Prenatal L-DOPA exposure produces lasting changes in brain dopamine content, cocaine-induced dopamine release and cocaine conditioned place preference

Abstract

Dopamine, its receptors and transporter are present in the brain beginning from early in the embryonic period. Dopamine receptor activation can influence developmental events including neurogenesis, neuronal migration and differentiation raising the possibility that dopamine imbalance in the fetal brain can alter development of the brain and behavior. We examined whether elevated dopamine levels during gestation can produce persisting changes in brain dopamine content and dopamine-mediated behaviors. We administered L-3,4-dihydroxyphenylalanine (L-DOPA) in drinking water to timed-pregnant CD1 mice from the 11th day of gestation until the day of parturition. The prenatal L-DOPA exposure led to significantly lower cocaine conditioned place preference, a behavioral test of reward, at postnatal day 60 (P60). However, in vivo microdialysis measurements showed significant increases in cocaine-induced dopamine release in the caudate putamen of P26 and P60 mice exposed to L-DOPA prenatally, ruling out attenuated dopamine release in the caudate putamen as a contributor to decreased conditioned place preference. Although dopamine release was induced in the nucleus accumbens of prenatally L-DOPA exposed mice at P60 by cocaine, the dopamine release in the nucleus accumbens was not significantly different between the L-DOPA and control groups. However, basal dopamine release was significantly higher in the prenatally L-DOPA exposed mice at P60 suggesting that the L-DOPA exposed mice may require a higher dose of cocaine for induction of cocaine place preference than the controls. The prenatal L-DOPA exposure did not alter cocaine-induced locomotor response, suggesting dissociation between the effects of prenatal L-DOPA exposure on conditioned place preference and locomotor activity. Tissue concentration of dopamine and its metabolites in the striatum and ventral midbrain were significantly affected by the L-DOPA exposure as well as by developmental changes over the P14-P60 period. Thus, elevation of dopamine levels during gestation can produce persisting changes in brain dopamine content, cocaine-induced dopamine release and cocaine conditioned place preference.

Figure 1

A diagrammatic representation of the experimental design. Timed-pregnant CD1 mice were assigned to 3 experimental groups beginning on the 11^th day of pregnancy: The mice in the L-DOPA group received drinking water containing L-DOPA (2 mg/ml) plus ascorbic acid (0.025%); the mice in the Ascorbic Acid group received drinking water containing 0.025% ascorbic acid; whereas the mice in the Water group received plain drinking water (i.e. drinking water without additives). On the day of parturition and during the nursing period the mice in all the 3 groups received the plain drinking water. Each of the 3 groups of mice was subjected to 3 types of experiments: 1. On postnatal day 60 (P60), we performed cocaine conditioned place preference and cocaine-induced locomotor activity assays; 2. On P26 and P60, we performed in vivo microdialysis to assay basal (un-stimulated) and cocaine-induced dopamine release in the striatum (P26), caudate putamen (P60) and nucleus accumbens (P60); and 3. On P14, P21 and P60 we analyzed tissue concentrations of dopamine, DOPAC and HVA in the striatum (caudate putamen and nucleus accumbens) and the midbrain using HPLC.

Figure 2

Cocaine conditioned place preference (A) and cocaine-induced locomotor activity (B) assays were performed in P60 mice exposed to drinking water containing L-DOPA plus ascorbic acid (L-DOPA), ascorbic acid alone (ASC) or plain drinking water (Water) from embryonic day 11 until the day of birth. The mice in the Water and ASC groups showed significant place preference by spending significantly longer periods of time in the cocaine-paired chamber during the test session compared to the preconditioning session (A). However, the mice in the L-DOPA group did not show place preference (A). Mice in all the 3 groups showed comparable locomotor activity upon a single intraperitoneal cocaine administration (B). * = p<0.05; ANOVA and Post-hoc Fisher’s PLSD test.

Figure 3

Microdialysis and HPLC analysis was performed in postnatal day 26 (P26) and P60 mice exposed to drinking water containing L-DOPA plus ascorbic acid (LDOPA), ascorbic acid alone (ASC) or plain drinking water (Water) from embryonic day 11 until the day of birth. Microdialysis probes were placed under stereotaxic guidance in the caudate putamen in awake, P26 and P60 mice and the nucleus accumbens in awake, P60 mice. The analysis in the nucleus accumbens was performed only in the L-DOPA and ASC groups. Samples were collected every 10 min for HPLC analysis. Following 30 min of sample collection for analysis of basal levels of dopamine and metabolites, cocaine (15 mg/kg) was administered intraperitoneally (arrows in A, B and C) and samples were collected at 10 min intervals for an additional 1 hr. The data showed that mice in all 3 groups showed a significant increase in dopamine release for ~ 30 min following the cocaine administration in the caudate putamen on P26 (A) and P60 (B) and nucleus accumbens on P60 (C). The increase in cocaine-induced dopamine release was significantly higher (* = p<0.05; ANOVA) in the L-DOPA group in the caudate putamen on P26 (A) and P60 (B). However, in the nucleus accumbens the dopamine release was not significantly different in the L-DOPA group compared to the ASC group.

Figure 4

Dopamine release prior to stimulation with cocaine (basal release) was analyzed in postnatal day 26 (P26) and P60 mice exposed to drinking water containing L-DOPA plus ascorbic acid (L-DOPA), ascorbic acid alone (ASC) or plain drinking water (Water) from embryonic day 11 until the day of birth. The analysis was performed by HPLC in samples collected from the caudate putamen on P26 (A) and P60 (B) and the nucleus accumbens on P60 (C) in awake mice by using microdialysis. The analysis in the nucleus accumbens was performed only in the L-DOPA and ASC mice. There was no statistically significant difference in dopamine release among the 3 groups of mice in the caudate putamen on P26 or P60. However, the dopamine release was significantly higher in the nucleus accumbens of L-DOPA mice compared to the ASC mice (C). * = P<0.05.

Figure 5

Tissue concentrations of dopamine (A) and its metabolites 3,4 dihydroxyphenylacetic acid (DOPAC; B) and 3-methoxy-4-hydroxyphenyl acetic acid (homovanillic acid, HVA; C) were measured in the striatum (caudate putamen plus nucleus accumbens) by HPLC in postnatal day 14 (P14), P21 and P60 mice exposed to drinking water containing L-DOPA plus ascorbic acid (L-DOPA), ascorbic acid alone (ASC) or plain drinking water (Water) from embryonic day 11 until the day of birth. The prenatal treatment significantly affected the concentrations of dopamine and DOPAC, with the L-DOPA group showing higher dopamine levels and lower DOPAC levels. The HVA concentration was unaffected by the prenatal treatment. The developmental stage also produced significant effects on all 3 measurements such that dopamine and DOPAC concentrations increased while the HVA concentration decreased with age.

Figure 6

Tissue concentrations of dopamine (A) and its metabolites 3,4 dihydroxyphenylacetic acid (DOPAC; B) and 3-methoxy-4-hydroxyphenyl acetic acid (homovanillic acid, HVA; C) were measured in ventral midbrain by HPLC in postnatal day 14 (P14), P21 and P60 mice exposed to drinking water containing L-DOPA plus ascorbic acid (L-DOPA), ascorbic acid alone (ASC) or plain drinking water (Water) from embryonic day 11 until the day of birth. The prenatal treatment significantly affected the concentrations of dopamine and HVA, with the L-DOPA group showing higher dopamine and lower HVA concentrations. Developmental stage also significantly affected all 3 measurements such that dopamine and DOPAC concentrations increased while the HVA concentration decreased with age.