Extracellular dopamine levels are lower in the medial prefrontal cortex of alcohol-preferring rats compared to Wistar rats

Abstract

Previous studies have identified deficiencies in the mesocorticolimbic dopamine (DA) systems of alcohol-preferring (P) rats. This study uses quantitative microdialysis to compare the extracellular levels of DA in the medial prefrontal cortex (MPF) of P rats and outbred Wistar rats and also compares the effects of systemic ethanol administration on DA levels in the MPF using traditional microdialysis. In experiment 1, male Wistar and P rats were implanted with loop-style microdialysis probes and later perfused at 0.5 μl/min with artificial cerebrospinal fluid for 120 min prior to five baseline (20-min) sample collections. Three concentrations (5, 10, and 20 nM) of DA were then perfused in random order for 100 min each. Samples (20-min) were collected and stored at −70°C until assayed using high performance liquid chromatography/electrochemical detection (HPLC/EC), and the data were analyzed using the quantitative no-net-flux (NNF) method. In experiment 2, male Wistar and P rats were implanted with dialysis probes aimed at the MPF. After collecting four baseline samples, all rats were injected (i.p.) with one dose of either 0.9% saline or 2.0 g/kg ethanol. Microdialysis samples were collected at 20-min intervals and stored at −70°C until analyzed by HPLC/EC. NNF microdialysis yielded significantly (P < .05) lower extracellular DA concentrations in the MPF of P rats compared to Wistar rats (2.0 ± 0.4 vs. 4.8 ± 0.4 nM, respectively). The extraction fractions were not different between the P and Wistar groups (69 ± 3 vs. 65 ± 3%, respectively). No significant change in extracellular DA levels was observed in P rats or Wistar rats after either saline or 2 g/kg ethanol. The lower extracellular concentrations of DA in the MPF of P rats compared to Wistar rats, without a difference in the extraction fraction, suggest that DA neurotransmission is lower in the MPF of the P rat. This lower DA neurotransmission could be a result of reduced activity of the DA neurons projecting to the MPF, reduced excitatory or increased inhibitory tone occurring locally within the MPF, and/or reduced DA innervation to the MPF. The lack of effect of systemic EtOH administration on extracellular DA levels in the MPF suggests that unlike the mesolimbic DA system, the mesocortical DA system is not responsive to acute EtOH administration.

Introduction

The medial prefrontal cortex (MPF) integrates a variety of sensory, limbic, and autonomic information from many brain areas (Groenewegen & Uylings, 2000) and is thought to subserve executive functions that are important for the optimal performance of certain goal-directed tasks (Koechlin et al., 2003, Robbins, 2000). The MPF receives dopamine (DA) projections as part of the mesocorticolimbic DA system and may play a role in mediating ethanol-drinking behavior. Recent studies have provided evidence of parallel systems in the dorsal and ventral MPF that are involved in mediating drug seeking and self-administration, respectively (McFarland et al., 2004).

Systemic administration of 2.0 g/kg ethanol reliably increases extracellular DA levels in the nucleus accumbens (NAC) (Yoshimoto et al., 1991) and ventral pallidum (Melendez et al., 2003) of Wistar rats. However, systemic administration of 0.5 g/kg ethanol significantly increased extracellular levels of DA in the NAC, but had no effect in the MPF (Bassareo et al., 1996). Similarly, Hegarty and Vogel (1993) observed that neither a 0.5 nor a 2.0 g/kg i.p. injection of ethanol had a significant effect on extracellular DA levels in the MPF. Together, these studies suggest that systemic administration of ethanol may not alter DA levels in the MPF and suggest that DA neurotransmission in this brain area may not be associated with the effects of ethanol. However, studies of DA mediation of ethanol self-administration indicate that DA D2/D3 receptors in the MPF are involved in ethanol self-administration behavior, as microinjections of the D2 antagonist Raclopride (0.05 and 1.0 μg/μl; bilateral) reduced the response rate and total responses for ethanol but had no effect on sucrose responding up to 20 μg/μl (Hodge et al., 1996). Moreover, recent work suggests that DA in the MPF is involved in mediating the onset and offset of ethanol drinking, whereas the NAC is more involved in the maintenance of ethanol self-administration (Samson & Chappell, 2003).

Some alcohol-preferring (P) lines of rats tend to have decreased tissue levels of DA in mesolimbic and cortical brain regions (Gongwer et al., 1989, Murphy et al., 1987). Moreover, immunohistological studies found a reduction in DA neuronal projections to mesolimbic areas, including the shell of the NAC and the cingulate cortex, in P rats as compared to alcohol-nonpreferring (NP) rats (Zhou et al., 1995). One study using quantitative microdialysis found that 5 days of ethanol administration (1.0 g/kg, i.p.) increased extracellular levels of DA in the NAC in both P and Wistar rats. However, no differences in basal DA levels were found between P, NP, and Wistar rats, suggesting that in this brain area, differences in DA content did not translate to differences in extracellular DA levels. Reduced DA neurotransmission in the MPF may result in an imbalance in the complex circuitry of the MPF, which integrates and processes autonomic, limbic, and sensory information important for the execution of goal-directed behavior (Groenewegen and Uylings, 2000, Robbins, 2000) and may contribute to the behavioral phenotypic profile of P rats, including alcohol preference (Murphy et al., 2002). Although several studies suggest that P rats may have a deficiency in the neurotransmitter DA in mesolimbic brain areas, no studies have directly examined extracellular levels in the MPF using a quantitative microdialysis approach, nor has the effect of systemic ethanol administration on extracellular MPF levels been assessed in P rats.

The differences in the alcohol-drinking phenotype of the P rat are thought to represent changes in the functional neurobiological makeup when compared with the outbred Wistar rat, which is the foundation stock for the P line of rats. Given the evidence implicating the MPF in executive function, goal-directed behavior, and the MPF DA system in ethanol self-administration behavior, coupled with low tissue levels of DA in the MPF, the hypothesis for this study was that DA neurotransmission would be reduced in the P rat as compared to the Wistar rat but would be more responsive to systemic alcohol administration. To test this hypothesis, quantitative no-net-flux (NNF) microdialysis was used to compare basal extracellular DA concentrations and conventional microdialysis was used to examine the effect of systemic administration of ethanol on the extracellular DA levels in the MPF of male Wistar and P rats.