TRANSIENT INCREASE IN PLATELET MONOAMINE OXIDASE B ACTIVITY DURING EARLY ABSTINENCE IN ALCOHOLICS: IMPLICATIONS FOR RESEARCH

Ulf Berggren, Claudia Fahlke1 and Jan Balldin*

Institute of Clinical Neuroscience, Department of Psychiatry and Neurochemistry, Sahlgren University Hospital/Mölndal, Göteborg University, SE-431 80 Mölndal and
1 Department of Psychology, Göteborg University, P.O. Box 500, SE-405 30 Göteborg, Sweden

Received 29 November 1999; in revised form 8 March 2000; accepted 27 March 2000


    ABSTRACT
 TOP
 FOOTNOTES
 ABSTRACT
 INTRODUCTION
 METHODS
 RESULTS
 DISCUSSION
 ACKNOWLEDGEMENTS
 REFERENCES
 
Some earlier studies have suggested that platelet monoamine oxidase (MAO)-B activity should be determined at time points other than early in the abstinence phase. However, the optimal times for blood sampling have not been precisely defined. We therefore assessed platelet MAO-B activity repeatedly in 13 male alcohol-dependent patients over the 2 months after the end of a period of heavy alcohol intake. Twelve healthy men were used as controls. In the alcohol-dependent patients, platelet MAO-B activity was transiently increased from 2 to 6 weeks after the end of alcohol intake and the values during this time period were not different from those of controls. Platelet MAO-B activity was, however, significantly lower in the alcohol-dependent patients at 1 week and at 2 months after the end of alcohol intake, in comparison to controls. It is concluded that the transient increase in platelet MAO-B activity after the end of alcohol intake in alcohol-dependent patients may conceal a difference from a control group. Therefore, it is suggested that when platelet MAO-B activity is determined, the preferential time point for obtaining those values in alcohol-dependent patients is after 2 months of abstinence.


    INTRODUCTION
 TOP
 FOOTNOTES
 ABSTRACT
 INTRODUCTION
 METHODS
 RESULTS
 DISCUSSION
 ACKNOWLEDGEMENTS
 REFERENCES
 
Platelet monoamine oxidase (MAO)-B activity has been investigated in alcoholism for more than two decades. Since the initial report by Gottfries et al. (1975), finding reduced MAO activity in post-mortem brains of alcoholics who had committed suicide, numerous studies have been performed using platelet MAO-B activity as a peripheral marker. Whether peripheral MAO-B activity represents central brain MAO-B activity has, however, been discussed. It is thus of note that in one study using positron emission tomography, brain and platelet MAO-B activity were shown to be highly correlated (Bench et al., 1991Go). In several studies on alcoholism, a low platelet MAO-B activity has been found and thus proposed as a trait marker for alcoholism, especially of ‘type 2’ alcoholism, which is associated with personality traits, such as monotony avoidance, impulsiveness, and sensation-seeking behaviour (Wiberg et al., 1977; Major and Murphy, 1978; Sullivan et al., 1978; von Knorring et al., 1984; Faraj et al., 1987; Pandey et al., 1988; Sullivan et al., 1990; Devor et al., 1993; Rommelspacher et al., 1994; Hallman et al., 1996; von Knorring and Oreland, 1996; Eriksson et al., work submitted). However, in some other studies, no such association was found (Tabakoff et al., 1988Go; Parsian et al., 1995Go; Anthenelli et al., 1998Go; Farren et al., 1998Go).

In some earlier studies, it has been shown that smokers also have low platelet MAO-B activity (for review see Oreland et al., 1999). In a recent study by Anthenelli et al. (1998), it was thus suggested that low platelet MAO-B activity is a state marker for cigarette smoking, rather than a trait marker for alcoholism or its subgroups. In a review on trait markers for alcoholism and their clinical utility by Farren and Tipton (1999), the authors discussed platelet MAO-B activity and alcoholism and concluded that ‘Probably the biggest problem has been the failure to address the potential confounder of cigarette smoking ...’ (Farren and Tipton, 1999, p. 652). However, they further concluded that ‘Despite this, it is difficult to ignore the literature suggesting an association between alcoholism and lowered platelet MAO-B activity. It is quite probable that some association does exist, but the nature of that association is not clear’ (Farren and Tipton, 1999, p. 652).

Consequently, the relationship between platelet MAO-B activity and alcoholism needs to be further investigated. From a methodological point of view, it appears that the length of the abstinence period is of particular importance. Platelet MAO-B activities have been reported to change during the early abstinence period and may be transiently increased during this time period (Wiberg et al., 1977Go; Alexopoulos et al., 1981Go; Major et al., 1981Go). Values for platelet MAO-B activity obtained during this period may therefore be falsely high and a difference in comparison to controls could not thus be obtained. Since the optimal time points for blood samplings have not been precisely defined, the aim of the present study was to investigate repeatedly platelet MAO-B activity in alcohol-dependent subjects for a period of 2 months after the end of alcohol intake. The patients were carefully controlled for sobriety during this time period. Values for platelet MAO-B activity were compared with that of a control group at each time point.


    METHODS
 TOP
 FOOTNOTES
 ABSTRACT
 INTRODUCTION
 METHODS
 RESULTS
 DISCUSSION
 ACKNOWLEDGEMENTS
 REFERENCES
 
Patients
This study was performed at an alcoholism treatment unit to which the patients had been committed by the social authorities after a long period of very heavy alcohol consumption. During the first week at this treatment unit, the patients were medicated for withdrawal symptoms. Most of them were given chlormethiazole (Heminevrin®) in decreasing doses and vitamins. In accordance with the general treatment scheme, the patients were cared for in the alcoholism treatment unit for a minimum of 2 months. To ensure their sobriety, they were carefully supervised by members of staff. Blood samplings for determination of liver functions and platelet MAO-B activity were performed once weekly during the first month and also 6 and 8 weeks after cessation of alcohol intake. The first blood samples (week 1) were drawn 1–3 days after withdrawal from alcohol.

Thirteen male patients with a diagnosis of alcohol dependence according to DSM-IV (American Psychiatric Association, 1994Go) were included in the study. Background data, including smoking status for the patients, are shown in Table 1Go.


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Table 1. Background data for 13 male patients
 
Twelve male subjects (age ± SD: 46 ± 10 years) were recruited as controls. Based on an interview, they were all considered physically and psychiatrically healthy. Results of routine laboratory analyses including liver parameters were all within the laboratory reference ranges. All controls were light social drinkers with a reported alcohol consumption of less than 100 g of pure alcohol weekly. The smoking status of the controls was not recorded.

Analysis of platelet MAO-B activity
Platelet-enriched plasma was obtained by centrifugation at 400 g for 10 min. The platelets were isolated by centrifugation at 2250 g for 15 min, then washed with saline and stored as a pellet at –70°C until analysis. MAO-B activity was determined according to Fowler et al. (1979) with ß-phenylethylamine as substrate. The substrate concentration was 12 µmol in a 0.013 mol/l phosphate buffer, pH 7.8, containing 0.03% (w/v) delipidized bovine serum albumin.

Statistics
All statistical calculations were performed using the statistical program Stat View (Abacus). Within-group (alcohol-dependent subjects) comparisons were performed with a paired t-test and between-group (alcohol-dependent subjects vs controls) comparisons with an unpaired t-test. In all tests, two-tailed levels of significance was used. The data are presented as means ± standard deviations (SD).

The study was approved by the Ethics Committee of the Göteborg University, Sweden and informed consent was obtained from all subjects.


    RESULTS
 TOP
 FOOTNOTES
 ABSTRACT
 INTRODUCTION
 METHODS
 RESULTS
 DISCUSSION
 ACKNOWLEDGEMENTS
 REFERENCES
 
In the alcohol-dependent patients, values of liver function tests were elevated at the beginning of the study, but found to decrease significantly during the period of investigation (Table 2Go).


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Table 2. Values for liver function parameters (mean ± SD) in 13 male alcohol-dependent patients
 
In the alcohol-dependent patients, platelet MAO-B activity did not differ between smokers (n = 3) and non-smokers (n = 10) at any time points. Thus, for the analyses, data from these two groups were combined. As seen in Fig. 1Go, the values for platelet MAO-B activity obtained at week 1 and at week 8 were not found to be significantly different. The platelet MAO-B activity was transiently increased between week 1 and week 8. Thus, there was a significant difference in platelet MAO-B activity when comparing week 1 with weeks 2, 3, and 4 (t = 2.63, P < 0.02; t = 2.24, P < 0.05; t = 2.34, P < 0.05, respectively). Moreover, there was a significant difference between week 2 compared to weeks 6 and 8 (t = 2.08, P < 0.05; t = 2.69, P < 0.02, respectively). The platelet MAO-B activity at week 4 was also found to be different from levels at week 8 (t = 2.57, P < 0.02). In total, 11 of the patients (85%) had a transient increase in platelet MAO-B activity with peak values between 2–4 weeks after the end of alcohol intake (at week 2: 7/11; week 3: 3/11; and week 4: 1/11).



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Fig. 1. Platelet monoamine oxidase (MAO)-B activity in 13 male alcohol-dependent patients repeatedly investigated over a period of 2 months after the end of heavy alcohol intake.

Values for 12 male controls are also shown. Values (in µkat/kg protein) are given as means ± SD. Within the group of alcohol-dependent subjects, there was a significant difference (paired t-test) in platelet MAO-B activity when comparing week 1 with weeks 2, 3, and 4 (P < 0.02, P < 0.05, P < 0.05), week 2 vs weeks 6 and 8 (P < 0.05, P < 0.02) and week 4 vs week 8 (P < 0.02). In comparison to controls, platelet MAO-B activity was significantly lower (unpaired t-test) at week 1 (P < 0.02) and at week 8 (P < 0.05).

 
In comparison to controls, the values for platelet MAO-B activity were significantly lower at week 1 (t = 2.67, P < 0.02) and at week 8 (t = 2.10, P < 0.05). The values from week 2 to week 6 were not different from that of controls.


    DISCUSSION
 TOP
 FOOTNOTES
 ABSTRACT
 INTRODUCTION
 METHODS
 RESULTS
 DISCUSSION
 ACKNOWLEDGEMENTS
 REFERENCES
 
In the present study, we repeatedly investigated platelet MAO-B activity in 13 male alcohol-dependent patients over 2 months after the end of a period of heavy alcohol intake. A limitation of the study is the relatively small number of subjects, which may reduce the validity of the findings. However, this patient group was extremely well controlled for sobriety during the investigation period. After the first week platelet MAO-B activity was transiently increased from week 2 until week 6. These results are in line with those obtained in earlier studies of alcoholics. Wiberg et al. (1977) reported a significant increase in platelet MAO-B activity during the first 10 days after the end of alcohol intake and a return to the original low level after 3 to 4 weeks. Major et al. (1981) also found a transient increase in platelet MAO-B activity from the initial value obtained within 72 h after the end of alcohol intake with a peak at about 1 week (66% increase above the initial value) and a return towards the initial value after 3 to 4 weeks, although still 15% above this value. Alexopoulos et al. (1981) found platelet MAO-B activity to be significantly higher in a group of alcoholics in withdrawal, 48 h after the end of alcohol intake, than in another group who had been abstinent for 6 weeks. Four of the patients were tested at both 48 h and 6 weeks and each of these had a transient increase in platelet MAO-B activity. This is consistent with results from the other studies and also with the present study, and thus strongly suggests a transient increase in platelet MAO-B activity after cessation of alcohol intake. The results from the studies of Wiberg et al. (1977), Major et al. (1981), and the present study further suggest that the peak in platelet MAO-B activity occurs between 1 and 2 weeks after the end of alcohol intake. The individual platelet MAO-B activities in the present study were transiently increased in 85% of the patients and mostly (64%) with a peak value 2 weeks after cessation of alcohol intake. Moreover, the present results suggest that the transient increase in platelet MAO-B activity may be relatively long-lasting. Major et al. (1981) suggested that studies comparing platelet MAO-B activity of alcoholics with controls should use blood samples obtained at least 6 weeks after the end of alcohol intake. The results in this study are in line with this suggestion, but also indicate that 2 months after the end of alcohol intake may be a preferential time point for blood sampling. In alcohol-dependent patients who cannot maintain sobriety for 2 months, platelet MAO-B activity obtained within 1 week of the end of alcohol intake might be used instead, since no significant difference between weeks 1 and 8 were observed.

The reason for the transitory increase in platelet MAO-B activity during early abstinence in alcoholics is not known with certainty. This increased activity after cessation of drinking has, however, been suggested to be due to either a temporary increase in the release of catecholamines in the blood, which occurs during the acute alcohol-withdrawal phase, or increased formation and release of newer platelets, containing higher levels of MAO-B activity (Wiberg et al., 1977Go; Alexopoulos et al., 1981Go; Major et al., 1981Go).

When comparing the values for platelet MAO-B activity at each time point during the 2 months of abstinence with those of a control group, we found that the patients had significantly lower platelet MAO-B activity at week 1 and at week 8 after cessation of alcohol intake. During the transient increase in platelet MAO-B activity from week 2 to week 6 the values were, on the other hand, not different from those of the control group. This latter finding clearly demonstrates that the transient increase in platelet MAO-B activity during early abstinence in alcohol-dependent patients may conceal a difference from a control group. Among studies that have found no difference in platelet MAO-B activity between alcoholics and controls, it is noteworthy that in the study by Tabakoff et al. (1988), the alcoholics had been abstinent for a mean period of 3 weeks. It is thus possible that a transient increase in platelet MAO-B activity, which occurs at this time point, may have concealed a difference from the control group, a possibility also discussed by the authors (Tabakoff et al., 1988Go). Regarding the study of Parsian et al. (1995), it is noted that the time points for determination of platelet MAO-B activity were recorded in only a subgroup of the subjects and the authors therefore concluded that the importance of the role of abstinence is difficult to evaluate in their study. In the multicentre investigation by Anthenelli et al. (1998), one factor that probably contributed to the results was, as also suggested by the authors, the inter-site differences where blood samples were collected at various lengths of abstinence from alcohol. On the other hand, in the study of Farren et al. (1998), the possibility of a transient increase in platelet MAO-B activity after the end of alcohol intake was carefully taken into consideration and thoroughly discussed. In that study, the alcoholics had been abstinent for an average time of about 4 months (16.7 weeks), which is a time point at which no transient changes in platelet MAO-B activity is expected (Farren et al., 1998Go).

Taken together, we found in this study that platelet MAO-B activity was transiently increased in alcohol-dependent patients from 2 to 6 weeks after the end of a period of heavy alcohol intake. During this period, the platelet MAO-B activity levels were not different from that of a control group. In this study, platelet MAO-B activities were, however, lower immediately after the end of the drinking period and also after 2 months of abstinence, compared to controls. This clearly demonstrates the necessity to use defined time points for collection of blood in an investigation of platelet MAO-B activity in alcoholism. The transient increase in platelet MAO-B activity in alcohol abstinence may conceal a difference from a control group. We thus suggest that the preferential time point for obtaining values for platelet MAO-B activity should be 2 months after the end of alcohol intake or, as mentioned earlier, immediately after the end of the drinking period.


    ACKNOWLEDGEMENTS
 TOP
 FOOTNOTES
 ABSTRACT
 INTRODUCTION
 METHODS
 RESULTS
 DISCUSSION
 ACKNOWLEDGEMENTS
 REFERENCES
 
This research was supported by the Swedish Medical Research Council (K98-21P-11842-03A).


    FOOTNOTES
 TOP
 FOOTNOTES
 ABSTRACT
 INTRODUCTION
 METHODS
 RESULTS
 DISCUSSION
 ACKNOWLEDGEMENTS
 REFERENCES
 
* Author to whom correspondence should be addressed. Back


    REFERENCES
 TOP
 FOOTNOTES
 ABSTRACT
 INTRODUCTION
 METHODS
 RESULTS
 DISCUSSION
 ACKNOWLEDGEMENTS
 REFERENCES
 
Alexopoulos, G. S., Lieberman, K. W., Frances, R. and Stokes, P. E. (1981) Platelet MAO during the alcohol withdrawal syndrome. American Journal of Psychiatry 138, 1254–1255.[Abstract]

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