis was activated after chronic treatment but not after 4 weeks. Values are mean ± sem. *p<0.05; **p<0.01; ***p<0.0001 morphine versus saline controls. ^#p<0.05 comparison between 1-week and 4-week abstinence groups within each treatment condition.

Figure 2

Social interaction deficits and depressive-like behavior develop during protracted abstinence. (A) Social interaction test. Social behaviors decreased after 4 weeks of abstinence. (B) Tail suspension test. Immobility duration increased after 4 weeks abstinence. (C, D) Open-field test. Morphine abstinence had no effect on locomotor activity (C) or anxiety-like behavior (D). Values are mean ± sem. *p<0.05; **p<0.01 morphine versus saline controls. ^#p<0.05; ^##p<0.01 comparison between 1-week and 4-week abstinence groups within each treatment condition (n=8/group).

Figure 3

5-HT turnover is increased in the dorsal raphe after protracted abstinence. 5-HT turnover, defined as the 5-HIAA/5-HT ratio, is shown. Chronic morphine treatment significantly increased 5-HT turnover in the prefrontal cortex (A), central amygdala (B) and hippocampus (C) but not in the dorsal raphe (D). After 4 weeks of abstinence, 5-HT turnover was increased specifically in the dorsal raphe. Values represent mean ± sem. **p<0.01; ***p<0.001 morphine versus saline controls. (n=5-6/group).

Figure 4

Fluoxetine prevents the development of depressive-like and social deficits. (A, B) Social interaction test. Fluoxetine prevented protracted morphine effects on social (A) and individual (B) behaviors. (C, D) Tail suspension test. Fluoxetine also normalized passive (C) and active (D) behaviors in morphine abstinent animals. Values represent mean ± sem. *p<0.05; **p<0.01 morphine versus saline controls. ^#p<0.05; ^##p<0.01 comparison between morphine/chow and morphine/FLX groups. (n=19-20/group).