Maternal deprivation increases vulnerability to morphine dependence and disturbs the enkephalinergic system in adulthood

Abstract

Maternal deprivation can trigger long-lasting molecular and cellular modifications in brain functions and might facilitate the appearance of pathogenic behaviors. This study focuses on the vulnerability to develop morphine dependence in adult rats that were separated from their mother and littermates for 3 h per day for 14 d after birth and examines the adaptive changes in the enkephalinergic pathways. Place-preference conditioning was observed with 2 mg/kg morphine in deprived rats, whereas 5 mg/kg morphine was necessary to induce conditioning in nondeprived animals. A prolonged morphine conditioning was shown in deprived rats. A strong increase in oral morphine self-administration behavior and preference was observed in deprived rats. Only a very slight increase in preference for sucrose solution, a more ethological reinforcer known to interact with the opioid system, was shown in deprived rats. These results indicate that this postnatal environment change leads to a hypersensitivity to the reinforcing properties of morphine and to the development of morphine dependence. A significant decrease in preproenkephalin mRNA expression was observed in the nucleus accumbens and the caudate-putamen nucleus of deprived rats. The basal extracellular levels of the Met-enkephalin-like immunoreactivity in the nucleus accumbens were significantly lower in deprived rats when compared with nondeprived animals, whereas no change in mu-opioid receptor binding occurred. These results strongly support that maternal deprivation leads to a basal hypoactivity of the enkephalinergic system and hypersensitivity to morphine effects. Together, our results suggest that maternal deprivation in pups likely represents a risk factor for morphine dependence in adult rats.

Figure 1.

Effects of 2 mg/kg morphine (ND, n = 14; D, n = 12) and 5 mg/kg morphine (intraperitoneally; ND, n = 20; D, n = 20) on the expression [day 8 (D8)] and extinction (D9 and D10) periods of the place-preference paradigm in ND and D rats. During the place-conditioning period, rats received morphine on days 1, 3, and 5 and saline on days 2, 4, and 6 immediately before confinement in the associated compartment. Rats were tested at 2.5-3 months of age. The results are expressed as a score (s), calculated as the difference between the postconditioning and preconditioning times spent in the compartment associated with the drug (ANOVA for 2 mg/kg; deprivation: F_(1,22) = 1.08; p = 0.30; treatment: F_(1,22) = 5.07; p = 0.03; interaction: F_(1,22) = 4.83; p = 0.03; time: F_(2,44) = 1.91; p > 0.05; interactions time-deprivation, time-treatment, and time-deprivation plus treatment, p > 0.05; ANOVA for 5 mg/kg; deprivation: F_(1,36) = 5.04; p = 0.03; treatment: F_(1,36) = 12.28; p = 0.001; interaction: F_(1,36) = 4.26; p = 0.04; time: F_(2,72) = 5.3; p = 0.007; interactions time-deprivation, time-treatment, and time-deprivation plus treatment, p > 0.05). Error bars represent SEM. ^*p < 0.05, ^***p < 0.001 versus the respective saline group; ⁺p < 0.05, ⁺⁺p < 0.01 versus 2 or 5 mg/kg morphine in the ND group (Newman-Keuls test).

Figure 2.

Oral morphine (25 mg/L) self-administration behavior using the two-bottle-choice paradigm in ND (n = 9) and D (n = 9) rats for 12 weeks. A, Morphine solution consumption. B, Morphine preference. The results are expressed as mean. (1), Habituation phase (consumption, deprivation: F_(1,16) = 1.62; p > 0.05; time: F_(5,80) = 7.9; p < 0.0001; interaction: F_(5,80) = 0.217; p > 0.05; preference, deprivation: F_(1,16) = 3.13; p > 0.05; time: F_(5,80) = 9.19; p < 0.0001; interaction: F_(5,80) = 0.16; p > 0.05); (2), installation phase (consumption, deprivation: F_(1,16) = 5.64; p = 0.03; time: F_(19,304) = 8.55; p < 0.0001; interaction: F_(19,304) = 1.64; p = 0.04; preference, deprivation: F_(1,16) = 5.44; p = 0.03; time: F_(19,304) = 5.49; p < 0.0001; interaction: F_(19,304) = 2.0; p = 0.008); (3), plateau phase (consumption, deprivation: F_(1,16) = 6.37; p = 0.02; time: F_(25,400) = 3.39; p < 0.0001; interaction: F_(25,400) = 0.76; p > 0.05; preference, deprivation: F_(1,16) = 5.73; p = 0.03; time: F_(25,400) = 2.49; p < 0.0001; interaction: F_(25,400) = 1; p > 0.05). ^*p < 0.05, ^**p < 0.01 versus the ND group (Newman-Keuls test).

Figure 3.

Oral morphine (25 mg/L) self-administration behavior using the two-bottle-choice paradigm in ND (n = 9) and D (n = 9) rats for 12 weeks. The long-term course of morphine solution consumption was subdivided into three phases: habituation, installation, and plateau. Means ± SEM within each of the three periods were calculated to smooth daily drinking variability. Error bars represent SEM. Two-way repeated-measures ANOVA showed significant effects of deprivation, time factors, and interaction (consumption, deprivation: F_(1,16) = 7.89; p = 0.01; time: F_(2,32) = 6.65; p = 0.003; interaction: F_(2,32) = 3.85; p = 0.03; preference, deprivation: F_(1,16) = 7.06; p = 0.01; time: F_(2,32) = 14.18; p < 0.0001; interaction: F_(2,32) = 3.71; p = 0.03). ^*p < 0.05 versus the ND group; ⁺p < 0.05 versus the D habituation phase (Newman-Keuls test).

Figure 4.

Percentage of ND and D rats showing a preference of >50% for the morphine solution during the habituation, installation, and plateau phases. ^***p < 0.001 versus the ND group (χ² test).

Figure 5.

Oral sucrose (0.025%) self-administration behavior using the two-bottle-choice paradigm in ND (n = 8) and D (n = 6) rats for 90 d. The results are expressed as means of preference (ANOVA; deprivation: F_(1,12) = 5.69; p = 0.03; time: F_(53,636) = 1.40; p = 0.03; interaction: F_(53,636) = 0.64; p > 0.05). ^*p < 0.05 versus the ND group (Newman-Keuls test).

Figure 6.

PPE mRNA hybridization signals in the rostral (R), core (Cr), cone (Cn), and shell (Sh) parts of the N.Acc.; in the anterior (A) and posterior (p) caudate-putamen nucleus (CPu); and in the central (Ce) and basolateral (BL) nucleus of the amygdala (Amyg) of ND (n = 5) and D (n = 7) rats. Rats were killed at 3 months of age. Values correspond to mean optical density × 10³± SEM. Error bars represent SEM. ^*p < 0.05, ^**p < 0.01 versus the ND group (Fisher's PLSD test).

Figure 7.

Autoradiograms of frontal brain sections showing the distribution of PPE mRNA expression in the striatum of ND and D rats.

Figure 8.

Autoradiograms of frontal brain sections showing the distribution of μ-opioid receptor binding ([³H]-DAGO) in the striatum and the mesencephalon of ND and D rats.

Figure 9.

Extracellular levels of Met-LI in the N.Acc. of ND (n = 5; ▪) and D (n = 6; □) rats. Values correspond to mean ± SEM (pg/60 μl; ANOVA; deprivation: F_(1,9) = 5.25; p < 0.05; time: F_(3,27) = 1.12; p > 0.05; interaction: F_(3,27) = 0.90; p > 0.05). Error bars represent SEM. ^*p < 0.05 versus the ND group (Newman-Keuls test).