Original Research Article

Molecular Psychiatry (2003) 8, 231–234. doi:10.1038/sj.mp.4001214

Serotonin transporter availability correlates with alcohol intake in non-human primates

A Heinz1, D W Jones1, J G Gorey1, A Bennet2, S J Suomi3, D R Weinberger1 and J D Higley2

  1. 1Clinical Brain Disorders Branch, NIMH, NIH, Bethesda, MD, USA
  2. 2Laboratory of Clinical Studies, DICBR, NIAAA, Bethesda, MD, USA
  3. 3Laboratory of Comparative Ethology, NICHD, Bethesda, MD, USA

Correspondence: A Heinz, Professor of Psychiatry, Chair, Department of Psychiatry of the Charité, Humboldt, University Berlin, Schumannstr. 20/21, 10117 Berlin, Germany. E-mail: andreas.heinz@charite.de

Received 3 January 2002; Revised 3 May 2002; Accepted 6 May 2002.

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Abstract

A low level of alcohol intoxication upon initial exposure and impulsive aggressiveness predispose humans to alcoholism. In non-human primates, central serotonin transporter availability and turnover rate were associated with aggressive behavior and a low response to initial alcohol exposure. We assessed the respective effects of these factors on alcohol intake in a free choice paradigm. Serotonin transporter availability in the raphe area, the origin of central serotonergic projections, was measured with single-photon emission computed tomography and the radioligand [123I]beta-CIT in 11 rhesus monkeys with low and high central serotonin turnover. The amount of alcohol intake in the 3-month observation period was positively correlated with serotonin transporter availability (R=0.76, P=0.006), but not with aggressiveness (R=0.19, P=0.6) or alcohol response upon first exposure (R=-0.48, P=0.2). In a linear multiple regression analysis with serotonin transporter availability, alcohol response, and aggressiveness as independent variables, 82% of the variance of alcohol intake was explained and serotonin transporter availability emerged as the only statistically significant factor (beta=7.81, P=0.006). These observations indicate that there may be a direct relationship between serotonin transporter availability and alcohol intake after controlling for aggression and alcohol response on first exposure.

Keywords:

disposition to alcoholism, alcohol response, molecular neuroimaging, beta-CIT SPECT, aggressiveness

Alcoholism is characterized by the inability of individuals to regulate their consumption of alcohol appropriately. The biological substrates remain to be elucidated and could lead to a better understanding of all forms of drug abuse.1,2 Several lines of evidence suggest that serotonergic dysfunction plays a role in the pathogenesis of alcoholism.3 In humans and non-human primates, reduced central nervous system (CNS) serotonin turnover rate, as measured by the serotonin metabolite 5-hydroxyindoleacetic acid (5-HIAA) in cerebrospinal fluid (CSF), has been implicated in the pathogenesis of alcohol abuse.4,5 In alcoholics, early onset of disease and impulsive aggressiveness were associated with low 5-HIAA concentrations in the CSF.4,6 In young human adults, reduced intoxication following alcohol consumption preceded later development of alcohol dependence,7 and was found among individuals with a certain genotypic variant of the serotonin transporter promoter8 that was associated with an increased density and functional capacity of serotonin uptake sites.9,10

Developmental factors have also been implicated in studies of adopted children of alcoholic parents, where early paternal neglect and later placement in adoptive families were associated with severity of adult alcohol consumption.11 In non-human primates, environmental stressors during early development such as parental absence reduce 5-HIAA concentrations in the CSF and increase alcohol consumption, aggression, and anxiety-like behavior during adolescence.5 In such primates, we previously observed that a low 5-HIAA concentration in the CSF and an increased availability of brainstem serotonin transporters correlated with two factors that may predispose to excessive alcohol intake: aggressive behavior and a low response to alcohol effects upon initial exposure.12 We now report the respective effects of these factors on alcohol intake of these primates in a free choice paradigm. Serotonin transporter availability in the brains of living rhesus monkeys was measured with single-photon emission computed tomography (SPECT) and the radioligand [123I]2beta-carbomethoxy-3beta-(4-iodophenyl) tropane ([123I]beta-CIT).13,14

Availability of serotonin transporters in the brainstem raphe area significantly and positively correlated with the amount of alcohol intake under free access conditions during the 3-month observation time (Spearman's R=0.76, P=0.006; Figure 1). Specifically, higher transporter expression in the area of the raphe nuclei was associated with greater alcohol use. The amount of alcohol intake in the free choice paradigm was not significantly correlated with aggressiveness (R=0.19, P=0.6) or with alcohol response upon first exposure (R=-0.48, P=0.2). Nor did we observe significant correlations between alcohol intake during the 3-month observation period and 5-HIAA (R=-0.35, P=0.3), HVA (R=0.03, P=0.9), or MHPG (R=-0.2, P=0.6) concentrations in the CSF.

Figure 1.
Figure 1 - Unfortunately we are unable to provide accessible alternative text for this. If you require assistance to access this image, please contact help@nature.com or the author

Correlation between the binding of the radioligand beta-CIT to brainstem serotonin transporters and average alcohol intake (g/kg and day) in a free access paradigm (Spearman's R=0.76, P=0.006). During an observation time of 3 months, subjects were allowed unfettered access to alcohol for 1 h per day for 4 days per week.

Full figure and legend (13K)

In a linear multiple regression analysis with the amount of alcohol intake as the dependent variable, the availability of brainstem serotonin transporters, alcohol intoxication, and aggressiveness as independent variables explained 82% of the variance in alcohol intake (F(3,6)=9.24, P=0.01). While other studies have shown the importance of all three variables in predicting alcohol consumption patterns,7,15 in the current analysis the availability of brainstem serotonin transporters was the only significant factor that statistically predicted excessive alcohol intake (beta=7.81, P=0.006), while neither aggressiveness (beta=0.30, P=0.2) nor the severity of alcohol intoxication upon first exposure (beta=0.34, P=0.2) reached statistical significance. No significant correlations between beta-CIT binding in any other region of interest outside of the raphe nuclei region and behavioral data were found.

We observed a positive correlation between serotonin transporter availability in the raphe area and alcohol intake in a free choice paradigm. The raphe nuclei contain the cell bodies of serotonergic neurons and represent a core region of the central serotonergic system, where all central serotonergic projections originate.16 In vivo microdialysis, serotonin depletion and restoration studies, and parallel assessment of CSF neurotransmitter metabolites in monkeys have shown that beta-CIT binding at monoamine transporters reflects extracellular monoamine concentrations.12,13,17 Extracellular serotonin concentrations in the raphe area interact with autoreceptors on serotonergic neurons and thus modulate serotonin release in cortical and subcortical projection areas.16 Concentrations of the serotonin metabolite 5-HIAA in the CSF, on the other hand, can also be influenced by spinal serotonin turnover and in this study were not associated with the amount of alcohol intake. Therefore, assessment of raphe serotonin transporters with brain imaging may be a preferable measure to reflect the overall functional state of central serotonergic neurotransmission.

We previously observed that a high availability of brainstem serotonin transporters is correlated with increased aggressiveness and a low response to alcohol upon first exposure.12 In this study, high availability of brainstem serotonin transporters, but not aggressiveness or a low level of alcohol response upon first exposure, was associated with high ethanol intake. This finding suggests that high alcohol intake is directly associated with serotonin transporter availability, while both aggression and alcohol response to first exposure may be independently related to serotonin function but may not significantly contribute to excessive alcohol consumption per se. Therefore, the univariate correlations that have previously been detected between alcohol consumption and initial alcohol response7,15 and alcohol consumption and aggressiveness5,6 may be a result of the underlying common correlation between serotonergic neurotransmission and the other variables.

The association between high serotonin transporter availability and a low level of alcohol intoxication upon first exposure is in striking agreement with the observations of Schuckit et al,8 where, in a prospective study, those who had a genetic disposition towards high serotonin transporters and who showed a low response to acute alcohol intoxication developed excessive alcohol intake and became alcoholic. The low response to acute alcohol intake may partly be mediated by a reduced GABAergic sedation among subjects with a low serotonin turnover rate.18

Together, our findings indicate that central serotonergic dysfunction in the area of the raphe nuclei is associated with reduced sensitivity to alcohol intoxication and excessive alcohol intake, factors implicated in the pathogenesis of alcoholism.7,15 These observations may aid in the development of preventive and therapeutic strategies that target specific monoaminergic dysfunctions.

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Methods

All protocols used in this study were reviewed and approved by animal care committees of the intramural research programs of the NIH. In total, 11 rhesus monkeys (macaca mulatta, age 5 years) underwent a series of four, 4-day social isolations at 6 months of age. One month after the social isolation, the subjects were placed in same-age social groups, where they remained until the present study. Concentrations of CSF 5-HIAA, homovanillic acid (HVA), and 3-methoxy-4-hydroxyphenylglycol (MHPG) were acquired periodically via cisternal puncture within 25 min of ketamine (10 mg/kg i.m.) administration and quantified with liquid chromatography and electrochemical detection as described in detail elsewhere.5 As previously reported,12 CSF 5-HIAA concentrations measured at ages 2 and 3 years in these primates were significantly correlated with 5-HIAA levels collected at age 5 years (R=0.65 and 0.77, P<0.05, respectively) and with beta-CIT binding assessed at age 5 years (R=-0.63 and -0.60, P<0.05, respectively), indicating a trait-like character of serotonergic neurotransmission in these primates. Serotonin transporter availability in the raphe area was not significantly correlated with dopamine or norepinephrine metabolites in the CSF (R=-0.1 and -0.6, P>0.05, respectively).

The monkeys were periodically assessed on standardized ratings of their social behavior and frequency of initiated aggression with Likert scales.5,12 Ratings of alcohol intoxication were based on sedation, body sway, and ataxia after a single administration of a standardized quantity of alcohol (10 ml/kg body weight of an 8.4% v/v alcohol solution). The animals were then exposed to a sweetened 8.4% v/v alcohol drink in a free access paradigm.5 Subjects were neither food nor water deprived throughout the experiment. The drinking station was a 'closet-like' Plexiglas-enclosed perch, where subjects could enter from the bottom and drink without the other subjects harassing them. After spout training, the color of the vehicle was changed. Two bottles of solution were freely accessible: one bottle of 8.4% alcohol solution sweetened with aspartame and one bottle of aspartame-sweetened water. Subjects were allowed unfettered access to the alcohol solution for 1 h each day (between 1300 and 1500 h) for 4 days per week. Alcohol consumption was recorded during an observation time of 3 months.

Serotonin transporters in the raphe area were assessed with [123I]beta-CIT and SPECT as described previously.12 The time period between alcohol exposure and SPECT scanning did not exceed 3 months. [123I]beta-CIT binds with high affinity to serotonin transporters in the brainstem.13 [123I]beta-CIT was prepared14 and administered intravenously (dose range: 11–14 mCi). Two SPECT scans (CERASPECT; Digital Scintigraphics, Waltham, MA; full-width at half-maximum 7.5 mm; 64 contiguous 1.67 mm slices; 30 min per scan) were acquired 2 and 3 h after injection, when equilibrium is established at brainstem serotonin transporters.19 Individual ROIs were drawn based on a standard atlas of rhesus macaque brain and comprised the basal ganglia, thalamus, brainstem, and cerebellum.12 The highest density of serotonin transporters is found in the brainstem area which contains the raphe system. ROI placement was confirmed on coregistered MRI scans for a subset of the subjects. Activity concentration (cpm/ml) was measured for the volume encompassed by each set of ROIs. Brainstem binding of [123I]beta-CIT was analysed using the ratio of specific to unspecific binding.19

Correlations between SPECT measurements and behavior and metabolite assessment were made by Spearman's correlation coefficient. A linear multiple regression analysis was used to assess the effects of the hypothesized variables (alcohol response, aggressiveness, and serotonin transporter availability) on alcohol intake.

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Acknowledgements

This work was inspired by Markku Linnoila, who died in 1999. Parts of this study were supported by the Deutsche Forschungsgemeinschaft (He 2597/7-1).

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