Drug and Chemical Toxicology
ISSN: 0148-0545 (Print) 1525-6014 (Online) Journal homepage: http://www.tandfonline.com/loi/idct20
Effect of Methionine on Regional CDF-1 Mouse Brain Monoamines F. S. Messiha To cite this article: F. S. Messiha (1990) Effect of Methionine on Regional CDF-1 Mouse Brain Monoamines, Drug and Chemical Toxicology, 13:4, 355-365, DOI: 10.3109/01480549009032292 To link to this article: http://dx.doi.org/10.3109/01480549009032292
Published online: 27 Sep 2008.
Submit your article to this journal
Article views: 4
View related articles
Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalInformation?journalCode=idct20 Download by: [Australian National University]
Date: 08 November 2015, At: 17:38
DRUG AND CHEMICAL TOXICOLOGY, 1 3 ( 4 ) , 3 5 5 - 3 6 5 (1990)
EFFECT OF METHIONINE ON REGIONAL CDF-1 Downloaded by [Australian National University] at 17:38 08 November 2015
MOUSE BRAIN MONOAMINES F.S.
Messiha
Department of Pharmacology University of North Dakota School of Medicine Grand Forks, ND 5 8 2 0 3
ABSTRACT
The effect of methionine on cerebral content of dopamine, serotonin and major acidic metabolites was studied in distinct CDF-1 mouse brain regions. Methionine was administered 3 0 mg/kg € o r five trials over 2 4 h and mice were sacrificed 30 min post the terminal treatment. Methionine exerted differential effects on regional brain concentrations of 3,4-dihydroxyphenylacetic acid and serotonin. The results suggest methionine-mediated decreases of dopamine turnover in the hippocampus and medulla compared to increases of serotonin turnover in the cerebral cortex and midbrain regions. The results indicate a central action for methionine as related to endogenous neurotransmitters measured which may precede the abnormal 0- or N-dimethylation ascribed to methionine and the insuing adverse effects postulated in schizophrenia. 355 Copyright 0 1990 by Marcel Dekker, Inc.
356
MESSIHA
INTRODUCTION
(MTN), a-amino-8-methylthio-n-butyric
Methionine
acid, is an essential sulfur amino acid which is not synthesized by the human body and provides
the main
Downloaded by [Australian National University] at 17:38 08 November 2015
source of sulfur requirement for the human organism.
It
is utilized as a methyl group donor for various transmethylation
reactions,
neurotransmitter activation
to
including methylation
substances1n2, subsequent
S-adenosyl
methionine.
of
some
enzymatic
Some
of
the
methylation processes also serve in the detoxification of certain pyridine analogues such as nicotinic acid, and in the biosyntheses of specific required compounds such as cholin
or creatine.
In addition,
abnormal
methylation of the neurotransmitters dopamine (DA) and 5-hydroxytryptamine (5-HT) has been hypothesized in the biology of schizophrenia3”.
This involves the enzymatic
0- and N-dimethylation of DA and 5-HT1’2’8’9. Enhancement
of such 0- or N-methylation by MTN may alter cerebral
neuronal activity of the biogenic amines which could dispose to neurotoxicity.
The present experiment reports
on the effect of MTN on regional brain DA, 5-HT and their major acidic metabolites by the adult male CDF-1 mouse strain which possesses high brain content of DA and 5-€IT among various mouse strains”.
357
EFFECT OF METHIONINE
METHODS Adult male mice (9-11 weeks old) of the CDF-1 strain were
purchased
Wilmington, MA.
from
Charles
River
Laboratories,
They were maintained in animal research
Downloaded by [Australian National University] at 17:38 08 November 2015
facility for at least 10-days before beginning of the experiments.
Animals had access to purina pellet food
and water ad lib in a facility supplied with controlled photoperiod of 12 hrs of illumination, beginning 7 : O O a.m., and 12 hrs of darkness. Methionine was dissolved in saline and was injected intraperitoneally (i.p.), 30 mg/kg, for five trials over 24
h.
a.m.
The initial 3 treatments were given between 10 and
4
p.m.
and
the
remaining
trials
were
administered next following day at 8:30 and 1O:OO a.m. The control were injected with saline at the same time intervals. The mice were then killed by decapitation 30 min after the fifth treatment.
The individual brains
were removed, placed on ice and were dissected into cerebral cortex, striatum, hippocampus, the medulla oblongata and midbrain regions. The brain specimens were weighed and individually homogenized in 2.0 ml ice cold 0.1M HC104 solution saturated with EDTA by a glass homogenizer supplied with a motor driven teflon pestle. The homogenate was immediately centrifuged f o r 30 min at 20,000
x g at
4 ' ~ and
the supernatant was similarly
358
MESSIHA
recentrifuged. The volume of the clear supernatant fluid obtained was measured and aliquoted in small vials which were kept frozen until assayed for the following compounds by high-performance liquid chromatography (HPLC). Downloaded by [Australian National University] at 17:38 08 November 2015
These were DA, 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) .
They were isolated and quantitated by
HPLC as previously described” by a programmable Waters Wisp Model 710B with an M-45 solvent delivery system attached to an electrochemical detector, LC-4B amperometric equipment and a spectra-physics SP4290 integrator. In brief, the compounds were separated on a reverse chromatography column, lOOm x 4.6 mm d.i. (Altech Inc., Deerfield, IL) with a 20% Carbon loading, at a flowrate of lml/min with an electrochemical operating at volts.
0.8
An acetonitrile mobile phase was used for the
determination of 5-HT, 5-HIAA and HVA.
A second mobile
phase, 0.1 M monochloracetic acid, was employed for the assays of DA and DOPAC. External standards derived from authentic compounds purchased from Sigma Co., St. Louis, MO and Aldrich Co., Milwaukee, WI.
Stock standard solutions of 1 mg/ml were
prepared, aliquoted in small vials and were kept frozen. Each standard was thawed and diluted for determination of the calibration curve which
&as
performed
from
359
EFFECT OF METHIONINE
replicate samples at 3 concentrations over the required ng range.
The diluted standard series were analyzed at
beginning, after each 10 to 12 brain samples and at the end o f each independent assay. Quantitation was made by Downloaded by [Australian National University] at 17:38 08 November 2015
integration in comparison to the external standards used. The results were analyzed by two tailed Student's & test for independent means for the assessment of the statistical significance.
RESULTS Table 1 lists means t SEM concentrations of DA and DOPAC
determined
in five mouse brain
function of the MTN treatment.
regions as a
A statistically insig-
nificant reduction of cortical DA and DOPAC compared to saline control was apparent by MTN.
The MTN treatment
decreased hippocampal content of DOPAC by approximately 70%
DA
(p
ISSN: 0148-0545 (Print) 1525-6014 (Online) Journal homepage: http://www.tandfonline.com/loi/idct20
Effect of Methionine on Regional CDF-1 Mouse Brain Monoamines F. S. Messiha To cite this article: F. S. Messiha (1990) Effect of Methionine on Regional CDF-1 Mouse Brain Monoamines, Drug and Chemical Toxicology, 13:4, 355-365, DOI: 10.3109/01480549009032292 To link to this article: http://dx.doi.org/10.3109/01480549009032292
Published online: 27 Sep 2008.
Submit your article to this journal
Article views: 4
View related articles
Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalInformation?journalCode=idct20 Download by: [Australian National University]
Date: 08 November 2015, At: 17:38
DRUG AND CHEMICAL TOXICOLOGY, 1 3 ( 4 ) , 3 5 5 - 3 6 5 (1990)
EFFECT OF METHIONINE ON REGIONAL CDF-1 Downloaded by [Australian National University] at 17:38 08 November 2015
MOUSE BRAIN MONOAMINES F.S.
Messiha
Department of Pharmacology University of North Dakota School of Medicine Grand Forks, ND 5 8 2 0 3
ABSTRACT
The effect of methionine on cerebral content of dopamine, serotonin and major acidic metabolites was studied in distinct CDF-1 mouse brain regions. Methionine was administered 3 0 mg/kg € o r five trials over 2 4 h and mice were sacrificed 30 min post the terminal treatment. Methionine exerted differential effects on regional brain concentrations of 3,4-dihydroxyphenylacetic acid and serotonin. The results suggest methionine-mediated decreases of dopamine turnover in the hippocampus and medulla compared to increases of serotonin turnover in the cerebral cortex and midbrain regions. The results indicate a central action for methionine as related to endogenous neurotransmitters measured which may precede the abnormal 0- or N-dimethylation ascribed to methionine and the insuing adverse effects postulated in schizophrenia. 355 Copyright 0 1990 by Marcel Dekker, Inc.
356
MESSIHA
INTRODUCTION
(MTN), a-amino-8-methylthio-n-butyric
Methionine
acid, is an essential sulfur amino acid which is not synthesized by the human body and provides
the main
Downloaded by [Australian National University] at 17:38 08 November 2015
source of sulfur requirement for the human organism.
It
is utilized as a methyl group donor for various transmethylation
reactions,
neurotransmitter activation
to
including methylation
substances1n2, subsequent
S-adenosyl
methionine.
of
some
enzymatic
Some
of
the
methylation processes also serve in the detoxification of certain pyridine analogues such as nicotinic acid, and in the biosyntheses of specific required compounds such as cholin
or creatine.
In addition,
abnormal
methylation of the neurotransmitters dopamine (DA) and 5-hydroxytryptamine (5-HT) has been hypothesized in the biology of schizophrenia3”.
This involves the enzymatic
0- and N-dimethylation of DA and 5-HT1’2’8’9. Enhancement
of such 0- or N-methylation by MTN may alter cerebral
neuronal activity of the biogenic amines which could dispose to neurotoxicity.
The present experiment reports
on the effect of MTN on regional brain DA, 5-HT and their major acidic metabolites by the adult male CDF-1 mouse strain which possesses high brain content of DA and 5-€IT among various mouse strains”.
357
EFFECT OF METHIONINE
METHODS Adult male mice (9-11 weeks old) of the CDF-1 strain were
purchased
Wilmington, MA.
from
Charles
River
Laboratories,
They were maintained in animal research
Downloaded by [Australian National University] at 17:38 08 November 2015
facility for at least 10-days before beginning of the experiments.
Animals had access to purina pellet food
and water ad lib in a facility supplied with controlled photoperiod of 12 hrs of illumination, beginning 7 : O O a.m., and 12 hrs of darkness. Methionine was dissolved in saline and was injected intraperitoneally (i.p.), 30 mg/kg, for five trials over 24
h.
a.m.
The initial 3 treatments were given between 10 and
4
p.m.
and
the
remaining
trials
were
administered next following day at 8:30 and 1O:OO a.m. The control were injected with saline at the same time intervals. The mice were then killed by decapitation 30 min after the fifth treatment.
The individual brains
were removed, placed on ice and were dissected into cerebral cortex, striatum, hippocampus, the medulla oblongata and midbrain regions. The brain specimens were weighed and individually homogenized in 2.0 ml ice cold 0.1M HC104 solution saturated with EDTA by a glass homogenizer supplied with a motor driven teflon pestle. The homogenate was immediately centrifuged f o r 30 min at 20,000
x g at
4 ' ~ and
the supernatant was similarly
358
MESSIHA
recentrifuged. The volume of the clear supernatant fluid obtained was measured and aliquoted in small vials which were kept frozen until assayed for the following compounds by high-performance liquid chromatography (HPLC). Downloaded by [Australian National University] at 17:38 08 November 2015
These were DA, 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) .
They were isolated and quantitated by
HPLC as previously described” by a programmable Waters Wisp Model 710B with an M-45 solvent delivery system attached to an electrochemical detector, LC-4B amperometric equipment and a spectra-physics SP4290 integrator. In brief, the compounds were separated on a reverse chromatography column, lOOm x 4.6 mm d.i. (Altech Inc., Deerfield, IL) with a 20% Carbon loading, at a flowrate of lml/min with an electrochemical operating at volts.
0.8
An acetonitrile mobile phase was used for the
determination of 5-HT, 5-HIAA and HVA.
A second mobile
phase, 0.1 M monochloracetic acid, was employed for the assays of DA and DOPAC. External standards derived from authentic compounds purchased from Sigma Co., St. Louis, MO and Aldrich Co., Milwaukee, WI.
Stock standard solutions of 1 mg/ml were
prepared, aliquoted in small vials and were kept frozen. Each standard was thawed and diluted for determination of the calibration curve which
&as
performed
from
359
EFFECT OF METHIONINE
replicate samples at 3 concentrations over the required ng range.
The diluted standard series were analyzed at
beginning, after each 10 to 12 brain samples and at the end o f each independent assay. Quantitation was made by Downloaded by [Australian National University] at 17:38 08 November 2015
integration in comparison to the external standards used. The results were analyzed by two tailed Student's & test for independent means for the assessment of the statistical significance.
RESULTS Table 1 lists means t SEM concentrations of DA and DOPAC
determined
in five mouse brain
function of the MTN treatment.
regions as a
A statistically insig-
nificant reduction of cortical DA and DOPAC compared to saline control was apparent by MTN.
The MTN treatment
decreased hippocampal content of DOPAC by approximately 70%
DA
(p
