Maturation of coordinated immediate early gene expression by cocaine during adolescence

Abstract

Adolescence may be a critical period for drug addiction. Young adolescent male rats have greater locomotor responses than adults after acute low dose cocaine administration. Further, repeated cocaine administration produces as much or more conditioned place preference but reduced locomotor sensitization in adolescents compared to adults. Acute activation of neurons by cocaine induces long-term changes in behavior by activating transcriptional complexes. The purpose of the present study was to correlate cocaine-induced locomotor activity with neuronal activation in subregions of the striatum and cortex by acute cocaine in young adolescent (postnatal (PN) 28) and adult (PN 65) male rats by measuring the induction of the plasticity-associated immediate early genes (IEGs) c-fos and zif268 using in situ hybridization. Animals were treated with saline, low (10 mg/kg), or high (40 mg/kg) dose cocaine in locomotor activity chambers and killed 30 min later. Low dose cocaine induced more locomotor activity and striatal c-fos expression in adolescents than adults whereas high dose cocaine induced more locomotor activity, striatal c-fos, and striatal zif268 expression in adults. Locomotor activity correlated with the expression of both genes in adults but correlated with striatal c-fos only in adolescents. Finally, there was a significant correlation between the expression of c-fos and zif268 in the adult striatum but not in adolescents. Our results suggest that the coordinated expression of transcription factors by cocaine continues to develop during adolescence. The immature regulation of transcription factors by cocaine could explain why adolescents show unique sensitivity to specific long-term behavioral alterations following cocaine treatment.

Figure 1. Regions analyzed

Schematic diagrams of the 29 subregions measured in four rostral-caudal coronal sections. In the most rostral section (+3.20 mm) we measured mRNA levels in the cingulate (CIN) and orbital cortex (ORB). We measured c-fos and zif268 mRNA levels from the same five cortical regions in each of the three remaining sections: the dorsal agranular cortex (AG), dorsal anterior cingulate (CD), ventral anterior cingulate (CV), sensorimotor cortex (SM), and the agranular insular cortex (AI). Cortical regions are indicated in the left hemispheres. We measured mRNA levels in 12 subregions of the striatum indicated in the right hemispheres. In the most rostral section corresponding to +1.60 mm, we measured mRNA levels in the dorsal caudate (DC), medial caudate (MC), lateral caudate (LC), nucleus accumbens core (C), medial nucleus accumbens shell (MS), and ventral nucleus accumbens shell (VS). In the middle section (+0.40 mm) we measured the dorsal caudate (DC), lateral caudate (LC), ventromedial caudate (MVC), and the ventrolateral caudate (LVC). In the caudal section (-0.80, bottom image) we measured in the dorsal caudate (DC) and the ventrolateral caudate (LVC).

Figure 2. Locomotor responses to cocaine

Locomotor responses in adolescents (filled symbols) and adults (open symbols) following saline (a), 10 mg/kg cocaine (b), or 40 mg/kg cocaine (c) are plotted against time. Symbols represent mean +/- SEM. * indicates P<0.05 greater than PN 65. ** indicates P<0.05 greater than PN 28. N=6-15 for each age × treatment group. Cocaine dose-dependently increased locomotor activity in adolescents and adults. Locomotion increased more in adolescents after 10 mg/kg and more in adults after 40 mg/kg.

Figure 3. Cocaine-induced increases in striatal c-fos expression

Top panels show the percent increase in c-fos expression (compared to saline) in adolescents (filled bars) and adults (open bars) after 10 mg/kg cocaine (a) or 40 mg/kg cocaine (b). The x axes indicate specific striatal regions abbreviated RDC (rostral dorsal caudate), RMC (rostral medial caudate), RLC (rostral lateral caudate), core (nucleus accumbens core), mshell (medial nucleus accumbens shell), vshell (ventral nucleus accumbens shell), MDC (medial dorsal caudate), MLC (medial lateral caudate), VMC (ventromedial caudate), and MVLC (medial ventrolateral caudate), CDC (caudal dorsal caudate), and CVLC (caudal ventrolateral caudate). Dashed lines separate regions measured in different rostral to caudal sections. *indicates PN 28 significantly greater than PN 65 in designated regions. **indicates PN 65 significantly greater than PN 28 in designated regions. Representative images from the most rostral section (+1.60 mm) from adolescents (top) and adults (bottom) are shown in (c). N=6-15 for each age × treatment group. Cocaine increased c-fos more in adolescents than adults in dorsal striatum after 10 mg/kg, and increased c-fos more in dorsal striatum after 40 mg/kg in adults than adolescents.

Figure 4. Cocaine-induced increases in striatal zif268 expression

Top panels show the percent increase in zif268 expression (compared to saline) in adolescents (filled bars) and adults (open bars) after 10 mg/kg cocaine (a) or 40 mg/kg cocaine (b). The x axes indicate specific striatal regions abbreviated RDC (rostral dorsal caudate), RMC (rostral medial caudate), RLC (rostral lateral caudate), core (nucleus accumbens core), mshell (medial nucleus accumbens shell), vshell (ventral nucleus accumbens shell), MDC (medial dorsal caudate), MLC (medial lateral caudate), VMC (ventromedial caudate), and MVLC (medial ventrolateral caudate), CDC (caudal dorsal caudate), and CVLC (caudal ventrolateral caudate). Dashed lines separate regions measured in different rostral to caudal sections. **indicates significantly greater than PN 28 in designated regions. Representative images from the most rostral section (+1.60 mm) from adolescents (top) and adults (bottom) are shown in (c). N=6-15 for each age × treatment group. Cocaine increased zif268 comparably in adolescents and adults in all areas after 10 mg/kg, and increased zif268 more in adults than adolescents in all areas after 40 mg/kg.

Figure 5. Effects of cocaine on cortical c-fos and zif268 expression

Top panels show the percent increase in c-fos expression (compared to saline) in adolescents (filled bars) and adults (open bars) following 10 mg/kg cocaine (a) or 40 mg/kg cocaine (b). Bottom panels show the percent increase in zif268 expression following 10 mg/kg cocaine (c) or 40 mg/kg cocaine (d). The x axes indicate specific cortical regions abbreviated CING (cingulate), ORB, (orbital cortex), AG (dorsal agranular cortex), ACD (dorsal anterior cingulate), ACV (ventral anterior cingulate), SM (sensorimotor cortex), and AI (agranular insular cortex). **indicates significantly greater than PN 28 in designated regions. N=6-15 for each age × treatment group. Cocaine increased c-fos comparably in adolescents and adults in all cortical areas, and increased zif268 more in ag, acd, acv, sm and ai in adults than in adolescents after 40 mg/kg.

Figure 6. Correlation between striatal c-fos and zif268 expression and locomotor activity

Correlations in adolescents are shown in (a) and adults are shown in (b). Open symbols represent animals treated with 10 mg/kg cocaine and closed symbols represent animals treated with 40 mg/kg cocaine. Regression lines are shown for significant correlations (P<0.05). N=6-15 for each age × treatment group. Cocaine-stimulated locomotion correlated with c-fos induction in both adolescents and adults but correlated with zif268 induction only in adults.

Figure 7. Correlation between striatal c-fos and zif268 mRNA levels

Correlations in adolescents are shown in (a) and adults are shown in (b). Open symbols represent animals treated with 10 mg/kg cocaine. Closed symbols represent animals treated with 40 mg/kg cocaine. Regression lines are shown for when significant correlations were observed. N=6-10 for each age × treatment group. Zif268 induction correlated with c-fos induction only in adults.

Figure 8. Time course of c-fos and zif268 induction by high dose cocaine in the adolescent and adult striatum

Cocaine induced a similar time course of expression for both c-fos (a) and zif268 (b) in young adolescent and adult male rats. Closed symbols represent young adolescents and open symbols represent adults. **indicates greater in PN 65 than PN 28. N=5-8 for all age × time groups. Induction of both c-fos and zif268 were greater in adults than adolescents both at 30 min and 60 min.