ACU leads to long-lasting behavioral deficits
Mice treated with CB1 agonist during adolescence show significant long-lasting deficits in sensorimotor gating and hippocampal-dependent contextual learning in adulthood. These deficits were not observed in mice treated with the CB1 agonist during adulthood.
: Mice treated with WIN 55,212-2 during adolescence display significant deficits in PPI. There is a significant interaction of age (F=12.37, df (1,36), P
=0.0012), no effect of drug (F=0.44, df (1,36), P
=0.51) and a significant age × drug interaction (F=6.09, df (2,27), P
=0.018). Post hoc
analyses show that ACU mice show reduced PPI at prepulse intensities 4
=2.1, df 18, P
=0.049) and 8
=2.2, df 18, P
=0.041) above background. Adult-CU mice, however, do not show any significant change in PPI at any of the prepulse intensities (all t
between 0.6 and 1.6, all P
>0.13) ( and ).
Figure 2 Adolescent cannabinoid treatment (ACU) leads to long-term behavioral deficits. Mice were administered WIN 55,212-2 (white bars) or vehicle (black bars) for 10 days during adolescence (a–d) or adulthood (e–h) and tested behaviorally as (more ...)
Fear conditioning: Mice treated with WIN 55,212-2 during adolescence display significant deficits in contextual learning. There is a significant interaction of age (F=72, df (1,36), P<0.0001), drug (F=8.1, df (1,36), P=0.007) and a significant age × drug interaction (F= 8.1, df, p=0.007). Post hoc analyses show that ACU mice show reduced freezing compared with controls (t=2.85, df 18, P=0.011). Adult-CU mice, however, do not show any significant change in the contextual fear conditioning (t=0.27, df 18, p=0.78), Similarly, two-way analysis of variance conducted for cued fear conditioning shows a significant interaction of age (F=57.3, df (1,36), P<0.001), drug (F=5.4, df (1,36), P=0.03 ) and a significant age × drug interaction (F=5.6, df (1,36), P=0.02). Post hoc analyses show that ACU mice show reduced freezing compared with controls (t=2.34, df 18, P=0.031). Adult-CU mice, however, do not show any significant change in cued fear conditioning (t=1.22, df 18, P=0.24,). Mice treated with WIN 55,212-2 during adolescence for a shorter duration between 3–5 days did not display any fear-conditioning deficits in the contextual (t=0.62, df 38, P=0.54) or cued paradigms (t=0.83, df 38, P=0.41) ( and ).
Social interaction: There were no significant differences in interaction time between groups. There were no interactions of age (F=0.78, df (1,36), P=0.38), drug (F=0.22, P=0.64) or age × drug interaction (F=0.80, df (1,36), P=0.38) ( and ).
Locomotor activity: There were no significant differences in total locomotor activity. There were no interactions of age (F=1.54, df (1,36), P=0.22), drug (F=1.14, df (1,36), P=0.29) or age × drug interaction (F=0.002, df (1,36), P=0.96) ( and ).
mGluR5 is critically involved in fear conditioning with mGluR5 knockout mice showing impairments in acquisition of fear, deficits in the ability to extinguish contextual fear and poor performance in novelty detection,27, 28
all tasks that involve the hippocampus. We measured mGluR5 protein expression in the hippocampus of the ACU mice and found significant reductions in mGluR5 levels (t
=3.02, df 18, P
=0.007) (). In addition, mGluR5 receptor activation is one mechanism for activating eCB synthesis and release, suggesting that eCB signaling might be altered in ACU mice. We measured CB1, DGL (synthetic enzyme for 2-AG), MGL (metabolic enzyme for 2-AG) and FAAH (metabolic enzyme for AEA) protein levels in the hippocampus of the experimental mice. We found significant increases in both MGL (t
=4.2, df 13, P
=0.001) and FAAH (t
=0.014) but no changes in CB1 (t
=0.86) or DGL (t
=0.59) between groups. Lastly, we quantified norbin, an endogenous mGluR5 ligand,29
which was not altered in ACU mice (t
=0.35, df 16, P
Figure 3 Adolescent cannabinoid treatment (ACU) leads to altered expression of genes involved in endocannabinoid signaling in the hippocampus of ACU and control (CON) mice in adulthood (n=10 mice per group). (a) Levels of hippocampal cannabinoid receptor 1 (CB1), (more ...)
Given the critical role for mGluR5 in contextual fear conditioning, we examined correlations between contextual freezing and mGluR5 protein levels. In ACU mice, we find strong significant correlations (r=0.66, P=0.039) that are not seen in vehicle-treated mice (r=0.20, P=0.58). There were no correlations between freezing behavior and hippocampal CB1 levels in ACU mice (r=0.31, P=0.39) or controls (r=0.35, P=0.32) ().
Figure 4 Correlations between contextual freezing behavior and hippocampal mGluR5 protein levels in adolescent cannabinoid treatment (ACU) and control (CON) mice (n=10 mice per group). Siginificant correlations are seen between contextual freezing in ACU mice (more ...)