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Plasma GABA in affective illness
Journal of Affective Elsevier/North-Holland
Disorders, 3 (1981) 339-343 Biomedical Press
339
PLASMA GABA IN AFFECTIVE ILLNESS A Preliminary Investigation
FREDERICK
PETTY
1 and MICHAEL
A. SCHLESSER
*
1 Department of Psychiatry, University of Iowa College of Medicine, University of Iowa, and Veterans Administration Medical Center, Iowa City, IA 52242 and * Department of Psychiatry, University of Texas Southwest Medical School, Dallas, TX 75235 (U.S.A.) (Received (Accepted
20 May, 1981) 4 June, 1981)
SUMMARY Plasma levels of y-aminobutyric acid (GABA) were determined in 34 clinically symptomatic patients with diagnosis of affective disorder and in 20 normal controls. Lowest levels were found in patients with familial pure depressive disease and in depression spectrum disease, while levels of patients with sporadic depressive disease, though significantly lower than control, were not as low. Plasma GABA levels in secondary depression and in bipolar depression were similar to control. Bipolar manic patients had plasma GABA levels that were significantly higher than control values.
INTRODUCTION
About eight years ago, derangements in the metabolism of y-aminobutyric acid (GABA), the major inhibitory neurotransmitter in mammalian brain, were reported in neurological disease. Perry et al. (1973) found decreased levels of GABA in postmortem basal ganglia tissue from patients with Huntington’s chorea. This report was subsequently confirmed by the finding of Bird and Iversen (1974) that glutamate dehydrogenase (GAD), the GABAsynthesizing enzyme, was also low in choreic basal ganglia tissue. More recently, Perry et al. (1979) reported that deficits of GABA in thalamus and nucleus accumbens occurred in brains of patients who had died from schizophrenia. This work was made possible by a Veterans Administration Research Career ment Award to F.P. and by a research Grant from the Department of Psychiatry, sity of Iowa.
01650327/81/0000-0000/$02.75
@ 1981
Elsevier/North-Holland
Biomedical
DevelopUniver-
Press
340
Improved techniques for quantitative determination of subnanomole levels of GABA have allowed extention of the original studies, which were performed with postmortem brain, to in vivo determinations on cerebral spinal fluid (CSF). Levels of GABA in CSF are significantly low in patients with Parkinson’s disease, Huntington’s disease, and dementia (Enna et al. 1977; Manyam et al. 1980), conditions which are significantly associated with depression. CSF GABA levels have also been found to be lower in patients with depression and schizoaffective disorder than in patients with schizophrenia or than neurologic controls (Gold et al. 1980). CSF GABA has also been reported to be elevated in patients with schizophrenia (McCarthy et al. 1981), however, this finding is difficult to interpret since another group reported normal CSF levels in schizophrenics, and that treatment with neuroleptic drugs increases CSF GABA (Zimmer et al. 1981). We have examined plasma levels of GABA in patients with affective disorder. METHODS
Clinical In this preliminary investigation, 34 patients were included who received diagnoses of affective disorder using the criteria of Feighner et al. (1972). Of these, 10 had primary bipolar affective disorder (mania n = 6, bipolar depression n = 4) and 17 had primary unipolar disorder. Seven patients were diagnosed as having secondary depression, and in 5 of these cases the primary diagnosis was schizophrenia. Patients with primary unipolar affective disorder were further categorized using the family history classification scheme of Winokur (Winokur et al. 1978; Schlesser et al. 1980). Eighteen of the patients were female and 16 were male. Most of the patients were receiving psychotropic medication at the time of venipuncture. After obtaining informed consent, 8 a.m. fasting blood samples were drawn into heparinized glass tubes. A normal control group was obtained from volunteers who were laboratory personnel at a university psychiatric hospital or staff at a community mental health center, in whom personal and family history of psychiatric illness was denied.
Laboratory The blood samples were promptly centrifuged at 1500 X g for 10 min, and the plasma was removed, quickly frozen, and stored at -20°C until time of assay, A loo-p1 aliquot was transferred into a 1.5-ml plastic test tube for amino acid analysis. Protein was precipitated with 100 ~1 of 3 N HC104, and after centrifugation for 1 min at 15 000 X g, the entire supematant was applied to a Rexin 101 cation exchange column, from which the amino acids were eluted with 1 N NH,OH. After lyophilization, the n-butyl trifluoroacyl
341
derivatives were prepared for gas chromatographic analysis using acidified butanol and trifluoroacetic anhydride. The assay was performed with a Shimadzu GC-7A gas chromatograph equipped with a flame thermionic detector, using a 1 m X 2 mm ID column packed with Tabsorb. Analyses were performed for alanine, valine, glycine, threonine, GABA, phenylalanine, aspartic acid, glutamic acid, and lysine. Plasma amino acid assays were performed blind to the patient’s diagnosis. RESULTS
Plasma GABA levels obtained for the normal controls were 66 + 14 ng/ ml (mean + SD, n = 20). No differences were noted between males and females (n = 10). For the patients, no significant differences between diagnostic categories were noted for any amino acid other than GABA (Fig. 1). Primary bipolar patients in the manic phase were noted to have significantly elevated levels (86 * 7 ng/ml), vs. control (t = 3.34, df 24, P < 0.01). Patients with bipolar depression and secondary depression had levels essentially the same as control (both 62 2 7 ng/ml). Patients in all categories of unipolar depression had levels significantly lower than control, with familial pure depressive disease (19 f. 5 ng/ml, t = 7.29, df 23, P < 0.01) and depression spectrum disease (21 + 4 ng/ml, t = 9.29, df 25, P < 0.01) having the lowest levels. Levels of patients with sporadic depressive disease were not as low (37 2 9 ng/ml), but still significantly lower than control, (t = 4.37, df 22, P < 0.01). These levels were also significantly higher than those of patients with familial pure depressive disease and depression spectrum disease, (t = 5.53, df 15, P < 0.01).
,Manic
Dep,
1’ Bipolor
%SDD FPDD DSD,
2ODep.
Control
I0 Unipolar
Fig. 1. Plasma GABA levels in patients with affective illness. Data are presented as mean * SD. Group size: Primary bipolar, manic (6); primary bipolar, depressed (4); primary unipolar, sporadic depressive disease (SDD) (5); familial pure depressive disease (FPDD) (5); depression spectrum disease (DSD) (7); secondary depression (7). Control values are those reported by Loscher (1979), which we have subsequently replicated.
342 DISCUSSION
To our knowledge, this is the first report of abnormal plasma levels of GABA in psychiatric illness. The values of plasma GABA that we obtain for normal controls are almost identical to those reported by Loscher (1979) but are somewhat higher than the values reported by Ferkany et al. (1978). Since both these authors used a radio-receptor assay, the discrepancy between them is difficult to understand; however, the chromatographic procedure employed in the present study is generally accepted as being a more selective procedure. The extent to which peripheral GABA reflects central GABAergic activity is, at this time, moot. Although approximately 93% of GAD activity in the mammalian body is in brain (Haber et al. 1970), and about 99% of human GABA is found in the central nervous system (Zachmann et al. 1966), the precise contribution of the non-CNS GAD to peripherally circulating GABA levels has yet to be ascertained. On the other hand, maneuvers which increase brain levels of GABA by 75% also lead to increases of plasma levels by a comparable proportion (Ferkany et al. 1978). Similarly, procedures which increase cerebral spinal fluid (CSF) GABA also cause comparable increases in plasma GABA (Loscher 1979). This being a preliminary study, no attempt was made to control for drug effects, but further research should clarify their effects (or lack of them) on plasma GABA. Regardless of whether plasma GABA measured in the present study reflects effects of peripheral GAD activity and/or medication, in our patient sample, patients with primary unipolar affective disorder had plasma GABA levels which were significantly lower than control. That GABA hypofunction should accompany the depressed state is a prediction of the learned helplessness animal model of depression, in which GABA has been demonstrated to play a controlling role, particularly in the hippocampus (Sherman and Petty 1980). The present research also supports the finding of Gold et al. (1980) of lowered CSF levels in depressed patients. It is interesting to note that patients with secondary depression had plasma GABA levels comparable to those of normal controls, since most of these patients had a diagnosis of schizophrenia, in which elevated CSF levels have been reported by one group (McCarthy et al. 1981), but in which normal GABA levels in CSF were reported by another (Zimmer et al. 1981). The finding that plasma GABA levels in patients with sporadic depressive disease were intermediate between those of familial pure depressive disease and depression spectrum disease, and control values, and which were significantly different from both, is of interest; since, in sporadic depressive disease, no familial component can be demonstrated. This further supports the notion that familial pure depressive disease, depression spectrum disease, and sporadic depressive disease are valid diagnostic entities. The findings with bipolars are difficult to interpret, but further research should clarify the specificity and selectivity of this preliminary work.
343 ACKNOWLEDGEMENT
We also thank Arnold D. Sherman,
Ph.D., for help with the GABA assay.
REFERENCES Bird, E.D. and Iversen, L.L., Huntington’s chorea - Post-mortem measurement of glutamic acid decarboxylase, choline acetyltransferase and dopamine in basal ganglia, Brain, 97 (1974) 457-472. Enna, S.J., Stern, L.Z., Wastek, G.J. and Yamamura, H.I., Cerebrospinal fluid ‘y-aminobutyric acid variations in neurological disorders, Arch. Neurol. (Chic.), 34 (1977) 683-685. Feighner, J.P., Robins, E., Guze, S.B., Woodruff, R.A., Winokur, G. and Munoz, R., Diagnostic criteria for use in psychiatric research, Arch. Gen. Psychiat., 26 (1972) 57-63. Ferkany, J.W., Smith, L.A., Seifert, Jr., W.E., Caprioli, R.M. and Enna, S.J., Measurement of gamma-aminobutyric acid (GABA) in blood, Life Sci., 22 (1978) 2121-2128. Gold, B.I., Bowers, M.B., Roth, R.H. and Sweeney, D.W., GABA levels in CSF of patients with psychiatric disorders, Amer. J. Psychiat., 137 (1980) 362-364. Haber, B., Kuriyama, K. and Roberts, E., An anion stimulated L-glutamic acid decarboxylase in non-neural tissues -occurrence and subcellular localization in mouse kidney and developing chick embryo brain, Biochem. Pharmacol., 19 (1970) 1119-1136. Liischer, W., GABA in plasma and cerebrospinal fluid of different species - Effects of y-acetylenic GABA, y-vinyl GABA and sodium valproate, J. Neurochem., 32 (1979) 1587-1591. McCarthy, B.W., Gomes, U.R., Neethling, A.C., Shanley, B.C., Taljaard, J.J.F., Potgieter, L. and Roux, J.T., y-Aminobutyric acid concentration in cerebrospinal fluid in schizophrenia, J. Neurochem., 36(4) (1981) 1406-1408. Manyam, N.V.B., Katz, L., Hare, T.A., Gerber, III, J.C. and Grossman, M.H., Levels of y-aminobutyric acid in cerebrospinal fluid in various neurologic disorders, Arch. Neurol. (Chic.), 37 (1980) 352-355. Perry, T.L., Hansen, S. and Kloster, M., Huntington’s chorea deficiency of y-aminobutyric acid in brain, N. Engl. J. Med., 288(7) (1973) 337-342. Perry, T.L., Buchanan, J., Kish, S.J. and Hansen, S., y-Aminobutyric-acid deficiency in brain of schizophrenic patients, Lancet, i (1979) 237-239. Schlesser, M.A., Winokur, G. and Sherman, B.M., Hypothalamic-pituitary-adrenal axis activity in depressive illness, Arch. Gen. Psychiat., 37 (1980) 737-743. Sherman, A.D. and Petty, F., Neurochemical basis of the action of antidepressants on learned helplessness, Behav. Neural Biol., 30 (1980) 119-134. Winokur, G., Behar, D., Vanvalkenburg, C. and Lowry, M., Is a familial definition of depression both feasible and valid? J. Nerv. Ment. Dis., 166(11) (1978) 764-768. Zachmann, M., Tocci, P. and Nyhan, W.L., The occurrence of y-aminobutyric acid in human tissues other than brain, J. Biol. Chem., 241 (1966) 1355-1358. Zimmer, R., Teeklen, A.W., Meier, K.D., Ackenheil, M. and Zander, K.J., Preliminary studies on CSF gamma-aminobutyric acid levels in psychiatric patients before and during the treatment with different psychotropic durgs, Progr. Neuro-Psychopharmacol., 4 (1981) 613-620.
Disorders, 3 (1981) 339-343 Biomedical Press
339
PLASMA GABA IN AFFECTIVE ILLNESS A Preliminary Investigation
FREDERICK
PETTY
1 and MICHAEL
A. SCHLESSER
*
1 Department of Psychiatry, University of Iowa College of Medicine, University of Iowa, and Veterans Administration Medical Center, Iowa City, IA 52242 and * Department of Psychiatry, University of Texas Southwest Medical School, Dallas, TX 75235 (U.S.A.) (Received (Accepted
20 May, 1981) 4 June, 1981)
SUMMARY Plasma levels of y-aminobutyric acid (GABA) were determined in 34 clinically symptomatic patients with diagnosis of affective disorder and in 20 normal controls. Lowest levels were found in patients with familial pure depressive disease and in depression spectrum disease, while levels of patients with sporadic depressive disease, though significantly lower than control, were not as low. Plasma GABA levels in secondary depression and in bipolar depression were similar to control. Bipolar manic patients had plasma GABA levels that were significantly higher than control values.
INTRODUCTION
About eight years ago, derangements in the metabolism of y-aminobutyric acid (GABA), the major inhibitory neurotransmitter in mammalian brain, were reported in neurological disease. Perry et al. (1973) found decreased levels of GABA in postmortem basal ganglia tissue from patients with Huntington’s chorea. This report was subsequently confirmed by the finding of Bird and Iversen (1974) that glutamate dehydrogenase (GAD), the GABAsynthesizing enzyme, was also low in choreic basal ganglia tissue. More recently, Perry et al. (1979) reported that deficits of GABA in thalamus and nucleus accumbens occurred in brains of patients who had died from schizophrenia. This work was made possible by a Veterans Administration Research Career ment Award to F.P. and by a research Grant from the Department of Psychiatry, sity of Iowa.
01650327/81/0000-0000/$02.75
@ 1981
Elsevier/North-Holland
Biomedical
DevelopUniver-
Press
340
Improved techniques for quantitative determination of subnanomole levels of GABA have allowed extention of the original studies, which were performed with postmortem brain, to in vivo determinations on cerebral spinal fluid (CSF). Levels of GABA in CSF are significantly low in patients with Parkinson’s disease, Huntington’s disease, and dementia (Enna et al. 1977; Manyam et al. 1980), conditions which are significantly associated with depression. CSF GABA levels have also been found to be lower in patients with depression and schizoaffective disorder than in patients with schizophrenia or than neurologic controls (Gold et al. 1980). CSF GABA has also been reported to be elevated in patients with schizophrenia (McCarthy et al. 1981), however, this finding is difficult to interpret since another group reported normal CSF levels in schizophrenics, and that treatment with neuroleptic drugs increases CSF GABA (Zimmer et al. 1981). We have examined plasma levels of GABA in patients with affective disorder. METHODS
Clinical In this preliminary investigation, 34 patients were included who received diagnoses of affective disorder using the criteria of Feighner et al. (1972). Of these, 10 had primary bipolar affective disorder (mania n = 6, bipolar depression n = 4) and 17 had primary unipolar disorder. Seven patients were diagnosed as having secondary depression, and in 5 of these cases the primary diagnosis was schizophrenia. Patients with primary unipolar affective disorder were further categorized using the family history classification scheme of Winokur (Winokur et al. 1978; Schlesser et al. 1980). Eighteen of the patients were female and 16 were male. Most of the patients were receiving psychotropic medication at the time of venipuncture. After obtaining informed consent, 8 a.m. fasting blood samples were drawn into heparinized glass tubes. A normal control group was obtained from volunteers who were laboratory personnel at a university psychiatric hospital or staff at a community mental health center, in whom personal and family history of psychiatric illness was denied.
Laboratory The blood samples were promptly centrifuged at 1500 X g for 10 min, and the plasma was removed, quickly frozen, and stored at -20°C until time of assay, A loo-p1 aliquot was transferred into a 1.5-ml plastic test tube for amino acid analysis. Protein was precipitated with 100 ~1 of 3 N HC104, and after centrifugation for 1 min at 15 000 X g, the entire supematant was applied to a Rexin 101 cation exchange column, from which the amino acids were eluted with 1 N NH,OH. After lyophilization, the n-butyl trifluoroacyl
341
derivatives were prepared for gas chromatographic analysis using acidified butanol and trifluoroacetic anhydride. The assay was performed with a Shimadzu GC-7A gas chromatograph equipped with a flame thermionic detector, using a 1 m X 2 mm ID column packed with Tabsorb. Analyses were performed for alanine, valine, glycine, threonine, GABA, phenylalanine, aspartic acid, glutamic acid, and lysine. Plasma amino acid assays were performed blind to the patient’s diagnosis. RESULTS
Plasma GABA levels obtained for the normal controls were 66 + 14 ng/ ml (mean + SD, n = 20). No differences were noted between males and females (n = 10). For the patients, no significant differences between diagnostic categories were noted for any amino acid other than GABA (Fig. 1). Primary bipolar patients in the manic phase were noted to have significantly elevated levels (86 * 7 ng/ml), vs. control (t = 3.34, df 24, P < 0.01). Patients with bipolar depression and secondary depression had levels essentially the same as control (both 62 2 7 ng/ml). Patients in all categories of unipolar depression had levels significantly lower than control, with familial pure depressive disease (19 f. 5 ng/ml, t = 7.29, df 23, P < 0.01) and depression spectrum disease (21 + 4 ng/ml, t = 9.29, df 25, P < 0.01) having the lowest levels. Levels of patients with sporadic depressive disease were not as low (37 2 9 ng/ml), but still significantly lower than control, (t = 4.37, df 22, P < 0.01). These levels were also significantly higher than those of patients with familial pure depressive disease and depression spectrum disease, (t = 5.53, df 15, P < 0.01).
,Manic
Dep,
1’ Bipolor
%SDD FPDD DSD,
2ODep.
Control
I0 Unipolar
Fig. 1. Plasma GABA levels in patients with affective illness. Data are presented as mean * SD. Group size: Primary bipolar, manic (6); primary bipolar, depressed (4); primary unipolar, sporadic depressive disease (SDD) (5); familial pure depressive disease (FPDD) (5); depression spectrum disease (DSD) (7); secondary depression (7). Control values are those reported by Loscher (1979), which we have subsequently replicated.
342 DISCUSSION
To our knowledge, this is the first report of abnormal plasma levels of GABA in psychiatric illness. The values of plasma GABA that we obtain for normal controls are almost identical to those reported by Loscher (1979) but are somewhat higher than the values reported by Ferkany et al. (1978). Since both these authors used a radio-receptor assay, the discrepancy between them is difficult to understand; however, the chromatographic procedure employed in the present study is generally accepted as being a more selective procedure. The extent to which peripheral GABA reflects central GABAergic activity is, at this time, moot. Although approximately 93% of GAD activity in the mammalian body is in brain (Haber et al. 1970), and about 99% of human GABA is found in the central nervous system (Zachmann et al. 1966), the precise contribution of the non-CNS GAD to peripherally circulating GABA levels has yet to be ascertained. On the other hand, maneuvers which increase brain levels of GABA by 75% also lead to increases of plasma levels by a comparable proportion (Ferkany et al. 1978). Similarly, procedures which increase cerebral spinal fluid (CSF) GABA also cause comparable increases in plasma GABA (Loscher 1979). This being a preliminary study, no attempt was made to control for drug effects, but further research should clarify their effects (or lack of them) on plasma GABA. Regardless of whether plasma GABA measured in the present study reflects effects of peripheral GAD activity and/or medication, in our patient sample, patients with primary unipolar affective disorder had plasma GABA levels which were significantly lower than control. That GABA hypofunction should accompany the depressed state is a prediction of the learned helplessness animal model of depression, in which GABA has been demonstrated to play a controlling role, particularly in the hippocampus (Sherman and Petty 1980). The present research also supports the finding of Gold et al. (1980) of lowered CSF levels in depressed patients. It is interesting to note that patients with secondary depression had plasma GABA levels comparable to those of normal controls, since most of these patients had a diagnosis of schizophrenia, in which elevated CSF levels have been reported by one group (McCarthy et al. 1981), but in which normal GABA levels in CSF were reported by another (Zimmer et al. 1981). The finding that plasma GABA levels in patients with sporadic depressive disease were intermediate between those of familial pure depressive disease and depression spectrum disease, and control values, and which were significantly different from both, is of interest; since, in sporadic depressive disease, no familial component can be demonstrated. This further supports the notion that familial pure depressive disease, depression spectrum disease, and sporadic depressive disease are valid diagnostic entities. The findings with bipolars are difficult to interpret, but further research should clarify the specificity and selectivity of this preliminary work.
343 ACKNOWLEDGEMENT
We also thank Arnold D. Sherman,
Ph.D., for help with the GABA assay.
REFERENCES Bird, E.D. and Iversen, L.L., Huntington’s chorea - Post-mortem measurement of glutamic acid decarboxylase, choline acetyltransferase and dopamine in basal ganglia, Brain, 97 (1974) 457-472. Enna, S.J., Stern, L.Z., Wastek, G.J. and Yamamura, H.I., Cerebrospinal fluid ‘y-aminobutyric acid variations in neurological disorders, Arch. Neurol. (Chic.), 34 (1977) 683-685. Feighner, J.P., Robins, E., Guze, S.B., Woodruff, R.A., Winokur, G. and Munoz, R., Diagnostic criteria for use in psychiatric research, Arch. Gen. Psychiat., 26 (1972) 57-63. Ferkany, J.W., Smith, L.A., Seifert, Jr., W.E., Caprioli, R.M. and Enna, S.J., Measurement of gamma-aminobutyric acid (GABA) in blood, Life Sci., 22 (1978) 2121-2128. Gold, B.I., Bowers, M.B., Roth, R.H. and Sweeney, D.W., GABA levels in CSF of patients with psychiatric disorders, Amer. J. Psychiat., 137 (1980) 362-364. Haber, B., Kuriyama, K. and Roberts, E., An anion stimulated L-glutamic acid decarboxylase in non-neural tissues -occurrence and subcellular localization in mouse kidney and developing chick embryo brain, Biochem. Pharmacol., 19 (1970) 1119-1136. Liischer, W., GABA in plasma and cerebrospinal fluid of different species - Effects of y-acetylenic GABA, y-vinyl GABA and sodium valproate, J. Neurochem., 32 (1979) 1587-1591. McCarthy, B.W., Gomes, U.R., Neethling, A.C., Shanley, B.C., Taljaard, J.J.F., Potgieter, L. and Roux, J.T., y-Aminobutyric acid concentration in cerebrospinal fluid in schizophrenia, J. Neurochem., 36(4) (1981) 1406-1408. Manyam, N.V.B., Katz, L., Hare, T.A., Gerber, III, J.C. and Grossman, M.H., Levels of y-aminobutyric acid in cerebrospinal fluid in various neurologic disorders, Arch. Neurol. (Chic.), 37 (1980) 352-355. Perry, T.L., Hansen, S. and Kloster, M., Huntington’s chorea deficiency of y-aminobutyric acid in brain, N. Engl. J. Med., 288(7) (1973) 337-342. Perry, T.L., Buchanan, J., Kish, S.J. and Hansen, S., y-Aminobutyric-acid deficiency in brain of schizophrenic patients, Lancet, i (1979) 237-239. Schlesser, M.A., Winokur, G. and Sherman, B.M., Hypothalamic-pituitary-adrenal axis activity in depressive illness, Arch. Gen. Psychiat., 37 (1980) 737-743. Sherman, A.D. and Petty, F., Neurochemical basis of the action of antidepressants on learned helplessness, Behav. Neural Biol., 30 (1980) 119-134. Winokur, G., Behar, D., Vanvalkenburg, C. and Lowry, M., Is a familial definition of depression both feasible and valid? J. Nerv. Ment. Dis., 166(11) (1978) 764-768. Zachmann, M., Tocci, P. and Nyhan, W.L., The occurrence of y-aminobutyric acid in human tissues other than brain, J. Biol. Chem., 241 (1966) 1355-1358. Zimmer, R., Teeklen, A.W., Meier, K.D., Ackenheil, M. and Zander, K.J., Preliminary studies on CSF gamma-aminobutyric acid levels in psychiatric patients before and during the treatment with different psychotropic durgs, Progr. Neuro-Psychopharmacol., 4 (1981) 613-620.